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Gene expression can be influenced by genetic variants that are closely linked to the expressed gene (cis eQTLs) and variants in other parts of the genome (trans eQTLs). We created a multiparental mapping population by sampling genotypes from a single natural population of Mimulus guttatus and scored gene expression in the leaves of 1,588 plants. We find that nearly every measured gene exhibits cis regulatory variation (91% have FDR < 0.05). cis eQTLs are usually allelic series with three or more functionally distinct alleles. The cis locus explains about two thirds of the standing genetic variance (on average) but varies among genes and tends to be greatest when there is high indel variation in the upstream regulatory region and high nucleotide diversity in the coding sequence. Despite mapping over 10,000 trans eQTL / affected gene pairs, most of the genetic variance generated by trans acting loci remains unexplained. This implies a large reservoir of trans acting genes with subtle or diffuse effects. Mapped trans eQTLs show lower allelic diversity but much higher genetic dominance than cis eQTLs. Several analyses also indicate that trans eQTLs make a substantial contribution to the genetic correlations in expression among different genes. They may thus be essential determinants of "gene expression modules," which has important implications for the evolution of gene expression and how it is studied by geneticists. Author summary: Mimulus guttatus (yellow monkeyflower) is a model for the study of quantitative trait evolution in natural populations. Research has focused mainly on whole organism traits like flower size or herbivore resistance, but the level of expression of a gene is also a quantitative trait. In this study, we dissect leaf transcriptome variation using a breeding design that estimates the contribution of individual loci to expression variation (eQTLs). We find rough agreement to the "oligogenic model" of inheritance where a major locus (the cis regulatory region) generates much of the genetic variation in the population. Associations studies usually characterize genetic effects as binary (e.g. the two alternatives at a single nucleotide polymorphism or "SNP"), but this description is insufficient for Mimulus. Most loci exhibit multiple, and in some cases, a continuum of alleles. We find that trans eQTLs have different features than cis eQTLs, both in terms of the diversity and genetic dominance of alleles. These genetic features of eQTLs are critical determinants of the "G matrix," the genetic variances and covariances among all genes which determine how gene expression will evolve under selection in response to changing environmental conditions. Our finding of large effect sizes and high allelic diversity suggests that the G matrix may be surprisingly malleable, even on ecological timescales. [ABSTRACT FROM AUTHOR]

The laboratory mouse has served as the premier animal model system for both basic and preclinical investigations for over a century. However, laboratory mice capture only a subset of the genetic variation found in wild mouse populations, ultimately limiting the potential of classical inbred strains to uncover phenotype-associated variants and pathways. Wild mouse populations are reservoirs of genetic diversity that could facilitate the discovery of new functional and disease-associated alleles, but the scarcity of commercially available, well-characterized wild mouse strains limits their broader adoption in biomedical research. To overcome this barrier, we have recently developed, sequenced, and phenotyped a set of 11 inbred strains derived from wild-caught Mus musculus domesticus. Each of these "Nachman strains" immortalizes a unique wild haplotype sampled from one of five environmentally distinct locations across North and South America. Whole genome sequence analysis reveals that each strain carries between 4.73–6.54 million single nucleotide differences relative to the GRCm39 mouse reference, with 42.5% of variants in the Nachman strain genomes absent from current classical inbred mouse strain panels. We phenotyped the Nachman strains on a customized pipeline to assess the scope of disease-relevant neurobehavioral, biochemical, physiological, metabolic, and morphological trait variation. The Nachman strains exhibit significant inter-strain variation in >90% of 1119 surveyed traits and expand the range of phenotypic diversity captured in classical inbred strain panels. These novel wild-derived inbred mouse strain resources are set to empower new discoveries in both basic and preclinical research. Author summary: Inbred laboratory mouse strains are integral tools for both preclinical and basic research. However, laboratory mice capture a small fraction of the genetic diversity found in wild mouse populations, a consideration that necessarily constrains the scope of possible discovery in studies restricted to lab mice. Further, laboratory mice were developed through programs of intense artificial selection for increased breeding performance, docility, and other traits of interest. Consequently, the genetic control of many complex traits in lab mice may not accurately reflect the mechanisms of their regulation in nature. To overcome these limitations, we developed a new inbred mouse strain resource founded from wild-caught mice subject to minimal laboratory selection. We show that strains in this "Nachman panel" harbor millions of genetic variants absent from current laboratory mouse models, including predicted deleterious alleles and gene-spanning structural variants. Paralleling this genetic diversity, we show that Nachman strains capture striking phenotypic variation across a multitude of disease-relevant biochemical, neurobehavioral, physiological, morphological, and metabolic traits, expanding the range of trait variation recovered in lab strains alone. Overall, our strain survey emphasizes the collective potential of the Nachman strains to advance discoveries into multiple disease areas and basic biology. [ABSTRACT FROM AUTHOR]

Symbiomonas scintillans Guillou et Chrétiennot-Dinet, 1999 is a tiny (1.4 μm) heterotrophic microbial eukaryote. The genus was named based on the presence of endosymbiotic bacteria in its endoplasmic reticulum, however, like most such endosymbionts neither the identity nor functional association with its host were known. We generated both amplification-free shotgun metagenomics and whole genome amplification sequencing data from S. scintillans strains RCC257 and RCC24, but were unable to detect any sequences from known lineages of endosymbiotic bacteria. The absence of endobacteria was further verified with FISH analyses. Instead, numerous contigs in assemblies from both RCC24 and RCC257 were closely related to prasinoviruses infecting the green algae Ostreococcus lucimarinus, Bathycoccus prasinos, and Micromonas pusilla (OlV, BpV, and MpV, respectively). Using the BpV genome as a reference, we assembled a near-complete 190 kbp draft genome encoding all hallmark prasinovirus genes, as well as two additional incomplete assemblies of closely related but distinct viruses from RCC247, and three similar draft viral genomes from RCC24, which we collectively call SsVs. A multi-gene tree showed the three SsV genome types branched within highly supported clades with each of BpV2, OlVs, and MpVs, respectively. Interestingly, transmission electron microscopy also revealed a 190 nm virus-like particle similar the morphology and size of the endosymbiont originally reported in S. scintillans. Overall, we conclude that S. scintillans currently does not harbour an endosymbiotic bacterium, but is associated with giant viruses. Author summary: Endosymbiotic bacteria are found in a wide variety of hosts across the tree of eukaryotes and have been proposed to be evolutionarily and ecologically significant, but in most cases, we know little to nothing about them. This is exemplified by the stramenopile flagellate Symbiomonas scintillans, where the bacterial endosymbiont that gave the genus its name remains unidentified and has no known function. Here we used multiple genomic sequencing methods on two strains of S. scintillans, and showed absence of the endobacteria belonging to common endosymbiotic lineages. Instead, we identified giant viruses similar to those infecting prasinophyte green algae. Although further experiments are needed to verify the nature of the viral association with S. scintillans, our study is reminiscent of how the first mimivirus (named for mimicking gram-negative bacteria) was discovered, and we speculate similar discoveries will follow with ever-increasing genomic data of protists. [ABSTRACT FROM AUTHOR]

Many disease-susceptible SNPs exhibit significant disparity in ancestral and derived allele frequencies across worldwide populations. While previous studies have examined population differentiation of alleles at specific SNPs, global ethnic patterns of ensembles of disease risk alleles across human diseases are unexamined. To examine these patterns, we manually curated ethnic disease association data from 5,065 papers on human genetic studies representing 1,495 diseases, recording the precise risk alleles and their measured population frequencies and estimated effect sizes. We systematically compared the population frequencies of cross-ethnic risk alleles for each disease across 1,397 individuals from 11 HapMap populations, 1,064 individuals from 53 HGDP populations, and 49 individuals with whole-genome sequences from 10 populations. Type 2 diabetes (T2D) demonstrated extreme directional differentiation of risk allele frequencies across human populations, compared with null distributions of European-frequency matched control genomic alleles and risk alleles for other diseases. Most T2D risk alleles share a consistent pattern of decreasing frequencies along human migration into East Asia. Furthermore, we show that these patterns contribute to disparities in predicted genetic risk across 1,397 HapMap individuals, T2D genetic risk being consistently higher for individuals in the African populations and lower in the Asian populations, irrespective of the ethnicity considered in the initial discovery of risk alleles. We observed a similar pattern in the distribution of T2D Genetic Risk Scores, which are associated with an increased risk of developing diabetes in the Diabetes Prevention Program cohort, for the same individuals. This disparity may be attributable to the promotion of energy storage and usage appropriate to environments and inconsistent energy intake. Our results indicate that the differential frequencies of T2D risk alleles may contribute to the observed disparity in T2D incidence rates across ethnic populations. [ABSTRACT FROM AUTHOR]

An interview with researcher Rebecca Cann is presented. When asked about biochemistry, she explains the elitism in the courses of anthropology and biochemistry. Cann thinks that only few high school girls made it through Catholic schools unscathed. Cann reveals that she had imagined getting mitochondrial DNA from the archaic people.

Cartilage-hair hypoplasia (CHH) is a pleiotropic disease caused by recessive mutations in the RMRP gene that result in a wide spectrum of manifestations including short stature, sparse hair, metaphyseal dysplasia, anemia, immune deficiency, and increased incidence of cancer. Molecular diagnosis of CHH has implications for management, prognosis, follow-up, and genetic counseling of affected patients and their families. We report 20 novel mutations in 36 patients with CHH and describe the associated phenotypic spectrum. Given the high mutational heterogeneity (62 mutations reported to date), the high frequency of variations in the region (eight single nucleotide polymorphisms in and around RMRP), and the fact that RMRP is not translated into protein, prediction of mutation pathogenicity is difficult. We addressed this issue by a comparative genomic approach and aligned the genomic sequences of RMRP gene in the entire class of mammals. We found that putative pathogenic mutations are located in highly conserved nucleotides, whereas polymorphisms are located in non-conserved positions. We conclude that the abundance of variations in this small gene is remarkable and at odds with its high conservation through species; it is unclear whether these variations are caused by a high local mutation rate, a failure of repair mechanisms, or a relaxed selective pressure. The marked diversity of mutations in RMRP and the low homozygosity rate in our patient population indicate that CHH is more common than previously estimated, but may go unrecognized because of its variable clinical presentation. Thus, RMRP molecular testing may be indicated in individuals with isolated metaphyseal dysplasia, anemia, or immune dysregulation. [ABSTRACT FROM AUTHOR]

Interviews Nicholas Wade, science journalist of the "New York Time." Description of the beats; Views on the readership of the newspaper; Length of service with the publication.

Cell intercalation is a highly directed cell rearrangement that is essential for animal morphogenesis. As such, intercalation requires orchestration of cell polarity across the plane of the tissue. CDC-42 is a Rho family GTPase with key functions in cell polarity, yet its role during epithelial intercalation has not been established because its roles early in embryogenesis have historically made it difficult to study. To circumvent these early requirements, in this paper we use tissue-specific and conditional loss-of-function approaches to identify a role for CDC-42 during intercalation of the Caenorhabditis elegans dorsal embryonic epidermis. CDC-42 activity is enriched in the medial tips of intercalating cells, which extend as cells migrate past one another. Moreover, CDC-42 is involved in both the efficient formation and orientation of cell tips during cell rearrangement. Using conditional loss-of-function we also show that the PAR complex functions in tip formation and orientation. Additionally, we find that the sole C. elegans Eph receptor, VAB-1, functions during this process in an Ephrin-independent manner. Using epistasis analysis, we find that vab-1 lies in the same genetic pathway as cdc-42 and is responsible for polarizing CDC-42 activity to the medial tip. Together, these data establish a previously uncharacterized role for polarized CDC-42, in conjunction with PAR-6, PAR-3 and an Eph receptor, during epithelial intercalation. [ABSTRACT FROM AUTHOR]

During the last few centuries D. melanogaster populations were invaded by several transposable elements, the most recent of which was thought to be the P-element between 1950 and 1980. Here we describe a novel TE, which we named Spoink, that has invaded D. melanogaster. It is a 5216nt LTR retrotransposon of the Ty3/gypsy superfamily. Relying on strains sampled at different times during the last century we show that Spoink invaded worldwide D. melanogaster populations after the P-element between 1983 and 1993. This invasion was likely triggered by a horizontal transfer from the D. willistoni group, much as the P-element. Spoink is probably silenced by the piRNA pathway in natural populations and about 1/3 of the examined strains have an insertion into a canonical piRNA cluster such as 42AB. Given the degree of genetic investigation of D. melanogaster it is perhaps surprising that Spoink was able to invade unnoticed. Author summary: Horizontal transfer of transposable elements (TE) is a major factor driving genome evolution. Yet well documented cases of such horizontal transfer events are rare. Most evidence is indirect, relying on sequence similarity of TEs between species. Based on strains sampled during the last decades we provide direct evidence that the retrotransposon Spoink was absent in worldwide D. melanogaster populations before 1983 but present in populations after 1993. We suggest that the Spoink invasion was triggered by a horizontal transfer from a Drosophila species of the willistoni group. [ABSTRACT FROM AUTHOR]

Cultural exchange of fermentation techniques has driven the spread of Saccharomyces cerevisiae across the globe, establishing natural populations in many countries. Despite this, Oceania is thought to lack native populations of S. cerevisiae, only being introduced after colonisation. Here we investigate the genomic landscape of 411 S. cerevisiae isolated from spontaneous grape fermentations in Australia across multiple locations, years, and grape cultivars. Spontaneous fermentations contained highly recombined mosaic strains that exhibited high levels of genome instability. Assigning genomic windows to putative ancestral origin revealed that few closely related starter lineages have come to dominate the genetic landscape, contributing most of the genetic variation. Fine-scale phylogenetic analysis of loci not observed in strains of commercial wine origin identified widespread admixture with European derived beer yeast along with three independent admixture events from potentially endemic Oceanic lineages that was associated with genome instability. Finally, we investigated Australian ecological niches for basal isolates, identifying phylogenetically distinct S. cerevisiae of non-European, non-domesticated origin associated with admixture loci. Our results illustrate the effect commercial use of microbes may have on local microorganism genetic diversity and demonstrates the presence of non-domesticated, potentially endemic lineages of S. cerevisiae in Australian niches that are actively admixing. Author summary: Saccharomyces cerevisiae yeast are utilized throughout the world to carry out fermentation and have been transferred between cultures for thousands of years. However, Australia is thought to lack natural populations of this yeast. In this study we investigated the genomic landscape of S. cerevisiae isolated from spontaneous wine fermentations in Australia, identifying widespread admixture and genome instability. Local phylogenetics identified clear signals for admixture between basal non-wine isolates, some of which were found to be associated with previously unidentified yeast from Australian natural environments. This discovery suggests that natural non-domesticated lineages of S. cerevisiae exist in the Australian environment that may predate colonisation. Finally, we investigated genetic contribution against a publicly available set of commercial starter and wine strains isolated in Australia over the past 70 years. Revealing that few related commercial starter lineages contribute most of the genetics within the standing variation of the spontaneously fermenting yeast analysed in this study. [ABSTRACT FROM AUTHOR]

Tissues are subject to multiple mechanical inputs at the cellular level that influence their overall shape and function. In the small intestine, actomyosin contractility can be induced by many physiological and pathological inputs. However, we have little understanding of how contractility impacts the intestinal epithelium on a cellular and tissue level. In this study, we probed the cell and tissue-level effects of contractility by using mouse models to genetically increase the level of myosin activity in the two distinct morphologic compartments of the intestinal epithelium, the crypts and villi. We found that increased contractility in the villar compartment caused shape changes in the cells that expressed the transgene and their immediate neighbors. While there were no discernable effects on villar architecture or cell polarity, even low levels of transgene induction in the villi caused non-cell autonomous hyperproliferation of the transit amplifying cells in the crypt, driving increased cell flux through the crypt-villar axis. In contrast, induction of increased contractility in the proliferating cells of the crypts resulted in nuclear deformations, DNA damage, and apoptosis. This study reveals the complex and diverse responses of different intestinal epithelial cells to contractility and provides important insight into mechanical regulation of intestinal physiology. Author summary: The small intestine epithelium is comprised of two main compartments: the villi which contain differentiated cells that function in nutrient absorption, and the crypts which are made up of undifferentiated cells which serve to replenish cells of the villi. Because of their physical location within the tissue, villi and crypts are subjected to different types of insults and mechanical forces. We sought to directly test how villi and crypts respond to mechanical changes in the epithelia by genetically inducing actomyosin contraction. Increasing contractility in villar cells resulted in cell shape changes without affecting their overall polarity or organization. However, it led to a non-autonomous increase in proliferation of the undifferentiated cells of the intestine. In contrast, increased contractility in the proliferative cells of the crypt resulted in nuclear shape changes, DNA damage and ultimately rapid cell death. Thus, our work demonstrates that the crypt and villi epithelia respond differently to mechanical changes and highlights long-range regulation between villi and crypt compartments. [ABSTRACT FROM AUTHOR]

The HostSeq initiative recruited 10,059 Canadians infected with SARS-CoV-2 between March 2020 and March 2023, obtained clinical information on their disease experience and whole genome sequenced (WGS) their DNA. We analyzed the WGS data for genetic contributors to severe COVID-19 (considering 3,499 hospitalized cases and 4,975 non-hospitalized after quality control). We investigated the evidence for replication of loci reported by the International Host Genetics Initiative (HGI); analyzed the X chromosome; conducted rare variant gene-based analysis and polygenic risk score testing. Population stratification was adjusted for using meta-analysis across ancestry groups. We replicated two loci identified by the HGI for COVID-19 severity: the LZTFL1/SLC6A20 locus on chromosome 3 and the FOXP4 locus on chromosome 6 (the latter with a variant significant at P < 5E-8). We found novel significant associations with MRAS and WDR89 in gene-based analyses, and constructed a polygenic risk score that explained 1.01% of the variance in severe COVID-19. This study provides independent evidence confirming the robustness of previously identified COVID-19 severity loci by the HGI and identifies novel genes for further investigation. Author summary: Host genetics determine how human genetics contribute to the response to infectious disease. HostSeq is a Canada-wide effort to contribute to our understanding of host genetics for COVID-19. The HostSeq study involves genetic and clinical data from individuals who tested positive for SARS-CoV-2 across Canada. This work examines locations in the human genome that have been reported to be involved in severe COVID-19 worldwide and determines if we also see the involvement of those genomic locations in Canadians. This work also explores several 'genetic quantities' such as a determination of how much of COVID-19 severity is due to genetics in Canada. [ABSTRACT FROM AUTHOR]

Fibrolamellar carcinoma (FLC) is a rare liver cancer that disproportionately affects adolescents and young adults. Currently, no standard of care is available and there remains a dire need for new therapeutics. Most patients harbor the fusion oncogene DNAJB1-PRKACA (DP fusion), but clinical inhibitors are not yet developed and it is critical to identify downstream mediators of FLC pathogenesis. Here, we identify long noncoding RNA LINC00473 among the most highly upregulated genes in FLC tumors and determine that it is strongly suppressed by RNAi-mediated inhibition of the DP fusion in FLC tumor epithelial cells. We show by loss- and gain-of-function studies that LINC00473 suppresses apoptosis, increases the expression of FLC marker genes, and promotes FLC growth in cell-based and in vivo disease models. Mechanistically, LINC00473 plays an important role in promoting glycolysis and altering mitochondrial activity. Specifically, LINC00473 knockdown leads to increased spare respiratory capacity, which indicates mitochondrial fitness. Overall, we propose that LINC00473 could be a viable target for this devastating disease. Schematic was created with BioRender.com. Author summary: Fibrolamellar carcinoma (FLC) is a lethal liver cancer that is characterized by the DNAJB1-PRKACA (DP fusion) oncogene. This disease is particularly challenging to treat as it lacks non-invasive diagnostic biomarkers, known risk factors, and effective therapeutic options. Patients with FLC have a median age of 22 years and the prognosis is poor. In this study, we examined FLC tumors from independent patient cohorts and multiple disease models and found consistently elevated levels of a primate-specific long noncoding RNA named LINC00473. By leveraging single-cell analysis on patient tumors, we found that LINC00473 is enriched in tumor epithelial cells. Using multiple FLC models, we show that the DP fusion drives LINC00473 expression and that LINC00473 promotes FLC growth by mitigating cell death. We also show that LINC00473 modulates FLC energetics by increasing glycolysis and altering mitochondrial fitness. Together, this study unveils an important mechanism downstream of the signature DP fusion oncogene in FLC pathogenesis. [ABSTRACT FROM AUTHOR]

Across the human genome, there are large-scale fluctuations in genetic diversity caused by the indirect effects of selection. This "linked selection signal" reflects the impact of selection according to the physical placement of functional regions and recombination rates along chromosomes. Previous work has shown that purifying selection acting against the steady influx of new deleterious mutations at functional portions of the genome shapes patterns of genomic variation. To date, statistical efforts to estimate purifying selection parameters from linked selection models have relied on classic Background Selection theory, which is only applicable when new mutations are so deleterious that they cannot fix in the population. Here, we develop a statistical method based on a quantitative genetics view of linked selection, that models how polygenic additive fitness variance distributed along the genome increases the rate of stochastic allele frequency change. By jointly predicting the equilibrium fitness variance and substitution rate due to both strong and weakly deleterious mutations, we estimate the distribution of fitness effects (DFE) and mutation rate across three geographically distinct human samples. While our model can accommodate weaker selection, we find evidence of strong selection operating similarly across all human samples. Although our quantitative genetic model of linked selection fits better than previous models, substitution rates of the most constrained sites disagree with observed divergence levels. We find that a model incorporating selective interference better predicts observed divergence in conserved regions, but overall our results suggest uncertainty remains about the processes generating fitness variation in humans. Author summary: Across the human genome, there are large-scale fluctuations in genetic diversity caused by the indirect effects of selection. This "linked selection signal" reflects the impact of selection according to the physical placement of functional regions and recombination rates along chromosomes. Previous work has shown that purifying selection acting against the steady influx of new deleterious mutations at functional portions of the genome shapes patterns of genomic variation. To date, statistical efforts to estimate purifying selection parameters from linked selection models have relied on classic Background Selection theory, which is only applicable when new mutations are so deleterious that they cannot fix in the population. Here, we develop a statistical method based on a quantitative genetics view of linked selection, that models how polygenic additive fitness variance distributed along the genome increases the rate of stochastic allele frequency change. By jointly predicting the equilibrium fitness variance and substitution rate due to both strong and weakly deleterious mutations, we estimate the distribution of fitness effects (DFE) and mutation rate across three geographically distinct human samples. While our model can accommodate weaker selection, we find evidence of strong selection operating similarly across all human samples. Although our quantitative genetic model of linked selection fits better than previous models, substitution rates of the most constrained sites disagree with observed divergence levels. We find that a model incorporating selective interference better predicts observed divergence in conserved regions, but overall our results suggest uncertainty remains about the processes generating fitness variation in humans. [ABSTRACT FROM AUTHOR]

Permanent heterozygous loci, such as sex- or mating-compatibility regions, often display suppression of recombination and signals of genomic degeneration. In Basidiomycota, two distinct loci confer mating compatibility. These loci encode homeodomain (HD) transcription factors and pheromone receptor (Pra)-ligand allele pairs. To date, an analysis of genome level mating-type (MAT) loci is lacking for obligate biotrophic basidiomycetes in the Pucciniales, an order containing serious agricultural plant pathogens. Here, we focus on four species of Puccinia that infect oat and wheat, including P. coronata f. sp. avenae, P. graminis f. sp. tritici, P. triticina and P. striiformis f. sp. tritici. MAT loci are located on two separate chromosomes supporting previous hypotheses of a tetrapolar mating compatibility system in the Pucciniales. The HD genes are multiallelic in all four species while the PR locus appears biallelic, except for P. graminis f. sp. tritici, which potentially has multiple alleles. HD loci are largely conserved in their macrosynteny, both within and between species, without strong signals of recombination suppression. Regions proximal to the PR locus, however, displayed signs of recombination suppression and genomic degeneration in the three species with a biallelic PR locus. Our observations support a link between recombination suppression, genomic degeneration, and allele diversity of MAT loci that is consistent with recent mathematical modelling and simulations. Finally, we confirm that MAT genes are expressed during the asexual infection cycle, and we propose that this may support regulating nuclear maintenance and pairing during infection and spore formation. Our study provides insights into the evolution of MAT loci of key pathogenic Puccinia species. Understanding mating compatibility can help predict possible combinations of nuclear pairs, generated by sexual reproduction or somatic recombination, and the potential evolution of new virulent isolates of these important plant pathogens. Author summary: Sexes in animals and some plants are determined by sex chromosomes. In fungi, mate compatibility is determined by mating-type (MAT) loci, which share some features with sex chromosomes including recombination suppression around heterozygous loci. Here, we study the MAT loci in fungal pathogens from the order Pucciniales, which cause rust diseases on many economically important plants, including wheat and oats. We show that one of the MAT loci is multiallelic, while the other is biallelic in most cases. The biallelic locus shows strong signs of recombination suppression and genetic deterioration with an increase in number of transposable elements and gene deserts surrounding the locus. Our findings on the genome biology of MAT loci in four economically important pathogens will improve predictions on potential novel virulent isolates that can lead to large scale pandemics in agriculture. [ABSTRACT FROM AUTHOR]

RNA sequencing (RNA-Seq) is widely used to capture transcriptome dynamics across tissues, biological entities, and conditions. Currently, few or no methods can handle multiple biological variables (e.g., tissues/ phenotypes) and their interactions simultaneously, while also achieving dimension reduction (DR). We propose INSIDER, a general and flexible statistical framework based on matrix factorization, which is freely available at https://github.com/kai0511/insider. INSIDER decomposes variation from different biological variables and their interactions into a shared low-rank latent space. Particularly, it introduces the elastic net penalty to induce sparsity while considering the grouping effects of genes. It can achieve DR of high-dimensional data (of > = 3 dimensions), as opposed to conventional methods (e.g., PCA/NMF) which generally only handle 2D data (e.g., sample × expression). Besides, it enables computing 'adjusted' expression profiles for specific biological variables while controlling variation from other variables. INSIDER is computationally efficient and accommodates missing data. INSIDER also performed similarly or outperformed a close competing method, SDA, as shown in simulations and can handle complex missing data in RNA-Seq data. Moreover, unlike SDA, it can be used when the data cannot be structured into a tensor. Lastly, we demonstrate its usefulness via real data analysis, including clustering donors for disease subtyping, revealing neuro-development trajectory using the BrainSpan data, and uncovering biological processes contributing to variables of interest (e.g., disease status and tissue) and their interactions. Author summary: The RNA-Seq studies aim to understand the contributing role of gene activities to phenotypes, especially human disorders, reveal important biological processes (BPs) that contribute to phenotypes, and understand the involvement of tissues (e.g., brain structures/regions) in disease development. However, our ability to understand these key issues in high-dimensional settings is hampered by the limitations of currently available methods for analyzing RNA-seq data. To address these issues, we propose a novel statistical method to jointly model variation from multiple biological variables (e.g., donor, tissue, phenotype), to introduce interactions (tissue × phenotype), and to reduce dimensionality. The favorable properties of our method are discussed below. Its advantages over one closest existing computational tool and its wide applicability are shown via simulation and applications. A computationally efficient implementation of our method is provided. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are at the interface of interactions between hosts and microbes and are therefore expected to be rapidly evolving in a coevolutionary arms race with pathogens. In contrast, previous work demonstrated that insect AMPs tend to evolve more slowly than the genome average. Metchikowin (Mtk) is a Drosophila AMP that has a single amino acid residue that segregates as either proline (P) or arginine (R) in populations of four different species, some of which diverged more than 10 million years ago. These results suggest that there is a distinct functional importance to each allele. The most likely hypotheses are driven by two main questions: does each allele have a different efficacy against different specific pathogens (specificity hypothesis)? Or, is one allele a more potent antimicrobial, but with a host fitness cost (autoimmune hypothesis)? To assess their functional differences, we created D. melanogaster lines with the P allele, R allele, or Mtk null mutation using CRISPR/Cas9 genome editing and performed a series of life history and infection assays to assess them. In males, testing of systemic immune responses to a repertoire of bacteria and fungi demonstrated that the R allele performs as well or better than the P and null alleles with most infections. Females show some results that contrast with males, with Mtk alleles either not contributing to survival or with the P allele outperforming the R allele. In addition, measurements of life history traits demonstrate that the R allele is more costly in the absence of infection for both sexes. These results are consistent with both the specificity hypothesis (either allele can perform better against certain pathogens depending on context), and the autoimmune hypothesis (the R allele is generally the more potent antimicrobial in males, and carries a fitness cost). These results provide strong in vivo evidence that differential fitness with or without infection and sex-based functional differences in alleles may be adaptive mechanisms of maintaining immune gene polymorphisms in contrast with expectations of rapid evolution. Therefore, a complex interplay of forces including pathogen species and host sex may lead to balancing selection for immune genotypes. Strikingly, this selection may act on even a single amino acid polymorphism in an AMP. Author summary: Antimicrobial peptides (AMPs) are small peptides produced by many living organisms as a frontline immune response to a variety of microbial pathogens. These peptides directly interface with pathogens to kill them shortly after infection. Importantly, multiple AMP alleles can be balanced within a population, but little is known about how this immune variation among individuals is maintained over time. Multiple species of Drosophila balance two specific alleles of the AMP, Metchnikowin (Mtk), a canonically antifungal and antibacterial peptide. Here, various life-history and infection assays assess the mechanism of Metchnikowin balancing selection using CRISPR-generated D. melanogaster with a null mutation or either of the two alleles, MtkR or MtkP. Findings demonstrate that the MtkR allele usually confers greater protection than the MtkP allele in males, but carries a fitness cost without infection. However, this pattern flips in females with certain infections, with the MtkP allele conferring greater protection than MtkR. This suggests that the two alleles are repeatedly maintained in fly populations due to differential fitness of hosts in the context of many variables including host genotype, sex, infection status, and pathogen species. It also suggests that maintenance of immune variation in a population can be based on just a single amino acid difference between individuals. [ABSTRACT FROM AUTHOR]

RpoS is an alternative sigma factor needed for the induction of the general stress response in many gammaproteobacteria. Tight regulation of RpoS levels and activity is required for bacterial growth and survival under stress. In Escherichia coli, various stresses lead to higher levels of RpoS due to increased translation and decreased degradation. During non-stress conditions, RpoS is unstable, because the adaptor protein RssB delivers RpoS to the ClpXP protease. RpoS degradation is prevented during stress by the sequestration of RssB by anti-adaptors, each of which is induced in response to specific stresses. Here, we examined how the stabilization of RpoS is reversed during recovery of the cell from stress. We found that RpoS degradation quickly resumes after recovery from phosphate starvation, carbon starvation, and when transitioning from stationary phase back to exponential phase. This process is in part mediated by the anti-adaptor IraP, known to promote RpoS stabilization during phosphate starvation via the sequestration of adaptor RssB. The rapid recovery from phosphate starvation is dependent upon a feedback loop in which RpoS transcription of rssB, encoding the adaptor protein, plays a critical role. Crl, an activator of RpoS that specifically binds to and stabilizes the complex between the RNA polymerase and RpoS, is also required for the feedback loop to function efficiently, highlighting a critical role for Crl in restoring RpoS basal levels. Author summary: In their native environments, bacteria are exposed to constant changes in nutrient availability, as well as other biotic and abiotic stressors. To adjust to these changes, bacteria must rewire gene expression to adapt to or avoid stress-induced damage. A key player in the global response to general stresses is the alternative sigma factor RpoS, a promoter specificity -determining subunit of RNA polymerase. RpoS levels increase with stress, due to increased translation and stabilization of the otherwise unstable RpoS protein. Here, we examine how the cell restores homeostasis after the stress has passed. We show that a negative feedback loop in which RpoS regulates the transcription of an adaptor for proteolysis poises the cell to rapidly resume RpoS degradation upon the exit from stress. [ABSTRACT FROM AUTHOR]

Hematophagous mosquitoes require vertebrate blood for their reproductive cycles, making them effective vectors for transmitting dangerous human diseases. Thus, high-intensity metabolism is needed to support reproductive events of female mosquitoes. However, the regulatory mechanism linking metabolism and reproduction in mosquitoes remains largely unclear. In this study, we found that the expression of estrogen-related receptor (ERR), a nuclear receptor, is activated by the direct binding of 20-hydroxyecdysone (20E) and ecdysone receptor (EcR) to the ecdysone response element (EcRE) in the ERR promoter region during the gonadotropic cycle of Aedes aegypti (named AaERR). RNA interference (RNAi) of AaERR in female mosquitoes led to delayed development of ovaries. mRNA abundance of genes encoding key enzymes involved in carbohydrate metabolism (CM)—glucose-6-phosphate isomerase (GPI) and pyruvate kinase (PYK)—was significantly decreased in AaERR knockdown mosquitoes, while the levels of metabolites, such as glycogen, glucose, and trehalose, were elevated. The expression of fatty acid synthase (FAS) was notably downregulated, and lipid accumulation was reduced in response to AaERR depletion. Dual luciferase reporter assays and electrophoretic mobility shift assays (EMSA) determined that AaERR directly activated the expression of metabolic genes, such as GPI, PYK, and FAS, by binding to the corresponding AaERR-responsive motif in the promoter region of these genes. Our results have revealed an important role of AaERR in the regulation of metabolism during mosquito reproduction and offer a novel target for mosquito control. Author summary: Following a blood meal in female mosquitoes, carbohydrate and lipid metabolism become activated, aligned with the high demands of reproductive requirements. Concurrently, the estrogen-related receptor (AaERR) mRNA level in Aedes aegypti rises dramatically. We found that AaERR is regulated by 20-hydroxyecdysone (20E), which is the main regulatory hormone during the post blood meal (PBM) reproductive phase. The 20E receptor, ecdysone receptor (EcR), directly binds to the AaERR gene promoter region, activating transcription of this gene. 20E promotes glycolysis and glycogen catabolism, accelerating carbohydrate utilization. AaERR stimulates fatty acid synthesis and leads to lipid accumulation. We confirmed the direct regulatory relationship between AaERR and metabolism-related enzymes. Further investigation revealed that AaERR depletion inhibited the expression of vitellogenin (Vg) in female mosquitoes and disrupted the development of the ovaries, affirming the critical role of AaERR in 20E-dependent metabolism. These findings highlight the essential part AaERR plays in maintaining metabolic homeostasis and promoting normal reproductive development in female mosquitoes. [ABSTRACT FROM AUTHOR]

Long terminal repeat retrotransposons (LTR-RTs) are powerful mutagens regarded as a major source of genetic novelty and important drivers of evolution. Yet, the uncontrolled and potentially selfish proliferation of LTR-RTs can lead to deleterious mutations and genome instability, with large fitness costs for their host. While population genomics data suggest that an ongoing LTR-RT mobility is common in many species, the understanding of their dual role in evolution is limited. Here, we harness the genetic diversity of 320 sequenced natural accessions of the Mediterranean grass Brachypodium distachyon to characterize how genetic and environmental factors influence plant LTR-RT dynamics in the wild. When combining a coverage-based approach to estimate global LTR-RT copy number variations with mobilome-sequencing of nine accessions exposed to eight different stresses, we find little evidence for a major role of environmental factors in LTR-RT accumulations in B. distachyon natural accessions. Instead, we show that loss of RNA polymerase IV (Pol IV), which mediates RNA-directed DNA methylation in plants, results in high transcriptional and transpositional activities of RLC_BdisC024 (HOPPLA) LTR-RT family elements, and that these effects are not stress-specific. This work supports findings indicating an ongoing mobility in B. distachyon and reveals that host RNA-directed DNA methylation rather than environmental factors controls their mobility in this wild grass model. Author summary: Long terminal repeat retrotransposons (LTR-RTs) are major components of plant genomes. Their 'copy-and-paste' replication mechanism allows them to rapidly increase in copy number, with potentially negative effects on host fitness. On the other hand, because they can rewire transcriptional networks and alter phenotypes, their mobility is an important driver of evolution. Ever since their discovery, LTR-RT activity has been linked to stress exposure, suggesting that LTR-RTs modulate the pace of evolution in response to the environment. In this study, we test this hypothesis by harnessing the genetic variation in a set of 320 natural accessions of the Mediterranean grass Brachypodium distachyon originating from diverse habitats. We find little evidence for the importance of stresses in activating B. distachyon LTR-RTs. Instead, we show that the loss of RNA polymerase IV, a component of plant retrotransposon silencing, leads to the activation and transposition of an LTR-RT family that we name HOPPLA. HOPPLA is the first LTR-RT family in B. distachyon shown to transpose in real-time. These findings open up new avenues for studying retrotransposon-mediated evolution in this close relative of staple crops, such as rice and wheat. [ABSTRACT FROM AUTHOR]

Succinate is a potent immune signalling molecule that is present in the mammalian gut and within macrophages. Both of these infection niches are colonised by the pathogenic bacterium Salmonella enterica serovar Typhimurium during infection. Succinate is a C4-dicarboyxlate that can serve as a source of carbon for bacteria. When succinate is provided as the sole carbon source for in vitro cultivation, Salmonella and other enteric bacteria exhibit a slow growth rate and a long lag phase. This growth inhibition phenomenon was known to involve the sigma factor RpoS, but the genetic basis of the repression of bacterial succinate utilisation was poorly understood. Here, we use an experimental evolution approach to isolate fast-growing mutants during growth of S. Typhimurium on succinate containing minimal medium. Our approach reveals novel RpoS-independent systems that inhibit succinate utilisation. The CspC RNA binding protein restricts succinate utilisation, an inhibition that is antagonised by high levels of the small regulatory RNA (sRNA) OxyS. We discovered that the Fe-S cluster regulatory protein IscR inhibits succinate utilisation by repressing the C4-dicarboyxlate transporter DctA. Furthermore, the ribose operon repressor RbsR is required for the complete RpoS-driven repression of succinate utilisation, suggesting a novel mechanism of RpoS regulation. Our discoveries shed light on the redundant regulatory systems that tightly regulate the utilisation of succinate. We speculate that the control of central carbon metabolism by multiple regulatory systems in Salmonella governs the infection niche-specific utilisation of succinate. Author summary: During infection, Salmonella enterica serovar Typhimurium must efficiently utilise the nutrients provided by the host and the microbiota. Within host niches colonised by Salmonella, key carbon source is succinate, which is abundant in both the gut and within macrophages. Although S. Typhimurium possesses all the molecular machinery to import and catabolise succinate, the utilisation of this carbon source is strongly inhibited under laboratory conditions. Here, we have used diverse genetic approaches to discover that several distinct genetic regulatory systems act in concert to repress succinate utilisation by Salmonella. Our study sheds new light on how carbon source utilisation is managed by this important model enteropathogenic bacterium. [ABSTRACT FROM AUTHOR]

To sustain growth in changing nutrient conditions, cells reorganize outputs of metabolic networks and appropriately reallocate resources. Signaling by reversible protein phosphorylation can control such metabolic adaptations. In contrast to kinases, the functions of phosphatases that enable metabolic adaptation as glucose depletes are poorly studied. Using a Saccharomyces cerevisiae deletion screen, we identified the PP2A-like phosphatase Ppg1 as required for appropriate carbon allocations towards gluconeogenic outputs—trehalose, glycogen, UDP-glucose, UDP-GlcNAc—after glucose depletion. This Ppg1 function is mediated via regulation of the assembly of the Far complex—a multi-subunit complex that tethers to the ER and mitochondrial outer membranes forming localized signaling hubs. The Far complex assembly is Ppg1 catalytic activity-dependent. Ppg1 regulates the phosphorylation status of multiple ser/thr residues on Far11 to enable the proper assembly of the Far complex. The assembled Far complex is required to maintain gluconeogenic outputs after glucose depletion. Glucose in turn regulates Far complex amounts. This Ppg1-mediated Far complex assembly, and Ppg1-Far complex dependent control of gluconeogenic outputs enables adaptive growth under glucose depletion. Our study illustrates how protein dephosphorylation is required for the assembly of a multi-protein scaffold present in localized cytosolic pools, to thereby alter gluconeogenic flux and enable cells to metabolically adapt to nutrient fluctuations. Author summary: Many organisms live in changing nutrient environments. To grow effectively, cells continuously adapt to changing nutrient conditions (such as glucose limitation) and reorganize their metabolic outputs. For this adaptation, cells employ 'signaling systems' that respond to nutrient conditions and regulate metabolism. Protein phosphatases in eukaryotic cells are part of one such signaling system which regulates metabolic outputs. They function by dephosphorylating target proteins to change their activity or function. In this study, we used budding yeast (Saccharomyces cerevisiae) as a model to study signaling-based adaptation to glucose limitation. Using a screen, we identified a protein phosphatase Ppg1, a member of PP2A family, as a regulator of metabolic outputs after glucose depletion. Surprisingly, we found that Ppg1 carried out this function by regulating the assembly of a large protein complex, Far complex. Ppg1 phosphatase dephosphorylated a key protein of this complex thereby allowing the complex to assemble. This assembled Far complex controlled the metabolic reorganization. This study therefore finds a novel mechanism through which cells fine-tune metabolism to thrive after glucose depletion, via the formation of a large protein complex in cells. This study deepens our understanding of signaling responses in changing environments required for appropriate metabolic adaptation. [ABSTRACT FROM AUTHOR]

We investigate the contribution of a candidate gene, fiz (fezzik), to complex polygenic adaptation to juvenile malnutrition in Drosophila melanogaster. Experimental populations maintained for >250 generations of experimental evolution to a nutritionally poor larval diet (Selected populations) evolved several-fold lower fiz expression compared to unselected Control populations. Here we show that this divergence in fiz expression is mediated by a cis-regulatory polymorphism. This polymorphism, originally sampled from a natural population in Switzerland, is distinct from a second cis-regulatory SNP previously identified in non-African D. melanogaster populations, implying that two independent cis-regulatory variants promoting high fiz expression segregate in non-African populations. Enzymatic analyses of Fiz protein expressed in E. coli demonstrate that it has ecdysone oxidase activity acting on both ecdysone and 20-hydroxyecdysone. Four of five fiz paralogs annotated to ecdysteroid metabolism also show reduced expression in Selected larvae, implying that malnutrition-driven selection favored general downregulation of ecdysone oxidases. Finally, as an independent test of the role of fiz in poor diet adaptation, we show that fiz knockdown by RNAi results in faster larval growth on the poor diet, but at the cost of greatly reduced survival. These results imply that downregulation of fiz in Selected populations was favored by selection on the nutritionally poor diet because of its role in suppressing growth in response to nutrient shortage. However, they suggest that fiz downregulation is only adaptive in combination with other changes evolved by Selected populations, which ensure that the organism can sustain the faster growth promoted by fiz downregulation. Author summary: Juvenile animals are particularly vulnerable to famines, which result in stunted growth and often lead to lifelong consequences for health and Darwinian fitness. Natural selection can alleviate consequences of juvenile undernutrition by improving the acquisition of scarce nutrients through more efficient digestion or minimizing their "waste" for non-essential tasks. However, mechanisms that regulate protein synthesis and cell division must also be fine-tuned so that the organism can grow as fast as is sustainable with the available nutrients without endangering survival. We show that the evolution of digestive and metabolic adaptations in experimental Drosophila populations exposed for >250 generations to a nutritionally poor diet was accompanied by a regulatory mutation in a gene (fiz) that modulates insect steroid hormones. Reduced expression of the fiz gene resulting from this change promotes faster larval growth on the poor diet. However, in the absence of other adaptations, the faster growth promoted by fiz downregulation cannot be sustained by larvae confronted with poor diet, resulting in very high mortality. Thus, evolution of improved tolerance to juvenile malnutrition appears to rely on a combination of multiple coordinated changes that may be detrimental in isolation. [ABSTRACT FROM AUTHOR]

The regulatory mechanism of gonadal sex differentiation, which is complex and regulated by multiple factors, remains poorly understood in teleosts. Recently, we have shown that compromised androgen and estrogen synthesis with increased progestin leads to all-male differentiation with proper testis development and spermatogenesis in cytochrome P450 17a1 (cyp17a1)-/- zebrafish. In the present study, the phenotypes of female-biased sex ratio were positively correlated with higher Fanconi anemia complementation group L (fancl) expression in the gonads of doublesex and mab-3 related transcription factor 1 (dmrt1)-/- and cyp17a1-/-;dmrt1-/- fish. The additional depletion of fancl in cyp17a1-/-;dmrt1-/- zebrafish reversed the gonadal sex differentiation from all-ovary to all-testis (in cyp17a1-/-;dmrt1-/-;fancl-/- fish). Luciferase assay revealed a synergistic inhibitory effect of Dmrt1 and androgen signaling on fancl transcription. Furthermore, an interaction between Fancl and the apoptotic factor Tumour protein p53 (Tp53) was found in vitro. The interaction between Fancl and Tp53 was observed via the WD repeat domain (WDR) and C-terminal domain (CTD) of Fancl and the DNA binding domain (DBD) of Tp53, leading to the K48-linked polyubiquitination degradation of Tp53 activated by the ubiquitin ligase, Fancl. Our results show that testis fate in cyp17a1-/- fish is determined by Dmrt1, which is thought to stabilize Tp53 by inhibiting fancl transcription during the critical stage of sexual fate determination in zebrafish. Author summary: Gonadal sex differentiation is known as the queen of problems in evolutionary biology, and the mechanisms that determine sexual fate vary widely among teleosts. Traditionally, Dmrt1 and androgen signaling have been essential for testis differentiation, and estrogen signaling has been essential for ovary differentiation. The all-testis phenotype observed in cyp17a1-/- zebrafish, has led to the conclusion that androgen signaling is dispensable for testis differentiation in zebrafish. By analyzing a series of mutant zebrafish lines, we show here that Dmrt1 sufficiently promotes all-testis differentiation in cyp17a1-deficient zebrafish albeit the compromised androgen and estrogen synthesis. In addition, we observed that zebrafish Dmrt1 and androgen signaling probably stabilize Tp53 by inhibiting the transcription of a ubiquitin ligase, Fancl. Our current study provides new insights into the interactive signals that regulate sexual fate determination in teleosts. [ABSTRACT FROM AUTHOR]

Bacteria have evolved complex transcriptional regulatory networks, as well as many diverse regulatory strategies at the RNA level, to enable more efficient use of metabolic resources and a rapid response to changing conditions. However, most RNA-based regulatory mechanisms are not well conserved across different bacterial species despite controlling genes important for virulence or essential biosynthetic processes. Here, we characterize the activity of, and assess the fitness benefit conferred by, twelve cis-acting regulatory RNAs (including several riboswitches and a T-box), in the opportunistic pathogen Streptococcus pneumoniae TIGR4. By evaluating native locus mutants of each regulator that result in constitutively active or repressed expression, we establish that growth defects in planktonic culture are associated with constitutive repression of gene expression, while constitutive activation of gene expression is rarely deleterious. In contrast, in mouse nasal carriage and pneumonia models, strains with either constitutively active and repressed gene expression are significantly less fit than matched control strains. Furthermore, two RNA-regulated pathways, FMN synthesis/transport and pyrimidine synthesis/transport display exceptional sensitivity to mis-regulation or constitutive gene repression in both planktonic culture and in vivo environments. Thus, despite lack of obvious phenotypes associated with constitutive gene expression in vitro, the fitness benefit conferred on bacteria via fine-tuned metabolic regulation through cis-acting regulatory RNAs is substantial in vivo, and therefore easily sufficient to drive the evolution and maintenance of diverse RNA regulatory mechanisms. Author summary: Regulation of gene expression is a nearly universal trait across all organisms that enables efficient utilization of resources, rapid reaction to a changing environment, or development of specialized subpopulations. However, the mechanisms that enable gene control are extraordinarily diverse. In bacteria, there are numerous regulatory mechanisms that act at the RNA level. Such mechanisms are frequently responsible for fine-tuning expression levels of critical metabolic processes or virulence factors, but are often not well conserved across divergent bacterial species. In this work, we seek to evaluate the selective pressures driving the evolution and maintenance of RNA cis-regulatory mechanisms. Using the opportunistic pathogen Streptococcus pneumoniae as a model, we measured the fitness of strains carrying native locus mutations to RNA regulators that result in constitutively active and repressed gene expression. We find that only constitutively repressed mutants show significant growth defects under stringent in vitro culture conditions, but both constitutively active and repressed mutants are significantly less fit within mouse models of infection and carriage. We also find that specific pathways such as pyrimidine and FMN synthesis/transport are very sensitive to mis-regulation and constitutive repression respectively. This study demonstrates that while individual variants may show mild fitness defects, when considered in aggregate, these defects correspond to significantly depressed fitness values that are more than sufficient to drive evolutionary trajectory for organisms with large population sizes such as S. pneumoniae. Thus, RNA-regulators responsible for fine-tuning metabolic genes are likely critical for S. pneumoniae success in a host. [ABSTRACT FROM AUTHOR]

Phenotypic plasticity is thought to be an important driver of diversification and adaptation to environmental variation, yet the genomic mechanisms mediating plastic trait development and evolution remain poorly understood. The Scarabaeinae, or true dung beetles, are a species-rich clade of insects recognized for their highly diversified nutrition-responsive development including that of cephalic horns—evolutionarily novel, secondary sexual weapons that exhibit remarkable intra- and interspecific variation. Here, we investigate the evolutionary basis for horns as well as other key dung beetle traits via comparative genomic and developmental assays. We begin by presenting chromosome-level genome assemblies of three dung beetle species in the tribe Onthophagini (> 2500 extant species) including Onthophagus taurus, O. sagittarius, and Digitonthophagus gazella. Comparing these assemblies to those of seven other species across the order Coleoptera identifies evolutionary changes in coding sequence associated with metabolic regulation of plasticity and metamorphosis. We then contrast chromatin accessibility in developing head horn tissues of high- and low-nutrition O. taurus males and females and identify distinct cis-regulatory architectures underlying nutrition- compared to sex-responsive development, including a large proportion of recently evolved regulatory elements sensitive to horn morph determination. Binding motifs of known and new candidate transcription factors are enriched in these nutrition-responsive open chromatin regions. Our work highlights the importance of chromatin state regulation in mediating the development and evolution of plastic traits, demonstrates gene networks are highly evolvable transducers of environmental and genetic signals, and provides new reference-quality genomes for three species that will bolster future developmental, ecological, and evolutionary studies of this insect group. Author summary: Phenotypic plasticity is the ability of a single genotype to produce multiple phenotypes in response to environmental variation and thus represents an important evolutionary mechanism for organismal adaptation and diversification. In this study, we investigate the genomic basis for phenotypic plasticity in Onthophagini horned dung beetles, a highly diverse (~2500 species) tribe of beetles renowned for their developmental plasticity. We assemble and annotate chromosome-level genome assembles for three Onthophagini species and identify examples of gene family and coding sequence evolution potentially associated with nutrition-responsiveness in this insect group. When then compare chromatin conformation underlying head horn development and specification in Onthophagus taurus, a species with dramatic nutrition- and sex-based differences in horn shape. We find chromatin accessibility likely plays a critical role in specifying nutritional and sexual horn dimorphisms in this species. Further, we show nutrition- and sex-responsive horn development are controlled by largely distinct, rather than shared, regulatory architectures, and the acquisition of lineage-specific regulatory elements may have played an outsized role in the evolution of nutrition-responsive development of this trait. Our results highlight the significance of chromatin accessibility and regulatory element activity in the regulation of plastic phenotypes and the highly-evolvable nature of developmental gene networks. [ABSTRACT FROM AUTHOR]

Bacterial type IV secretion systems (T4SSs) are a versatile family of macromolecular translocators, collectively able to recruit diverse DNA and protein substrates and deliver them to a wide range of cell types. Presently, there is little understanding of how T4SSs recognize substrate repertoires and form productive contacts with specific target cells. Although T4SSs are composed of a number of conserved subunits and adopt certain conserved structural features, they also display considerable compositional and structural diversity. Here, we explored the structural bases underlying the functional versatility of T4SSs through systematic deletion and subunit swapping between two conjugation systems encoded by the distantly-related IncF plasmids, pED208 and F. We identified several regions of intrinsic flexibility among the encoded T4SSs, as evidenced by partial or complete functionality of chimeric machines. Swapping of VirD4-like TraD type IV coupling proteins (T4CPs) yielded functional chimeras, indicative of relaxed specificity at the substrate—TraD and TraD—T4SS interfaces. Through mutational analyses, we further delineated domains of the TraD T4CPs contributing to recruitment of cognate vs heterologous DNA substrates. Remarkably, swaps of components comprising the outer membrane core complexes, a few F-specific subunits, or the TraA pilins supported DNA transfer in the absence of detectable pilus production. Among sequenced enterobacterial species in the NCBI database, we identified many strains that harbor two or more F-like plasmids and many F plasmids lacking one or more T4SS components required for self-transfer. We confirmed that host cells carrying co-resident, non-selftransmissible variants of pED208 and F elaborate chimeric T4SSs, as evidenced by transmission of both plasmids. We propose that T4SS plasticity enables the facile assembly of functional chimeras, and this intrinsic flexibility at the structural level can account for functional diversification of this superfamily over evolutionary time and, on a more immediate time-scale, to proliferation of transfer-defective MGEs in nature. Author summary: Mobile genetic elements (MGEs) comprise a diverse group of extrachromosomal plasmids or integrated DNA fragments that are widely distributed among many bacterial species. MGEs typically encode conjugation systems dedicated to their transmission to other bacteria, and also code for resistance to antibiotics or virulence or other fitness traits. The conjugation systems, along with an equally medically important group of translocators devoted to the interkingdom delivery of protein effectors by pathogenic species, comprise the superfamily of type IV secretion systems (T4SSs). Recent studies have defined many mechanistic and structural features of the T4SSs, yet there remains little understanding of how T4SSs recruit specific DNA or protein substrates, elaborate functional channels, and in some cases build attachment organelles termed conjugative pili. We explored the mechanics of T4SS machine function by systematically exchanging individual components between two distinct conjugation systems functioning in enterobacterial species. Through construction of chimeric machines, and further mutational analyses, we identified subunits or protein domains of conjugation machines specifying recruitment of different DNA substrates or selectively contributing to assembly of translocation channels or conjugative pili. Such features of T4SSs are prime targets for development of inhibitory strategies aimed at blocking T4SS functions for therapeutic intervention. [ABSTRACT FROM AUTHOR]

The honey bee trypanosomatid parasite, Lotmaria passim, contains two genes that encode the flagellar calcium binding protein (FCaBP) through tandem duplication in its genome. FCaBPs localize in the flagellum and entire body membrane of L. passim through specific N-terminal sorting sequences. This finding suggests that this is an example of protein subcellular relocalization resulting from gene duplication, altering the intracellular localization of FCaBP. However, this phenomenon may not have occurred in Leishmania, as one or both of the duplicated genes have become pseudogenes. Multiple copies of the FCaBP gene are present in several Trypanosoma species and Leptomonas pyrrhocoris, indicating rapid evolution of this gene in trypanosomatid parasites. The N-terminal flagellar sorting sequence of L. passim FCaBP1 is in close proximity to the BBSome complex, while that of Trypanosoma brucei FCaBP does not direct GFP to the flagellum in L. passim. Deletion of the two FCaBP genes in L. passim affected growth and impaired flagellar morphogenesis and motility, but it did not impact host infection. Therefore, FCaBP represents a duplicated gene with a rapid evolutionary history that is essential for flagellar structure and function in a trypanosomatid parasite. Author summary: Protein subcellular relocalization (PSR) was proposed as a mechanism for the functional divergence and retention of duplicate genes. We explored this hypothesis using flagellar calcium binding protein (FCaBP) genes duplicated in many trypanosomatid parasites. FCaBPs localize in the flagellum and entire body membrane of honey bee trypanosomatid parasite, Lotmaria passim through specific N-terminal sorting sequences. However, this is not common with all trypanosomatid parasites since one or both of the duplicated genes have become pseudogenes in Leishmania. N-terminal flagellar sorting sequence of FCaBP1 interacts with BBSome complex, and thus they must have co-evolved in each trypanosomatid species. FCaBPs are essential for the normal growth, flagellar morphogenesis, and motility but not host infection of L. passim. Our findings demonstrate that duplicated FCaBP genes have undergone PSR by changing amino acids in the N-terminal end with some but not all trypanosomatid parasites. In the parasites with multiple FCaBP genes, they play critical roles for the growth control as well as flagellar structure and function. [ABSTRACT FROM AUTHOR]

The basement membrane (BM) is an essential structural element of tissues, and its diversification participates in organ morphogenesis. However, the traffic routes associated with BM formation and the mechanistic modulations explaining its diversification are still poorly understood. Drosophila melanogaster follicular epithelium relies on a BM composed of oriented BM fibrils and a more homogenous matrix. Here, we determined the specific molecular identity and cell exit sites of BM protein secretory routes. First, we found that Rab10 and Rab8 define two parallel routes for BM protein secretion. When both routes were abolished, BM production was fully blocked; however, genetic interactions revealed that these two routes competed. Rab10 promoted lateral and planar-polarized secretion, whereas Rab8 promoted basal secretion, leading to the formation of BM fibrils and homogenous BM, respectively. We also found that the dystrophin-associated protein complex (DAPC) and Rab10 were both present in a planar-polarized tubular compartment containing BM proteins. DAPC was essential for fibril formation and sufficient to reorient secretion towards the Rab10 route. Moreover, we identified a dual function for the exocyst complex in this context. First, the Exo70 subunit directly interacted with dystrophin to limit its planar polarization. Second, the exocyst complex was also required for the Rab8 route. Altogether, these results highlight important mechanistic aspects of BM protein secretion and illustrate how BM diversity can emerge from the spatial control of distinct traffic routes. Author summary: Diversification of the basement membrane (BM) is crucial for tissue structure and organ development, yet the mechanisms governing its formation and diversification remain unclear. We focused on the Drosophila melanogaster follicular epithelium, characterizing BM protein secretion pathways. We identified Rab10 and Rab8 as key regulators, orchestrating parallel secretion routes. Rab10 promotes lateral and planar-polarized secretion, while Rab8 facilitates basal secretion. As a result of this highly polarized system, Rab10 route induces the formation of BM fibrils while Rab8 route generate homogenous BM. Additionally, dystrophin-associated protein complex (DAPC) is involved in fibril formation and secretion route orientation towards Rab10. Furthermore, we discovered the involvement of the exocyst complex, notably its Exo70 subunit that it interacts with dystrophin, impacting its planar polarization, and that is crucial for the Rab8 route. These findings deepen our understanding of BM protein secretion mechanisms, emphasizing the role of distinct traffic routes in BM diversity emergence. Moreover, it reveals new functions and molecular interactions for the DAPC that could be relevant for the many congenital diseases involving this complex. [ABSTRACT FROM AUTHOR]

Recent human genome-wide association studies have identified common missense variants in MARC1, p.Ala165Thr and p.Met187Lys, associated with lower hepatic fat, reduction in liver enzymes and protection from most causes of cirrhosis. Using an exome-wide association study we recapitulated earlier MARC1 p.Ala165Thr and p.Met187Lys findings in 540,000 individuals from five ancestry groups. We also discovered novel rare putative loss of function variants in MARC1 with a phenotype similar to MARC1 p.Ala165Thr/p.Met187Lys variants. In vitro studies of recombinant human MARC1 protein revealed Ala165Thr substitution causes protein instability and aberrant localization in hepatic cells, suggesting MARC1 inhibition or deletion may lead to hepatoprotection. Following this hypothesis, we generated Marc1 knockout mice and evaluated the effect of Marc1 deletion on liver phenotype. Unexpectedly, our study found that whole-body Marc1 deficiency in mouse is not protective against hepatic triglyceride accumulation, liver inflammation or fibrosis. In attempts to explain the lack of the observed phenotype, we discovered that Marc1 plays only a minor role in mouse liver while its paralogue Marc2 is the main Marc family enzyme in mice. Our findings highlight the major difference in MARC1 physiological function between human and mouse. Author summary: Non-alcoholic fatty liver disease (NAFLD) affects almost a third population worldwide and is the main cause of chronic liver disease and cirrhosis. Recent human genetics studies identified several common variants in MARC1 gene, associated with lower hepatic fat, reduction in liver enzymes and protection from most causes of cirrhosis, pointing to MARC1 as a potential therapeutic target. Yet, the hepatoprotective mechanism of MARC1 loss of function is unknown. In this study, we expand human genetic analysis to 540,000 individuals. We confirm previous findings and identify novel rare variants in MARC1 that similarly associated with reduction in liver fat, liver enzymes and protection from cirrhosis. We also confirm that most common variant in MARC1 is a true loss of function. We knockdown Marc1 in mice and challenge them with NAFLD-causing diets to understand its role in liver disease. We find loss of Marc1 in mice did not lead to hepatoprotective effect observed in humans. To understand the discrepancy, we knockdown Marc1 and its paralogue Marc2 from primary hepatocytes and discover that Marc1 plays only a minor role in mouse liver while its paralogue Marc2 is the main Marc family enzyme in mice. [ABSTRACT FROM AUTHOR]

During meiosis, genetic recombination is initiated by the formation of many DNA double-strand breaks (DSBs) catalysed by the evolutionarily conserved topoisomerase-like enzyme, Spo11, in preferred genomic sites known as hotspots. DSB formation activates the Tel1/ATM DNA damage responsive (DDR) kinase, locally inhibiting Spo11 activity in adjacent hotspots via a process known as DSB interference. Intriguingly, in S. cerevisiae, over short genomic distances (<15 kb), Spo11 activity displays characteristics of concerted activity or clustering, wherein the frequency of DSB formation in adjacent hotspots is greater than expected by chance. We have proposed that clustering is caused by a limited number of sub-chromosomal domains becoming primed for DSB formation. Here, we provide evidence that DSB clustering is abolished when meiotic prophase timing is extended via deletion of the NDT80 transcription factor. We propose that extension of meiotic prophase enables most cells, and therefore most chromosomal domains within them, to reach an equilibrium state of similar Spo11-DSB potential, reducing the impact that priming has on estimates of coincident DSB formation. Consistent with this view, when Tel1 is absent but Ndt80 is present and thus cells are able to rapidly exit meiotic prophase, genome-wide maps of Spo11-DSB formation are skewed towards pericentromeric regions and regions that load pro-DSB factors early—revealing regions of preferential priming—but this effect is abolished when NDT80 is deleted. Our work highlights how the stochastic nature of Spo11-DSB formation in individual cells within the limited temporal window of meiotic prophase can cause localised DSB clustering—a phenomenon that is exacerbated in tel1Δ cells due to the dual roles that Tel1 has in DSB interference and meiotic prophase checkpoint control. Author summary: Genetic variation arises in sexually reproducing organisms via the combination of two processes: outbreeding and meiosis. Outbreeding ensures that individuals are unique composites of their genetically distinct parents, whereas meiosis—a specialised form of cell division—creates genetic variation within the gametes used during breeding (eggs and sperm in humans). During the early stages of meiosis—termed prophase—hundreds of DNA breaks are generated within parental chromosomes. Repair of these DNA breaks generates novel combinations of gene types (alleles). Despite many steps of meiosis being evolutionarily conserved from yeast to humans, it is poorly understood how DNA break formation is regulated to ensure that DNA breaks are evenly spread across all chromosomes in order to generate genetic diversity. Here, we utilise the model eukaryote, Saccharomyces cerevisiae, (budding yeast) to investigate the interplay between a key protein involved in DNA break recognition (Tel1, the yeast version of the mammalian gene, ATM), and the length of meiotic prophase controlled by the transcription factor Ndt80. Our observations indicate that the clustering of DNA breaks that happens when Tel1 is absent is suppressed by extending the time window of meiotic prophase, providing novel insight into the regulation of genetic variation within sexually reproducing organisms. [ABSTRACT FROM AUTHOR]

The use of artificial intelligence (AI) for facial diagnostics is increasingly used in the genetics clinic to evaluate patients with potential genetic conditions. Current approaches focus on one type of AI called Deep Learning (DL). While DL- based facial diagnostic platforms have a high accuracy rate for many conditions, less is understood about how this technology assesses and classifies (categorizes) images, and how this compares to humans. To compare human and computer attention, we performed eye-tracking analyses of geneticist clinicians (n = 22) and non-clinicians (n = 22) who viewed images of people with 10 different genetic conditions, as well as images of unaffected individuals. We calculated the Intersection-over-Union (IoU) and Kullback–Leibler divergence (KL) to compare the visual attentions of the two participant groups, and then the clinician group against the saliency maps of our deep learning classifier. We found that human visual attention differs greatly from DL model's saliency results. Averaging over all the test images, IoU and KL metric for the successful (accurate) clinician visual attentions versus the saliency maps were 0.15 and 11.15, respectively. Individuals also tend to have a specific pattern of image inspection, and clinicians demonstrate different visual attention patterns than non-clinicians (IoU and KL of clinicians versus non-clinicians were 0.47 and 2.73, respectively). This study shows that humans (at different levels of expertise) and computer vision model examine images differently. Understanding these differences can improve the design and use of AI tools, and lead to more meaningful interactions between clinicians and AI technologies. Author summary: Artificial intelligence (AI) is increasingly used in medicine. In clinical practice, medical geneticists often use AI tools to help them examine the facial features of a patient who might have a genetic condition. While these tools are very popular, less is understood about how they work, and which parts of the face are most important to the AI tools, especially compared to medical geneticists. To address this, we performed a study where medical geneticists (as well as non-clinicians) looked at pictures of people with and without genetic conditions. We used eye-tracking tools to visualize which parts of the images the medical geneticists preferentially looked at. We compared this to the parts of the image that were the most important to the AI tools. We found that the medical geneticists and the AI tools tended to pay attention to very different features. We also found that medical geneticists and non-clinicians look at different parts of the images. Understanding how AI tools examine images compared to clinicians can help ensure the tools function properly, and could also help perform tasks like alerting medical geneticists to important clinical findings that could otherwise be overlooked. [ABSTRACT FROM AUTHOR]

TOP2 inhibitors (TOP2i) are effective drugs for breast cancer treatment. However, they can cause cardiotoxicity in some women. The most widely used TOP2i include anthracyclines (AC) Doxorubicin (DOX), Daunorubicin (DNR), Epirubicin (EPI), and the anthraquinone Mitoxantrone (MTX). It is unclear whether women would experience the same adverse effects from all drugs in this class, or if specific drugs would be preferable for certain individuals based on their cardiotoxicity risk profile. To investigate this, we studied the effects of treatment of DOX, DNR, EPI, MTX, and an unrelated monoclonal antibody Trastuzumab (TRZ) on iPSC-derived cardiomyocytes (iPSC-CMs) from six healthy females. All TOP2i induce cell death at concentrations observed in cancer patient serum, while TRZ does not. A sub-lethal dose of all TOP2i induces limited cellular stress but affects calcium handling, a function critical for cardiomyocyte contraction. TOP2i induce thousands of gene expression changes over time, giving rise to four distinct gene expression response signatures, denoted as TOP2i early-acute, early-sustained, and late response genes, and non-response genes. There is no drug- or AC-specific signature. TOP2i early response genes are enriched in chromatin regulators, which mediate AC sensitivity across breast cancer patients. However, there is increased transcriptional variability between individuals following AC treatments. To investigate potential genetic effects on response variability, we first identified a reported set of expression quantitative trait loci (eQTLs) uncovered following DOX treatment in iPSC-CMs. Indeed, DOX response eQTLs are enriched in genes that respond to all TOP2i. Next, we identified 38 genes in loci associated with AC toxicity by GWAS or TWAS. Two thirds of the genes that respond to at least one TOP2i, respond to all ACs with the same direction of effect. Our data demonstrate that TOP2i induce thousands of shared gene expression changes in cardiomyocytes, including genes near SNPs associated with inter-individual variation in response to DOX treatment and AC-induced cardiotoxicity. Author summary: Anthracycline drugs such as Doxorubicin are effective treatments for breast cancer; however, they can cause cardiotoxicity in some women. It is unclear whether women would experience the same toxicity for all drugs in this class, or whether specific drugs would be better tolerated in specific individuals. We used an in vitro system of induced pluripotent stem cell-derived cardiomyocytes from six healthy females to test the effects of five breast cancer drugs on cell heath and global gene expression. We identified a strong shared cellular and gene expression response to drugs from the same class. However, there is more variation in gene expression levels between individuals following treatment with each anthracycline compared to untreated cells. We found that many genes in regions previously associated with Doxorubicin-induced cardiotoxicity in cancer patients, respond to at least two drugs in the class. This suggests that drugs in the same class induce similar effects on an individual's heart. This work contributes to our understanding of how drug response, in the context of off-target effects, varies across individuals. [ABSTRACT FROM AUTHOR]

Colorectal cancer (CRC) is a major cause of cancer mortality and a serious health problem worldwide. Mononuclear phagocytes are the main immune cells in the tumor microenvironment of CRC with remarkable plasticity, and current studies show that macrophages are closely related to tumor progression, invasion and dissemination. To understand the immunological function of mononuclear phagocytes comprehensively and deeply, we use single-cell RNA sequencing and classify mononuclear phagocytes in CRC into 6 different subsets, and characterize the heterogeneity of each subset. We find that tissue inhibitor of metalloproteinases (TIMPs) involved in the differentiation of proinflammatory and anti-inflammatory mononuclear phagocytes. Trajectory of circulating monocytes differentiation into tumor-associated macrophages (TAMs) and the dynamic changes at levels of transcription factor (TF) regulons during differentiation were revealed. We also find that C5 subset, characterized by activation of lipid metabolism, is in the terminal state of differentiation, and that the abundance of C5 subset is negatively correlated with CRC patients' prognosis. Our findings advance the understanding of circulating monocytes' differentiation into macrophages, identify a new subset associated with CRC prognosis, and reveal a set of TF regulons regulating mononuclear phagocytes differentiation, which are expected to be potential therapeutic targets for reversing immunosuppressive tumor microenvironment. Author summary: Colorectal cancer (CRC) is a serious health problem worldwide. Mononuclear phagocytes are the main immune cells in the tumor microenvironment of CRC with remarkable plasticity. In this study, single-cell RNA sequencing technology was used to study the subsets and characteristics of mononuclear phagocytes in CRC. We classify mononuclear phagocytes in CRC into 6 different subsets and demonstrate the heterogeneity in infiltrating mononuclear phagocytes of CRC. We find that tissue inhibitor of metalloproteinases (TIMPs) involved in the differentiation of proinflammatory and anti-inflammatory mononuclear phagocytes and reveal the trajectory and characteristics of circulating monocytes differentiation into tumor-associated macrophages (TAMs). We also find a new immunosuppressive subset of mononuclear phagocytes, which is associated with the prognosis of CRC patients. Our findings identify the heterogeneity in infiltrating mononuclear phagocytes of CRC, find a new subset associated with CRC prognosis. Also, we reveal the differentiation of mononuclear phagocytes would lead to an immunosuppressive tumor microenvironment and provide potential targets for reversing the process. [ABSTRACT FROM AUTHOR]

The eye is instrumental for controlling circadian rhythms in mice and human. Here, we address the conservation of this function in the zebrafish, a diurnal vertebrate. Using lakritz (lak) mutant larvae, which lack retinal ganglion cells (RGCs), we show that while a functional eye contributes to masking, it is largely dispensable for the establishment of circadian rhythms of locomotor activity. Furthermore, the eye is dispensable for the induction of a phase delay following a pulse of white light at CT 16 but contributes to the induction of a phase advance upon a pulse of white light at CT21. Melanopsin photopigments are important mediators of photoentrainment, as shown in nocturnal mammals. One of the zebrafish melanopsin genes, opn4xa, is expressed in RGCs but also in photosensitive projection neurons in the pineal gland. Pineal opn4xa+ projection neurons function in a LIGHT ON manner in contrast to other projection neurons which function in a LIGHT OFF mode. We generated an opn4xa mutant in which the pineal LIGHT ON response is impaired. This mutation has no effect on masking and circadian rhythms of locomotor activity, or for the induction of phase shifts, but slightly modifies period length when larvae are subjected to constant light. Finally, analysis of opn4xa;lak double mutant larvae did not reveal redundancy between the function of the eye and opn4xa in the pineal for the control of phase shifts after light pulses. Our results support the idea that the eye is not the sole mediator of light influences on circadian rhythms of locomotor activity and highlight differences in the circadian system and photoentrainment of behaviour between different animal models. Author summary: Experiments performed in mice have established a crucial role for the eye in general and melanopsin expressing cells in particular in the control of circadian rhythms most notably during photoentrainment, by which circadian rhythms adapt to a changing light environment. In marked contrast to this, we show that in zebrafish the eye and photosensitivity dependent on one of the melanopsin genes, opn4xa, which is expressed in both the eye and the pineal gland, are largely dispensable for correct circadian rhythms. These results provide insight that the light sensors orchestrating circadian rhythms of locomotor activity are different between animal models revealing that vertebrates employ different molecular/cellular circuits for photoentrainment of behavior depending on their phylogeny and/or temporal niche. [ABSTRACT FROM AUTHOR]

Defects in protein homeostasis can induce proteotoxic stress, affecting cellular fitness and, consequently, overall tissue health. In various growing tissues, cell competition based mechanisms facilitate detection and elimination of these compromised, often referred to as 'loser', cells by the healthier neighbors. The precise connection between proteotoxic stress and competitive cell survival remains largely elusive. Here, we reveal the function of an endoplasmic reticulum (ER) and Golgi localized protein Rer1 in the regulation of protein homeostasis in the developing Drosophila wing epithelium. Our results show that loss of Rer1 leads to proteotoxic stress and PERK-mediated phosphorylation of eukaryotic initiation factor 2α. Clonal analysis showed that rer1 mutant cells are identified as losers and eliminated through cell competition. Interestingly, we find that Rer1 levels are upregulated upon Myc-overexpression that causes overgrowth, albeit under high proteotoxic stress. Our results suggest that increased levels of Rer1 provide cytoprotection to Myc-overexpressing cells by alleviating the proteotoxic stress and thereby supporting Myc-driven overgrowth. In summary, these observations demonstrate that Rer1 acts as a novel regulator of proteostasis in Drosophila and reveal its role in competitive cell survival. Author summary: In developing tissues, cells can stochastically acquire defects that can reduce their fitness. To maintain the overall health of tissues, these unfit cells are identified by the healthier neighboring cells and eliminated via a juxtacrine-acting cellular fitness sensing mechanism called cell competition. An example of such physiological regulation of cellular fitness is the maintenance of proteostasis. Defects in maintaining proteostasis cause proteotoxic stress. Interestingly, proteotoxic stress is observed not only in the unfit loser cells but also in the overgrowing super-competitor cells, for instance, cells with higher levels of Myc. How cell competition is linked to the maintenance of proteostasis is poorly understood. In this study, we have characterized for the first time the function of Drosophila Rer1 protein in development. We demonstrate that Rer1 is essential for maintaining protein homeostasis and loss of Rer1 activates stress-induced unfolded protein responses. Cells lacking Rer1 are identified as unfit cells and become losers when juxtaposed to the normal neighboring cells. Moreover, we show that Myc-overexpressing cells upregulate Rer1 levels, which allows them to maintain a higher demand for stress regulation, caused by increased protein translation. In this work, we propose that Rer1 functions as a stress regulator and that modulating its levels could provide cytoprotection under stress conditions. [ABSTRACT FROM AUTHOR]

A growing body of evidence suggests that gene flow between closely related species is a widespread phenomenon. Alleles that introgress from one species into a close relative are typically neutral or deleterious, but sometimes confer a significant fitness advantage. Given the potential relevance to speciation and adaptation, numerous methods have therefore been devised to identify regions of the genome that have experienced introgression. Recently, supervised machine learning approaches have been shown to be highly effective for detecting introgression. One especially promising approach is to treat population genetic inference as an image classification problem, and feed an image representation of a population genetic alignment as input to a deep neural network that distinguishes among evolutionary models (i.e. introgression or no introgression). However, if we wish to investigate the full extent and fitness effects of introgression, merely identifying genomic regions in a population genetic alignment that harbor introgressed loci is insufficient—ideally we would be able to infer precisely which individuals have introgressed material and at which positions in the genome. Here we adapt a deep learning algorithm for semantic segmentation, the task of correctly identifying the type of object to which each individual pixel in an image belongs, to the task of identifying introgressed alleles. Our trained neural network is thus able to infer, for each individual in a two-population alignment, which of those individual's alleles were introgressed from the other population. We use simulated data to show that this approach is highly accurate, and that it can be readily extended to identify alleles that are introgressed from an unsampled "ghost" population, performing comparably to a supervised learning method tailored specifically to that task. Finally, we apply this method to data from Drosophila, showing that it is able to accurately recover introgressed haplotypes from real data. This analysis reveals that introgressed alleles are typically confined to lower frequencies within genic regions, suggestive of purifying selection, but are found at much higher frequencies in a region previously shown to be affected by adaptive introgression. Our method's success in recovering introgressed haplotypes in challenging real-world scenarios underscores the utility of deep learning approaches for making richer evolutionary inferences from genomic data. Author summary: It is now known that a sizeable fraction of species occasionally hybridize with related species. Thus, many species harbor genetic material that traces its ancestry to closely related species. For example, many humans contain DNA that was "introgressed" from Neanderthals. The growing appreciation of the commonality of introgression has sparked a keen interest in determining which portions of the genome were introgressed. Several statistical approaches have been devised for identifying the population genetic signatures of introgression, but the most powerful techniques for this task take advantage of modern machine learning techniques. Here, we describe a deep learning method for identifying segments of introgressed DNA. This method is based on neural networks used to determine which pixels in an image belong to which type of object. By treating a matrix of genotypes from a sample of individuals from two closely related species, we can use this deep learning approach to accurately infer which portions of which genomes from the first population were introgressed from the second, and vice-versa. We show that our method, which we have released as an open-source software package, is highly accurate using a variety of simulated scenarios and a real test case from the genus Drosophila. [ABSTRACT FROM AUTHOR]

Common genetic variants in the repressive GATA-family transcription factor (TF) TRPS1 locus are associated with breast cancer risk, and luminal breast cancer cell lines are particularly sensitive to TRPS1 knockout. We introduced an inducible degron tag into the native TRPS1 locus within a luminal breast cancer cell line to identify the direct targets of TRPS1 and determine how TRPS1 mechanistically regulates gene expression. We acutely deplete over 80 percent of TRPS1 from chromatin within 30 minutes of inducing degradation. We find that TRPS1 regulates transcription of hundreds of genes, including those related to estrogen signaling. TRPS1 directly regulates chromatin structure, which causes estrogen receptor alpha (ER) to redistribute in the genome. ER redistribution leads to both repression and activation of dozens of ER target genes. Downstream from these primary effects, TRPS1 depletion represses cell cycle-related gene sets and reduces cell doubling rate. Finally, we show that high TRPS1 activity, calculated using a gene expression signature defined by primary TRPS1-regulated genes, is associated with worse breast cancer patient prognosis. Taken together, these data suggest a model in which TRPS1 modulates the genomic distribution of ER, both activating and repressing transcription of genes related to cancer cell fitness. Author summary: Breast cancer is the most common cancer among women. The majority of cases are luminal, which tend to be estrogen receptor alpha (ER)-positive. ER is well-studied among transcription factors (TFs) because it is ligand-activated. This allows for the rapid induction of ER activity and the identification of primary estrogen-responsive genes. Most TFs have not been so extensively characterized, because their activity is not so rapidly perturbable. TRPS1 is an atypical GATA family TF that is associated with corepressor complexes and transcriptional repression. Here, we use an inducible degron tag system to rapidly deplete endogenously-tagged TRPS1 in luminal breast cancer cells within 30 minutes. We find that TRPS1 directly decreases local chromatin accessibility. This decreases ER binding intensity at TRPS1-proximal ER binding sites. As an indirect effect, ER binding intensity distal to TRPS1 increases in intensity. These effects on ER binding are associated with changes in ER target gene transcription, repressing or activating these genes in concordance with the effect on ER binding intensity. Our work is consistent with a model in which TFs either exclusively activate or exclusively repress transcription of their direct target genes, with indirect primary response genes changing due to the redistribution of limiting activating TFs or coactivators. [ABSTRACT FROM AUTHOR]

Filamentous fungi display allorecognition genes that trigger regulated cell death (RCD) when strains of unlike genotype fuse. Podospora anserina is one of several model species for the study of this allorecognition process termed heterokaryon or vegetative incompatibility. Incompatibility restricts transmission of mycoviruses between isolates. In P. anserina, genetic analyses have identified nine incompatibility loci, termed het loci. Here we set out to clone the genes controlling het-B incompatibility. het-B displays two incompatible alleles, het-B1 and het-B2. We find that the het-B locus encompasses two adjacent genes, Bh and Bp that exist as highly divergent allelic variants (Bh1/Bh2 and Bp1/Bp2) in the incompatible haplotypes. Bh encodes a protein with an N-terminal HET domain, a cell death inducing domain bearing homology to Toll/interleukin-1 receptor (TIR) domains and a C-terminal domain with a predicted lectin fold. The Bp product is homologous to PII-like proteins, a family of small trimeric proteins acting as sensors of adenine nucleotides in bacteria. We show that although the het-B system appears genetically allelic, incompatibility is in fact determined by the non-allelic Bh1/Bp2 interaction while the reciprocal Bh2/Bp1 interaction plays no role in incompatibility. The highly divergent C-terminal lectin fold domain of BH determines recognition specificity. Population studies and genome analyses indicate that het-B is under balancing selection with trans-species polymorphism, highlighting the evolutionary significance of the two incompatible haplotypes. In addition to emphasizing anew the central role of TIR-like HET domains in fungal RCD, this study identifies novel players in fungal allorecognition and completes the characterization of the entire het gene set in that species. Author summary: Many cellular life forms display genetic systems that protect individuality and discriminate conspecific self from non-self. In filamentous fungi, cell fusion events between strains are under check by specific allorecognition genes that trigger regulated cell death upon detection of non-self. The role of incompatibility is to restrict mycovirus transmission and conspecific parasitism. Podospora anserina, a good model for the study of this form of allorecognition, harbors nine incompatibility het loci. Previous studies have revealed that these genes can be homologous to genes with immune functions in other phyla including bacteria, plants and animals. We have cloned het-B, the last of the nine het genes that remained to be identified and found that it is a complex locus comprising two adjacent genes Bh and Bp. BH displays an N-terminal HET domain (related to TIR domains) and a C-terminal domain with a predicted lectin fold. BP is homologous to PII-like proteins, known bacterial metabolite sensors. Intriguingly, despite apparent genetic allelism, incompatibility is dictated by the non-allelic Bh/Bp interaction. This study stresses the reoccurring involvement of HET domains in fungal RCD and signs completion of the characterization of the entire set of het loci in that species, enabling a comparative analysis of the different genetic architectures underlying allorecognition. [ABSTRACT FROM AUTHOR]

Lipid dyshomeostasis has been implicated in a variety of diseases ranging from obesity to neurodegenerative disorders such as Neurodegeneration with Brain Iron Accumulation (NBIA). Here, we uncover the physiological role of Nazo, the Drosophila melanogaster homolog of the NBIA-mutated protein–c19orf12, whose function has been elusive. Ablation of Drosophila c19orf12 homologs leads to dysregulation of multiple lipid metabolism genes. nazo mutants exhibit markedly reduced gut lipid droplet and whole-body triglyceride contents. Consequently, they are sensitive to starvation and oxidative stress. Nazo is required for maintaining normal levels of Perilipin-2, an inhibitor of the lipase–Brummer. Concurrent knockdown of Brummer or overexpression of Perilipin-2 rescues the nazo phenotype, suggesting that this defect, at least in part, may arise from diminished Perilipin-2 on lipid droplets leading to aberrant Brummer-mediated lipolysis. Our findings potentially provide novel insights into the role of c19orf12 as a possible link between lipid dyshomeostasis and neurodegeneration, particularly in the context of NBIA. Author summary: Lipids such as triacylglycerols comprise a class of biomolecules that serve important biological functions and are therefore fundamental to life. Thus, it is not surprising that defects in lipid biology cause multiple human diseases such as obesity and diabetes. As lipids perform crucial structural and functional roles in the animal brain, disruption of lipid metabolism is also implicated in various neurodegenerative diseases including Alzheimer's and Parkinson's disease. In this study, we decipher the function of the Drosophila melanogaster gene nazo, whose human counterpart–c19orf12, is mutated in a familial neurodegenerative disorder Neurodegeneration with Brain Iron Accumulation (NBIA). The function of this gene is largely unknown. Drosophila is a powerful genetic system that has informed us about various aspects of human biology. Using this organism, we show that nazo is required for maintenance of lipid droplets, cell organelles that store triacylglycerols and other lipids. Loss of this gene has significant impacts on the life span and health of the animal. Our study will provide the foundation for future investigations into the role of human c19orf12 and the cause of NBIA. [ABSTRACT FROM AUTHOR]

Genome-wide genealogies of multiple species carry detailed information about demographic and selection processes on individual branches of the phylogeny. Here, we introduce TRAILS, a hidden Markov model that accurately infers time-resolved population genetics parameters, such as ancestral effective population sizes and speciation times, for ancestral branches using a multi-species alignment of three species and an outgroup. TRAILS leverages the information contained in incomplete lineage sorting fragments by modelling genealogies along the genome as rooted three-leaved trees, each with a topology and two coalescent events happening in discretized time intervals within the phylogeny. Posterior decoding of the hidden Markov model can be used to infer the ancestral recombination graph for the alignment and details on demographic changes within a branch. Since TRAILS performs posterior decoding at the base-pair level, genome-wide scans based on the posterior probabilities can be devised to detect deviations from neutrality. Using TRAILS on a human-chimp-gorilla-orangutan alignment, we recover speciation parameters and extract information about the topology and coalescent times at high resolution. Author summary: DNA sequences can be compared to reconstruct the evolutionary history of different species. While the ancestral history is usually represented by a single phylogenetic tree, speciation is a more complex process, and, due to the effect of recombination, different parts of the genome might follow different genealogies. For example, even though humans are more closely related to chimps than to gorillas, around 15% of our genome is more similar to the gorilla genome than to the chimp one. Even for those parts of the genome that do follow the same human-chimp topology, we might encounter a last common ancestor at different time points in the past for different genomic fragments. Here, we present TRAILS, a new framework that utilizes the information contained in all these genealogies to reconstruct the speciation process. TRAILS infers unbiased estimates of the speciation times and the ancestral effective population sizes, improving the accuracy when compared to previous methods. TRAILS also reconstructs the genealogy at the highest resolution, inferring, for example, when common ancestry was found for different parts of the genome. This information can also be used to detect deviations from neutrality, effectively inferring natural selection that happened millions of years ago. We validate the method using extensive simulations, and we apply TRAILS to a human-chimp-gorilla multiple genome alignment, from where we recover speciation parameters that are in good agreement with previous estimates. [ABSTRACT FROM AUTHOR]

The detrimental health effects of smoking are well-known, but the impact of regular nicotine use without exposure to the other constituents of tobacco is less clear. Given the increasing daily use of alternative nicotine delivery systems, such as e-cigarettes, it is increasingly important to understand and separate the effects of nicotine use from the impact of tobacco smoke exposure. Using a multivariable Mendelian randomisation framework, we explored the direct effects of nicotine compared with the non-nicotine constituents of tobacco smoke on health outcomes (lung cancer, chronic obstructive pulmonary disease [COPD], forced expiratory volume in one second [FEV-1], forced vital capacity [FVC], coronary heart disease [CHD], and heart rate [HR]). We used Genome-Wide Association Study (GWAS) summary statistics from Buchwald and colleagues, the GWAS and Sequencing Consortium of Alcohol and Nicotine, the International Lung Cancer Consortium, and UK Biobank. Increased nicotine metabolism increased the risk of COPD, lung cancer, and lung function in the univariable analysis. However, when accounting for smoking heaviness in the multivariable analysis, we found that increased nicotine metabolite ratio (indicative of decreased nicotine exposure per cigarette smoked) decreases heart rate (b = -0.30, 95% CI -0.50 to -0.10) and lung function (b = -33.33, 95% CI -41.76 to -24.90). There was no clear evidence of an effect on the remaining outcomes. The results suggest that these smoking-related outcomes are not due to nicotine exposure but are caused by the other components of tobacco smoke; however, there are multiple potential sources of bias, and the results should be triangulated using evidence from a range of methodologies. Author summary: Although we know that smoking tobacco negatively impacts health, we know relatively little about whether nicotine plays a role in causing poor health outcomes. When used for short periods of time, nicotine appears to have little impact on health. However, until recently, nicotine has rarely been used for long periods without accompanying exposure to tobacco smoke, so it is hard to disentangle the effects of regular nicotine use from the effects of tobacco smoke exposure. In this study, we aimed to dissect the effects of nicotine versus non-nicotine constituents of tobacco smoke on heart and lung health using multivariable Mendelian randomisation and data from a range of sources (including UK Biobank). We found that nicotine does not appear to be an independent cause of poor lung function, lung cancer, chronic obstructive pulmonary disease, or coronary heart disease, but does increase heart rate. These results support previous evidence which suggests that nicotine on its own does not directly cause poor health outcomes, with the exception of increasing heart rate. [ABSTRACT FROM AUTHOR]

The presence of large protein inclusions is a hallmark of neurodegeneration, and yet the precise molecular factors that contribute to their formation remain poorly understood. Screens using aggregation-prone proteins have commonly relied on downstream toxicity as a readout rather than the direct formation of aggregates. Here, we combined a genome-wide CRISPR knockout screen with Pulse Shape Analysis, a FACS-based method for inclusion detection, to identify direct modifiers of TDP-43 aggregation in human cells. Our screen revealed both canonical and novel proteostasis genes, and unearthed SRRD, a poorly characterized protein, as a top regulator of protein inclusion formation. APEX biotin labeling reveals that SRRD resides in proximity to proteins that are involved in the formation and breakage of disulfide bonds and to intermediate filaments, suggesting a role in regulation of the spatial dynamics of the intermediate filament network. Indeed, loss of SRRD results in aberrant intermediate filament fibrils and the impaired formation of aggresomes, including blunted vimentin cage structure, during proteotoxic stress. Interestingly, SRRD also localizes to aggresomes and unfolded proteins, and rescues proteotoxicity in yeast whereby its N-terminal low complexity domain is sufficient to induce this affect. Altogether this suggests an unanticipated and broad role for SRRD in cytoskeletal organization and cellular proteostasis. Author summary: The presence of large protein inclusions is a hallmark of many neurodegenerative diseases, yet the precise mechanisms by which cells compartmentalize unfolded proteins is still not completely understood. Here we used a novel screening approach that enables FACS-based inclusion detection, to identify direct modifiers of TDP-43 aggregation in human cells. Our screen revealed both canonical and novel proteostasis genes, and unearthed SRRD, a poorly characterized protein, as a top regulator of protein inclusion formation. In follow up experiments, using both proximity labeling and imaging, we show that SRRD is involved in the regulation of intermediate filaments (IF) spatial organization. Loss of SRRD results in fragmented IF network and the reduced formation of aggresomes, which are transient compartments to which cells transfer unfolded proteins under cellular stress. Interestingly, without SRRD, aggresomes completely lack vimentin cages, which are typical structures that encapsulate aggresomes and have been associated with the recruitment of protein degradation and chaperones machinery to these structures. Altogether, we present a novel screening approach for the identification of gene associated with cellular proteostasis. We reveal several novel protein factors, including SRRD, which our data suggests has a broad and previously overlooked role cytoskeletal organization and protein quality control. [ABSTRACT FROM AUTHOR]

Endocytosis and endolysosomal trafficking are essential for almost all aspects of physiological functions of eukaryotic cells. As our understanding on these membrane trafficking events are mostly from studies in yeast and cultured mammalian cells, one challenge is to systematically evaluate the findings from these cell-based studies in multicellular organisms under physiological settings. One potentially valuable in vivo system to address this challenge is the vitellogenic oocyte in Drosophila, which undergoes extensive endocytosis by Yolkless (Yl), a low-density lipoprotein receptor (LDLR), to uptake extracellular lipoproteins into oocytes and package them into a specialized lysosome, the yolk granule, for storage and usage during later development. However, by now there is still a lack of sufficient understanding on the molecular and cellular processes that control yolk granule biogenesis. Here, by creating genome-tagging lines for Yl receptor and analyzing its distribution in vitellogenic oocytes, we observed a close association of different endosomal structures with distinct phosphoinositides and actin cytoskeleton dynamics. We further showed that Rab5 and Rab11, but surprisingly not Rab4 and Rab7, are essential for yolk granules biogenesis. Instead, we uncovered evidence for a potential role of Rab7 in actin regulation and observed a notable overlap of Rab4 and Rab7, two Rab GTPases that have long been proposed to have distinct spatial distribution and functional roles during endolysosomal trafficking. Through a small-scale RNA interference (RNAi) screen on a set of reported Rab5 effectors, we showed that yolk granule biogenesis largely follows the canonical endolysosomal trafficking and maturation processes. Further, the data suggest that the RAVE/V-ATPase complexes function upstream of or in parallel with Rab7, and are involved in earlier stages of endosomal trafficking events. Together, our study provides s novel insights into endolysosomal pathways and establishes vitellogenic oocyte in Drosophila as an excellent in vivo model for dissecting the highly complex membrane trafficking events in metazoan. Author summary: Endocytosis and endolysosomal trafficking are membrane-based package and delivery systems in eukaryotes for material exchanges intracellularly in-between membrane-enclosed compartments and extracellular with surrounding milieu. Current understanding of these exchange mechanisms are mainly from studies in yeast and cultured mammalian cells, but exactly how they operate in multicellular organisms under physiological conditions remain unclear. Here we focus on vitellogenic oocytes in Drosophila, which uptake large quantity of extracellular lipoproteins by a low-density lipoprotein receptor called Yolkless into the oocyte and package them into large yolk granules, a specialized lysosome, for storage and usage in later development. Using novel fly lines that allows faithful detection and manipulation of endogenous Yolkless receptor and known endosomal regulators, we show that the formation and maturation of yolk granules largely follows the canonical endolysosomal trafficking pathways, including the critical involvement of small GTPases Rab5 and Rab11 as well as distinct phospholipid species and actin networks, although the results also raise questions on the roles of other regulators including Rab4 and Rab7 in granule biogenesis. Together, this study provides novel insights into the highly complex membrane trafficking events in multicellular organisms and supports Drosophila oocyte as a useful in vivo model for such studies in the future. [ABSTRACT FROM AUTHOR]

Single-strand annealing (SSA) is initiated when a double strand break (DSB) occurs between two flanking repeated sequences, resulting in a deletion that leaves a single copy of the repeat. We studied budding yeast strains carrying two 200-bp URA3 sequences separated by 2.6 kb of spacer DNA (phage lambda) in which a site-specific DSB can be created by HO or Cas9 endonucleases. Repeat-mediated deletion requires removal of long 3'-ended single-stranded tails (flaps) by Rad1-Rad10 with the assistance of Msh2-Msh3, Saw1 and Slx4. A natural 3% divergence of unequally spaced heterologies between these repeats (designated F and A) causes a significant reduction in the frequency of SSA repair. This decrease is caused by heteroduplex rejection in which mismatches (MMs) in the annealed intermediate are recognized by the MutS (Msh2 and Msh6) components of the MM repair (MMR) pathway coupled to unwinding of the duplex by the Sgs1-Rmi1-Top3 helicase. MutL homologs, Mlh1-Pms1 (MutL), are not required for rejection but play their expected role in mismatch correction. Remarkably, heteroduplex rejection is very low in strains where the divergent repeats were immediately adjacent (Tailless strains) and the DSB was induced by Cas9. These results suggest that the presence of nonhomologous tails strongly stimulates heteroduplex rejection in SSA. DNA sequencing analysis of SSA products from the FA Tailed strain showed a gradient of correction favoring the sequence opposite each 3' end of the annealed strand. Mismatches located in the center of the repair intermediate were corrected by Msh2-Msh6 mediated mismatch correction, while correction of MMs at the extremity of the SSA intermediate often appears to use a different mechanism, possibly by 3' nonhomologous tail removal that includes part of the homologous sequence. In contrast, in FA Tailless strains there was a uniform repair of the MMs across the repeat. A distinctive pattern of correction was found in the absence of MSH2, in both Tailed and Tailless strains, different from the spectrum seen in a msh3Δ msh6Δ double mutant. Previous work has shown that SSA is Rad51-independent but dependent on the strand annealing activity of Rad52. However Rad52 becomes dispensable in a Tailless construct where the DSB is induced by Cas9 or in transformation of a plasmid where SSA occurs in the absence of nonhomologous tails. Author summary: Deletions between dispersed, divergent, repeated sequences are frequently found among cancer mutations. We have studied deletion formation by single-strand annealing (SSA) between naturally divergent 200 bp repeats in budding yeast. SSA was initiated by creating a site-specific double-strand break (DSB) in the region between the two repeats, either using HO endonuclease or guide RNA-directed Cas9. In this process, the ends are resected 5' to 3', leaving long 3'-ended single-stranded sequences that can anneal. After annealing any 3'-ended nonhomologous tails are clipped off by the Rad1-Rad10 endonuclease. The presence of nonhomologous tails triggers a Msh2-Msh3-dependent and Sgs1-dependent heteroduplex rejection when there are 7 heterologies within the 200-bp repeats being annealed, whereas a "tailless" construct is not rejected. These results suggest that the presence of nonhomologous tails strongly stimulates heteroduplex rejection in SSA. Mismatch repair within the heteroduplex is Msh2-Msh6-dependent; however, a msh2 deletion proved have a spectrum of mismatch repair different from a msh3 msh6 double mutant, implying that Msh2 may have some additional role when the intermediates have nonhomologous 3' tails. "Tailed" repeats yielded a gradient of repair that favored sequences in the "left" repeat, but "Tailless" SSA yielded a uniform correction of all 7 heterologies in the 200-bp region. A cluster of repeats near the left end of the sequence provokes correction of these markers in a Msh6-independent fashion; this correction is not attributable to 3' to 5' exonuclease activity of DNA polymerase δ and may reflect instances where part of the homologous sequence itself is clipped off by Rad1-Rad10. The presence or absence of nonhomologous tails also affected the requirement for the Rad52 strand annealing protein. Both in the chromosomal SSA assay and in a plasmid assay, Rad52 was largely dispensable when there were no nonhomologous tails after strand annealing. [ABSTRACT FROM AUTHOR]

Changes in gene regulation have long been appreciated as a driving force of adaptive evolution, however the relative contributions of cis- and trans-acting changes to gene regulation over short evolutionary timescales remain unclear. Instances of recent, parallel phenotypic evolution provide an opportunity to assess whether parallel patterns are seen at the level of gene expression, and to assess the relative contribution of cis- and trans- changes to gene regulation in the early stages of divergence. Here, we studied gene expression in liver and brown adipose tissue in two wild-derived strains of house mice that independently adapted to cold, northern environments, and we compared them to a strain of house mice from a warm, tropical environment. To investigate gene regulatory evolution, we studied expression in parents and allele-specific expression in F1 hybrids of crosses between warm-adapted and cold-adapted strains. First, we found that the different cold-adapted mice showed both unique and shared changes in expression, but that the proportion of shared changes (i.e. parallelism) was greater than expected by chance. Second, we discovered that expression evolution occurred largely at tissue-specific and cis-regulated genes, and that these genes were over-represented in parallel cases of evolution. Finally, we integrated the expression data with scans for selection in natural populations and found substantial parallelism in the two northern populations for genes under selection. Furthermore, selection outliers were associated with cis-regulated genes more than expected by chance; cis-regulated genes under selection influenced phenotypes such as body size, immune functioning, and activity level. These results demonstrate that parallel patterns of gene expression in mice that have independently adapted to cold environments are driven largely by tissue-specific and cis-regulatory changes, providing insight into the mechanisms of adaptive gene regulatory evolution at the earliest stages of divergence. Author summary: The parallel movement of organisms into novel environments provides an opportunity to understand the molecular basis of adaptation and the repeatability of this process. Mutations affecting the expression of genes are known to underlie much of adaptive evolution. Such mutations can arise in cis- (near the gene of interest) or in trans- (at a distant locus), but the relative contribution of these different kinds of changes to adaptation is poorly understood, especially during very recent divergence. Here, we compared evolved gene expression differences between a warm-adapted house mouse strain and two different cold-adapted strains that have independently evolved similar phenotypic traits, such as increased body size and decreased extremity length during the last few hundred years. Using crosses between warm-adapted and cold-adapted mice, we found that mutations acting in a context specific manner (cis-regulatory and tissue-specific changes) predominate expression divergence and are more likely to be involved in parallel evolution. We used population level selection scans in wild animals to identify regions of the genome under selection and combined these findings with the gene expression data to identify candidate genes underlying adaptation to novel environments. Together, our work describes the gene regulatory dynamics of rapid environmental adaptation, and the repeatability of these patterns over multiple instances of adaptation. [ABSTRACT FROM AUTHOR]

Genome evolution is partly driven by the mobility of transposable elements (TEs) which often leads to deleterious effects, but their activity can also facilitate genetic novelty and catalyze local adaptation. We explored how the intraspecific diversity of TE polymorphisms might contribute to the broad geographic success and adaptive capacity of the emerging oil crop Thlaspi arvense (field pennycress). We classified the TE inventory based on a high-quality genome assembly, estimated the age of retrotransposon TE families and comprehensively assessed their mobilization potential. A survey of 280 accessions from 12 regions across the Northern hemisphere allowed us to quantify over 90,000 TE insertion polymorphisms (TIPs). Their distribution mirrored the genetic differentiation as measured by single nucleotide polymorphisms (SNPs). The number and types of mobile TE families vary substantially across populations, but there are also shared patterns common to all accessions. Ty3/Athila elements are the main drivers of TE diversity in T. arvense populations, while a single Ty1/Alesia lineage might be particularly important for transcriptome divergence. The number of retrotransposon TIPs is associated with variation at genes related to epigenetic regulation, including an apparent knockout mutation in BROMODOMAIN AND ATPase DOMAIN-CONTAINING PROTEIN 1 (BRAT1), while DNA transposons are associated with variation at the HSP19 heat shock protein gene. We propose that the high rate of mobilization activity can be harnessed for targeted gene expression diversification, which may ultimately present a toolbox for the potential use of transposition in breeding and domestication of T. arvense. Author summary: Transposable elements (TEs) are often considered genomic parasites, but they can also generate phenotypic novelty that helps organisms to adapt to new environments. To understand how TEs might contribute to phenotypic diversity and adaptive potential in the emerging oilseed crop Thlaspi arvense (field pennycress), we examined the dynamics of TE variation in a geographically diverse sample of this species. By surveying almost 300 wild accessions from North America and Eurasia we discovered over 90,000 polymorphic TE insertions. We identified not only genetic factors that vary between populations and that are associated with TE mobilization, but also TE families that are most likely to generate genetic diversity of interest to breeders. [ABSTRACT FROM AUTHOR]

Fungal pathogens cause devastating disease in crops. Understanding the evolutionary origin of pathogens is essential to the prediction of future disease emergence and the potential of pathogens to disperse. The fungus Pyrenophora teres f. teres causes net form net blotch (NFNB), an economically significant disease of barley. In this study, we have used 104 P. teres f. teres genomes from four continents to explore the population structure and demographic history of the fungal pathogen. We showed that P. teres f. teres is structured into populations that tend to be geographically restricted to different regions. Using Multiple Sequentially Markovian Coalescent and machine learning approaches we demonstrated that the demographic history of the pathogen correlates with the history of barley, highlighting the importance of human migration and trade in spreading the pathogen. Exploring signatures of natural selection, we identified several population-specific selective sweeps that colocalized with genomic regions enriched in putative virulence genes, and loci previously identified as determinants of virulence specificities by quantitative trait locus analyses. This reflects rapid adaptation to local hosts and environmental conditions of P. teres f. teres as it spread with barley. Our research highlights how human activities can contribute to the spread of pathogens that significantly impact the productivity of field crops. Author summary: Population genetic and genomics studies of several crop pathogens have revealed that human activities, such us domestication, trade, and migration have played a pivotal role in the emergence and spread of plant diseases. In this study, we employed cutting-edge genetic analysis techniques and machine learning tools to shed light on the population structure and historical dispersal patterns of a major fungal pathogen of barley, Pyrenophora teres f. teres. We found that barley domestication during the Neolithic era potentially gave rise to the pathogen which has since co-evolved with barley to become one of the most devastating barley diseases we face today. In addition, we identified a large number of genomic regions evolving under strong positive selection that were specific to different populations. This finding suggests that populations are evolving fast, becoming well adapted to their local host availability and environmental conditions. [ABSTRACT FROM AUTHOR]

Females from many mosquito species feed on blood to acquire nutrients for egg development. The oogenetic cycle has been characterized in the arboviral vector Aedes aegypti, where after a bloodmeal, the lipid transporter lipophorin (Lp) shuttles lipids from the midgut and fat body to the ovaries, and a yolk precursor protein, vitellogenin (Vg), is deposited into the oocyte by receptor-mediated endocytosis. Our understanding of how the roles of these two nutrient transporters are mutually coordinated is however limited in this and other mosquito species. Here, we demonstrate that in the malaria mosquito Anopheles gambiae, Lp and Vg are reciprocally regulated in a timely manner to optimize egg development and ensure fertility. Defective lipid transport via Lp knockdown triggers abortive ovarian follicle development, leading to misregulation of Vg and aberrant yolk granules. Conversely, depletion of Vg causes an upregulation of Lp in the fat body in a manner that appears to be at least partially dependent on target of rapamycin (TOR) signaling, resulting in excess lipid accumulation in the developing follicles. Embryos deposited by Vg-depleted mothers are completely inviable, and are arrested early during development, likely due to severely reduced amino acid levels and protein synthesis. Our findings demonstrate that the mutual regulation of these two nutrient transporters is essential to safeguard fertility by ensuring correct nutrient balance in the developing oocyte, and validate Vg and Lp as two potential candidates for mosquito control. Author summary: Female mosquitoes bite humans to acquire nutrients for egg development. The shuttling of nutrients from the gut (where the blood is digested) to the ovaries (where eggs are produced) relies on several nutrient transporters to move through the mosquito circulation, including Lipophorin (Lp), a fat transporter expressed early and Vitellogenin (Vg), a large protein expressed later that transports amino acids. We know relatively little of how these two nutrient transporters are successfully coordinated in any mosquito species, so we undertook to examine their interplay in the Anopheles malaria mosquito. We found that without Lp expression, Vg is incorrectly distributed within ovarian follicles and egg production is aborted, whereas without Vg, fat transport by Lp is not switched off in a timely manner. This results in excess fat and minimal protein deposition in eggs, rendering females completely infertile. We also find evidence that the mutual regulation of these transporters may be mediated by TOR signaling. As well as providing further insight into the regulation of essential reproductive processes, these results may aid in the development of malaria control strategies that aim to reduce the size of mosquito populations. [ABSTRACT FROM AUTHOR]

In the presence of recombination, the evolutionary relationships between a set of sampled genomes cannot be described by a single genealogical tree. Instead, the genomes are related by a complex, interwoven collection of genealogies formalized in a structure called an ancestral recombination graph (ARG). An ARG extensively encodes the ancestry of the genome(s) and thus is replete with valuable information for addressing diverse questions in evolutionary biology. Despite its potential utility, technological and methodological limitations, along with a lack of approachable literature, have severely restricted awareness and application of ARGs in evolution research. Excitingly, recent progress in ARG reconstruction and simulation have made ARG-based approaches feasible for many questions and systems. In this review, we provide an accessible introduction and exploration of ARGs, survey recent methodological breakthroughs, and describe the potential for ARGs to further existing goals and open avenues of inquiry that were previously inaccessible in evolutionary genomics. Through this discussion, we aim to more widely disseminate the promise of ARGs in evolutionary genomics and encourage the broader development and adoption of ARG-based inference. [ABSTRACT FROM AUTHOR]