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Down syndrome cell adhesion molecule ( Dscam ) gene encodes a cell adhesion molecule required for neuronal wiring. A remarkable feature of arthropod Dscam is massive alternative splicing generating thousands of different isoforms from three variable clusters of alternative exons. Dscam expression and diversity arising from alternative splicing have been studied during development, but whether they exert functions in adult brains has not been determined. Here, using honey bees, we find that Dscam expression is critically linked to memory retention as reducing expression by RNAi enhances memory after reward learning in adult worker honey bees. Moreover, alternative splicing of Dscam is altered in all three variable clusters after learning. Since identical Dscam isoforms engage in homophilic interactions, these results suggest a mechanism to alter inclusion of variable exons during memory consolidation to modify neuronal connections for memory retention., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Ustaoglu, McQuarrie, Rochet, Dix, Haussmann, Arnold, Devaud and Soller.)

Nuclear pore proteins (Nups) prominently are among the few genes linked to speciation from hybrid incompatibility in Drosophila. These studies have focused on coding sequence evolution of Nup96 and Nup160 and shown evidence of positive selection driving nucleoporin evolution. Intriguingly, channel Nup54 functionality is required for neuronal wiring underlying the female post-mating response induced by male-derived sex-peptide. A region of rapid evolution in the core promoter of Nup54 suggests a critical role for general transcriptional regulatory elements at the onset of speciation, but whether this is a general feature of Nup genes has not been determined. Consistent with findings for Nup54, additional channel Nup58 and Nup62 promoters also rapidly accumulate insertions/deletions (indels). Comprehensive examination of Nup upstream regions reveals that core Nup complex gene promoters accumulate indels rapidly. Since changes in promoters can drive changes in expression, these results indicate an evolutionary mechanism driven by indel accumulation in core Nup promoters. Compensation of such gene expression changes could lead to altered neuronal wiring, rapid fixation of traits caused by promoter changes and subsequently the rise of new species. Hence, the nuclear pore complex may act as a nexus for species-specific changes via nucleo-cytoplasmic transport regulated gene expression., (© 2023. The Author(s).)

Animal, protist and viral messenger RNAs (mRNAs) are most prominently modified at the beginning by methylation of cap-adjacent nucleotides at the 2'-O-position of the ribose (cOMe) by dedicated cap methyltransferases (CMTrs). If the first nucleotide of an mRNA is an adenosine, PCIF1 can methylate at the N 6 -position (m 6 A), while internally the Mettl3/14 writer complex can methylate. These modifications are introduced co-transcriptionally to affect many aspects of gene expression including localisation to synapses and local translation. Of particular interest, transcription start sites of many genes are heterogeneous leading to sequence diversity at the beginning of mRNAs, which together with cOMe and m 6 Am could constitute an extensive novel layer of gene expression control. Given the role of cOMe and m 6 A in local gene expression at synapses and higher brain functions including learning and memory, such code could be implemented at the transcriptional level for lasting memories through local gene expression at synapses., (© 2022 The Authors. BioEssays published by Wiley Periodicals LLC.)

Marek's Disease (MD) has a significant impact on both the global poultry economy and animal welfare. The disease pathology can include neurological damage and tumour formation. Sexual dimorphism in immunity and known higher susceptibility of females to MD makes the chicken Z chromosome (GGZ) a particularly attractive target to study the chicken MD response. Previously, we used a Hy-Line F 6 population from a full-sib advanced intercross line to map MD QTL regions (QTLRs) on all chicken autosomes. Here, we mapped MD QTLRs on GGZ in the previously utilized F 6 population with individual genotypes and phenotypes, and in eight elite commercial egg production lines with daughter-tested sires and selective DNA pooling (SDP). Four MD QTLRs were found from each analysis. Some of these QTLRs overlap regions from previous reports. All QTLRs were tested by individuals from the same eight lines used in the SDP and genotyped with markers located within and around the QTLRs. All QTLRs were confirmed. The results exemplify the complexity of MD resistance in chickens and the complex distribution of p -values and Linkage Disequilibrium (LD) pattern and their effect on localization of the causative elements. Considering the fragments and interdigitated LD blocks while using LD to aid localization of causative elements, one must look beyond the non-significant markers, for possible distant markers and blocks in high LD with the significant block. The QTLRs found here may explain at least part of the gender differences in MD tolerance, and provide targets for mitigating the effects of MD.

Cap methyltransferases (CMTrs) O methylate the 2' position of the ribose (cOMe) of cap-adjacent nucleotides of animal, protist, and viral mRNAs. Animals generally have two CMTrs, whereas trypanosomes have three, and many viruses encode one in their genome. In the splice leader of mRNAs in trypanosomes, the first four nucleotides contain cOMe, but little is known about the status of cOMe in animals. Here, we show that cOMe is prominently present on the first two cap-adjacent nucleotides with species- and tissue-specific variations in Caenorhabditis elegans , honeybees, zebrafish, mouse, and human cell lines. In contrast, Drosophila contains cOMe primarily on the first cap-adjacent nucleotide. De novo RoseTTA modeling of CMTrs reveals close similarities of the overall structure and near identity for the catalytic tetrad, and for cap and cofactor binding for human, Drosophila and C. elegans CMTrs. Although viral CMTrs maintain the overall structure and catalytic tetrad, they have diverged in cap and cofactor binding. Consistent with the structural similarity, both CMTrs from Drosophila and humans methylate the first cap-adjacent nucleotide of an AGU consensus start. Because the second nucleotide is also methylated upon heat stress in Drosophila , these findings argue for regulated cOMe important for gene expression regulation., (© 2022 Dix et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)

Cap-adjacent nucleotides of animal, protist and viral mRNAs can be O-methylated at the 2' position of the ribose (cOMe). The functions of cOMe in animals, however, remain largely unknown. Here we show that the two cap methyltransferases (CMTr1 and CMTr2) of Drosophila can methylate the ribose of the first nucleotide in mRNA. Double-mutant flies lack cOMe but are viable. Consistent with prominent neuronal expression, they have a reward learning defect that can be rescued by conditional expression in mushroom body neurons before training. Among CMTr targets are cell adhesion and signaling molecules. Many are relevant for learning, and are also targets of Fragile X Mental Retardation Protein (FMRP). Like FMRP, cOMe is required for localization of untranslated mRNAs to synapses and enhances binding of the cap binding complex in the nucleus. Hence, our study reveals a mechanism to co-transcriptionally prime mRNAs by cOMe for localized protein synthesis at synapses., (© 2022. The Author(s).)

Alternative splicing increases neuronal transcriptomic complexity throughout animal phylogeny. To delve into the mechanisms controlling the assembly and evolution of this regulatory layer, we characterized the neuronal microexon program in Drosophila and compared it with that of mammals. In nonvertebrate bilaterians, this splicing program is restricted to neurons by the posttranscriptional processing of the enhancer of microexons (eMIC) domain in Srrm234 . In Drosophila , this processing is dependent on regulation by Elav/Fne. eMIC deficiency or misexpression leads to widespread neurological alterations largely emerging from impaired neuronal activity, as revealed by a combination of neuronal imaging experiments and cell type-specific rescues. These defects are associated with the genome-wide skipping of short neural exons, which are strongly enriched in ion channels. We found no overlap of eMIC-regulated exons between flies and mice, illustrating how ancient posttranscriptional programs can evolve independently in different phyla to affect distinct cellular modules while maintaining cell-type specificity.

Changes in gene expression are a hallmark of learning and memory consolidation. Little is known about how alternative mRNA processing, particularly abundant in neuron-specific genes, contributes to these processes. Prototype RNA binding proteins of the neuronally expressed ELAV/Hu family are candidates for roles in learning and memory, but their capacity to cross-regulate and take over each other's functions complicate substantiation of such links. Honey bees Apis mellifera have only one elav/Hu family gene elavl2, that has functionally diversified by increasing alternative splicing including an evolutionary conserved microexon. RNAi knockdown demonstrates that ELAVL2 is required for learning and memory in bees. ELAVL2 is dynamically expressed with altered alternative splicing and subcellular localization in mushroom bodies, but not in other brain regions. Expression and alternative splicing of elavl2 change during memory consolidation illustrating an alternative mRNA processing program as part of a local gene expression response underlying memory consolidation., (© 2021. The Author(s).)

Background: Female reproductive behaviors and physiology change profoundly after mating. The control of pregnancy-associated changes in physiology and behaviors are largely hard-wired into the brain to guarantee reproductive success, yet the gene expression programs that direct neuronal differentiation and circuit wiring at the end of the sex determination pathway in response to mating are largely unknown. In Drosophila, the post-mating response induced by male-derived sex-peptide in females is a well-established model to elucidate how complex innate behaviors are hard-wired into the brain. Here, we use a genetic approach to further characterize the molecular and cellular architecture of the sex-peptide response in Drosophila females., Results: Screening for mutations that affect the sensitivity to sex-peptide, we identified the channel nuclear pore protein Nup54 gene as an essential component for mediating the sex-peptide response, with viable mutant alleles leading to the inability of laying eggs and reducing receptivity upon sex-peptide exposure. Nup54 directs correct wiring of eight adult brain neurons that express pickpocket and are required for egg-laying, while additional channel Nups also mediate sexual differentiation. Consistent with links of Nups to speciation, the Nup54 promoter is a hot spot for rapid evolution and promoter variants alter nucleo-cytoplasmic shuttling., Conclusions: These results implicate nuclear pore functionality to neuronal wiring underlying the sex-peptide response and sexual differentiation as a response to sexual conflict arising from male-derived sex-peptide to direct the female post-mating response., (© 2021. The Author(s).)

N 6 -methyladenosine (m 6 A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m 6 A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila and human cells. In Drosophila, its depletion results in reduced m 6 A levels and altered m 6 A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerization and interaction with other members of the m 6 A machinery, while its catalytic activity is dispensable. Finally, we demonstrate that the loss of Hakai destabilizes several subunits of the methyltransferase complex, resulting in impaired m 6 A deposition. Our work adds functional and molecular insights into the mechanism of the m 6 A mRNA writer complex.

Background: In the face of significant distress among physicians, access to counseling is critical., Objective: An on-site wellness program for physicians-in-training and faculty was assessed by examining (a) were participants representative of those eligible for services and (b) demographic and trainee vs. faculty differences in burnout, distress, suicide risk, and presenting concerns of participants who utilized services., Methods: From 2013-2018, 73% (N = 468; 316 residents/fellows, 152 faculty) of individuals seeking services also consented to research. At intake, participants completed a distress measure (ACORN) and two items from the Maslach Burnout Inventory (MBI), and clinicians categorized presenting concerns and suicide risk. Using Chi-square analyses, participants' characteristics were compared to physicians eligible for treatment. The association between demographics, faculty vs. trainee status, specialty, and distress, burnout, suicide risk, and presenting concerns was evaluated with ANOVAs and logistic regressions., Results: Women, trainees, and primary care physicians were more likely to access services. On the ACORN, 63% were in the clinical range (M =1.7, SD =0.6). On the MBI, 36% scored in the clinical range. Clinicians rated 9% of participants with suicide risk. Neither gender, racial/ethnic minority status, nor specialty were associated with distress, burnout or suicide risk. Trainees reported greater distress than faculty ( F (1,447) = 8.42, P  =   .004, η p 2  = .018). Participants reported multiple presenting concerns (M = 3.0, SD = 1.18) with faculty more commonly endorsing work-related issues. Trainees more commonly reported new or worsening psychological symptoms, performance and family concerns., Conclusions: Two physician groups which often report higher levels of burnout and distress when surveyed, women physicians and residents/fellows, were the most likely to get professional help in an on-site wellness program. Physician wellness programs need to be prepared to address work and personal stressors and different levels of distress and risk., Competing Interests: Declaration of Conflicting Interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s) 2021.)

The nuclear pore complex (NPC) is the principal gateway between nucleus and cytoplasm that enables exchange of macromolecular cargo. Composed of multiple copies of ~30 different nucleoporins (Nups), the NPC acts as a selective portal, interacting with factors which individually license passage of specific cargo classes. Here we show that two Nups of the inner channel, Nup54 and Nup58, are essential for transposon silencing via the PIWI-interacting RNA (piRNA) pathway in the Drosophila ovary. In ovarian follicle cells, loss of Nup54 and Nup58 results in compromised piRNA biogenesis exclusively from the flamenco locus, whereas knockdowns of other NPC subunits have widespread consequences. This provides evidence that some Nups can acquire specialised roles in tissue-specific contexts. Our findings consolidate the idea that the NPC has functions beyond simply constituting a barrier to nuclear/cytoplasmic exchange as genomic loci subjected to strong selective pressure can exploit NPC subunits to facilitate their expression., Competing Interests: MM, VL, AP, SM, SB, IH, MS, GH, BC No competing interests declared, (© 2021, Munafò et al.)

Maximizing crop yields relies on the use of agrochemicals to control insect pests. One of the most widely used classes of insecticides are neonicotinoids that interfere with signalling of the neurotransmitter acetylcholine, but these can also disrupt crop-pollination services provided by bees. Here, we analysed whether chronic low dose long-term exposure to the neonicotinoid thiamethoxam alters gene expression and alternative splicing in brains of Africanized honey bees, Apis mellifera, as adaptation to altered neuronal signalling. We find differentially regulated genes that show concentration-dependent responses to thiamethoxam, but no changes in alternative splicing. Most differentially expressed genes have no annotated function but encode short Open Reading Frames, a characteristic feature of anti-microbial peptides. As this suggested that immune responses may be compromised by thiamethoxam exposure, we tested the impact of thiamethoxam on bee immunity by injecting bacteria. We show that intrinsically sub-lethal thiamethoxam exposure makes bees more vulnerable to normally non-pathogenic bacteria. Our findings imply a synergistic mechanism for the observed bee population declines that concern agriculturists, conservation ecologists and the public.

Modified nucleotides in mRNA are an essential addition to the standard genetic code of four nucleotides in animals, plants, and their viruses. The emerging field of epitranscriptomics examines nucleotide modifications in mRNA and their impact on gene expression. The low abundance of nucleotide modifications and technical limitations, however, have hampered systematic analysis of their occurrence and functions. Selective chemical and immunological identification of modified nucleotides has revealed global candidate topology maps for many modifications in mRNA, but further technical advances to increase confidence will be necessary. Single-molecule sequencing introduced by Oxford Nanopore now promises to overcome such limitations, and we summarize current progress with a particular focus on the bioinformatic challenges of this novel sequencing technology., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Accounts of frontline health care workers experiencing distress in the midst of the COVID-19 pandemic highlight the need for accessible psychological support for them. Prior to the pandemic, medical residents and physicians often experienced difficulty receiving counseling due to concerns about confidentiality, stigma, cost, time, and reportability to licensure/credentialing bodies. Since 2004, the OHSU Resident and Faculty Wellness Program (RFWP) team has sought to reduce these barriers by providing on-site free, confidential, individual counseling and medication management. Utilization of this program is high with over 500 physicians a year seeking care; 38% of all OHSU residents/fellows and 7% of all faculty eligible for our services participated in 2019-20. In the present essay, we describe how our model of care for trainees and faculty was a key wellness resource during COVID-19. Similar to other accounts of lower help-seeking by health professionals initially during the pandemic, we experienced a slight downturn in utilization rates during the initial weeks of when the pandemic struck our area, but quickly returned to normal and exceeded prior levels. All appointments shifted to telehealth visits and a number of physicians expressed gratitude for the opportunity to talk through concerns and strengthen coping. A number of physicians requested medication consultations to address severe insomnia, anxiety, and depression. We hope that being present in our physicians' lives when they are exposed to COVID-related stress or trauma will keep them safe, help them cope with difficult experiences and losses, and ultimately facilitate both recovery and post-traumatic growth., Competing Interests: Declaration of Conflicting Interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s) 2020.)

The tissue-specific deployment of highly extended neural 3' UTR isoforms, generated by alternative polyadenylation (APA), is a broad and conserved feature of metazoan genomes. However, the factors and mechanisms that control neural APA isoforms are not well understood. Here, we show that three ELAV/Hu RNA binding proteins (Elav, Rbp9, and Fne) have similar capacities to induce a lengthened 3' UTR landscape in an ectopic setting. These factors promote accumulation of chromatin-associated, 3' UTR-extended, nascent transcripts, through inhibition of proximal polyadenylation site (PAS) usage. Notably, Elav represses an unannotated splice isoform of fne, switching the normally cytoplasmic Fne toward the nucleus in elav mutants. We use genomic profiling to reveal strong and broad loss of neural APA in elav/fne double mutant CNS, the first genetic background to largely abrogate this distinct APA signature. Overall, we demonstrate how regulatory interplay and functionally overlapping activities of neural ELAV/Hu RBPs drives the neural APA landscape., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Marek's disease (MD) represents a significant global economic and animal welfare issue. Marek's disease virus (MDV) is a highly contagious oncogenic and highly immune-suppressive α-herpes virus, which infects chickens, causing neurological effects and tumour formation. Though partially controlled by vaccination, MD continues to have a profound impact on animal health and on the poultry industry. Genetic selection provides an alternative and complementary method to vaccination. However, even after years of study, the genetic mechanisms underlying resistance to MDV remain poorly understood. The Major Histocompatability Complex (MHC) is known to play a role in disease resistance, along with a handful of other non-MHC genes. In this study, one of the largest to date, we used a multi-facetted approach to identify QTL regions (QTLR) influencing resistance to MDV, including an F 6 population from a full-sib advanced intercross line (FSIL) between two elite commercial layer lines differing in resistance to MDV, RNA-seq information from virus challenged chicks, and genome wide association study (GWAS) from multiple commercial lines. Candidate genomic elements residing in the QTLR were further tested for association with offspring mortality in the face of MDV challenge in eight pure lines of elite egg-layer birds. Thirty-eight QTLR were found on 19 chicken chromosomes. Candidate genes, miRNAs, lncRNAs and potentially functional mutations were identified in these regions. Association tests were carried out in 26 of the QTLR, using eight pure lines of elite egg-layer birds. Numerous candidate genomic elements were strongly associated with MD resistance. Genomic regions significantly associated with resistance to MDV were mapped and candidate genes identified. Various QTLR elements were shown to have a strong genetic association with resistance. These results provide a large number of significant targets for mitigating the effects of MDV infection on both poultry health and the economy, whether by means of selective breeding, improved vaccine design, or gene-editing technologies.

Backgrounds and Purpose- Although new methods for genetic analyses are rapidly evolving, there are currently knowledge gaps in how to detect Mendelian forms of stroke. Methods- We performed whole-exome sequencing in 22 probands, under 56 years at their first ischemic stroke episode, from multi-incident stroke families. With the use of a comprehensive stroke-gene panel, we searched for variants in stroke-related genes. The probands' clinical stroke subtype was related to clinical characteristics previously associated with pathogenic variants in these genes. Relatives were genotyped in 7 families to evaluate stroke-gene variants of unknown significance. In 2 larger families with embolic stroke of unknown source, whole-exome sequencing was performed in additional members to examine the possibility of identifying new stroke genes. Results- Six of 22 probands carried pathogenic or possibly pathogenic variants in genes reported to be associated with their stroke subtype. A known pathogenic variant in NOTCH3 and a possibly pathogenic variant in ACAD9 gene were identified. A novel JAK2 :c.3188G>A (p.Arg1063His) mutation was seen in a proband with embolic stroke of undetermined source and prothrombotic status. However, penetrance in the family was incomplete. COL4A2: c.3368A>G (p.Glu1123Gly) was detected in 2 probands but did not cosegregate with the disease in their families. Whole-exome sequencing in multiple members of 2 pedigrees with embolic stroke of undetermined source revealed possibly pathogenic variants in genes not previously associated with stroke, GPR142: c.148C>G (p.Leu50Val), and PTPRN2 :c.2416A>G (p.Ile806Val); LRRC1 c.808A>G (p.Ile270Val), SLC7A10 c.1294dupG (p.Val432fs), IKBKB : c.1070C>T (p.Ala357Val), and OXGR1 c.392G>A (p.Arg131His), respectively. Conclusions- Screening with whole-exome sequencing using a comprehensive stroke-gene panel may identify rare monogenic forms of stroke, but careful evaluation of clinical characteristics and potential pathogenicity of novel variants remain important. In our study, the majority of individuals with familial aggregation of stroke lacked any identified genetic causes.

The distribution of assembled, and potentially translating, ribosomes within cells can be visualised in Drosophila by using Bimolecular Fluorescence Complementation (BiFC) to monitor the interaction between tagged pairs of 40S and 60S ribosomal proteins (RPs) that are close neighbours across inter-subunit junctions in the assembled 80S ribosome. Here we describe transgenes expressing two novel RP pairs tagged with Venus-based BiFC fragments that considerably increase the sensitivity of this technique we termed Ribo-BiFC. This improved method should provide a convenient way of monitoring the local distribution of ribosomes in most Drosophila cells and we suggest that it could be implemented in other organisms. We visualised 80S ribosomes in different neurons, particularly photoreceptors in the larva, pupa and adult brain. Assembled ribosomes are most abundant in the various neuronal cell bodies, but they are also present along the full length of axons. They are concentrated in growth cones of developing photoreceptors and are apparent at the terminals of mature larval photoreceptors targeting the larval optical neuropil. Surprisingly, there is relatively less puromycin incorporation in the distal portion of axons in the larval optic stalk, suggesting that some of the ribosomes that have initiated translation may not be engaged in elongation in growing axons.This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

Securing food supply for a growing population is a major challenge and heavily relies on the use of agrochemicals to maximize crop yield. It is increasingly recognized, that some neonicotinoid insecticides have a negative impact on non-target organisms, including important pollinators such as the European honeybee Apis mellifera. Toxicity of neonicotinoids may be enhanced through simultaneous exposure with additional pesticides, which could help explain, in part, the global decline of honeybee colonies. Here we examined whether exposure effects of the neonicotinoid thiamethoxam on bee viability are enhanced by the commonly used fungicide carbendazim and the herbicide glyphosate. We also analysed alternative splicing changes upon pesticide exposure in the honeybee. In particular, we examined transcripts of three genes: (i) the stress sensor gene X box binding protein-1 (Xbp1), (ii) the Down Syndrome Cell Adhesion Molecule (Dscam) gene and iii) the embryonic lethal/abnormal visual system (elav) gene, which are important for neuronal function. Our results showed that acute thiamethoxam exposure is not enhanced by carbendazim, nor glyphosate. Toxicity of the compounds did not trigger stress-induced, alternative splicing in the analysed mRNAs, thereby leaving dormant a cellular response pathway to these man-made environmental perturbations.

The Collaborative Cross (CC) mouse resource is a next-generation mouse genetic reference population (GRP) designed for high-resolution mapping of quantitative trait loci (QTL) of large effect affecting complex traits during health and disease. The CC resource consists of a set of 72 recombinant inbred lines (RILs) generated by reciprocal crossing of five classical and three wild-derived mouse founder strains. Complex traits are controlled by variations within multiple genes and environmental factors, and their mutual interactions. These traits are observed at multiple levels of the animals' systems, including metabolism, body weight, immune profile, and susceptibility or resistance to the development and progress of infectious or chronic diseases. Herein, we present general guidelines for design of QTL mapping experiments using the CC resource-along with full step-by-step protocols and methods that were implemented in our lab for the phenotypic and genotypic characterization of the different CC lines-for mapping the genes underlying host response to infectious and chronic diseases. © 2019 by John Wiley & Sons, Inc. Basic Protocol 1: CC lines for whole body mass index (BMI) Basic Protocol 2: A detailed assessment of the power to detect effect sizes based on the number of lines used, and the number of replicates per line Basic Protocol 3: Obtaining power for QTL with given target effect by interpolating in Table 1 of Keele et al. (2019)., (© 2019 John Wiley & Sons, Inc.)

Ethanol tolerance, a polygenic trait of the yeast Saccharomyces cerevisiae , is the primary factor determining industrial bioethanol productivity. Until now, genomic elements affecting ethanol tolerance have been mapped only at low resolution, hindering their identification. Here, we explore the genetic architecture of ethanol tolerance, in the F6 generation of an Advanced Intercrossed Line (AIL) mapping population between two phylogenetically distinct, but phenotypically similar, S. cerevisiae strains (a common laboratory strain and a wild strain isolated from nature). Under ethanol stress, 51 quantitative trait loci (QTLs) affecting growth and 96 QTLs affecting survival, most of them novel, were identified, with high resolution, in some cases to single genes, using a High-Resolution Mapping Package of methodologies that provided high power and high resolution. We confirmed our results experimentally by showing the effects of the novel mapped genes: MOG1 , MGS1 , and YJR154W. The mapped QTLs explained 34% of phenotypic variation for growth and 72% for survival. High statistical power provided by our analysis allowed detection of many loci with small, but mappable effects, uncovering a novel "quasi-infinitesimal" genetic architecture. These results are striking demonstration of tremendous amounts of hidden genetic variation exposed in crosses between phylogenetically separated strains with similar phenotypes; as opposed to the more common design where strains with distinct phenotypes are crossed. Our findings suggest that ethanol tolerance is under natural evolutionary fitness-selection for an optimum phenotype that would tend to eliminate alleles of large effect. The study provides a platform for development of superior ethanol-tolerant strains using genome editing or selection., (Copyright © 2019 Haas, Horev, Lipkin, Kesten, Portnoy, Buhnik-Rosenblau, Soller and Kashi.)

Alternative splicing of pre-mRNA is a major mechanism to diversify protein functionality in metazoans from a limited number of genes. The Drosophila melanogaster Down syndrome cell adhesion molecule ( Dscam ) gene, which is important for neuronal wiring and phagocytosis of bacteria, can generate up to 38,016 isoforms by mutually exclusive alternative splicing in four clusters of variable exons. However, it is not understood how a specific exon is chosen from the many variables and how variable exons are prevented from being spliced together. A main role in the regulation of Dscam alternative splicing has been attributed to RNA binding proteins (RBPs), but how they impact on exon selection is not well understood. Serine-arginine rich (SR) proteins and hnRNP proteins are the two main types of RBPs with major roles in exon definition and splice site selection. Here, we analyzed the role of SR and hnRNP proteins in Dscam exon 9 alternative splicing in mutant Drosophila melanogaster embryos because of their essential function for development. Strikingly, loss or overexpression of canonical SR and hnRNP proteins even when multiple proteins are depleted together, does not affect Dscam alternative exon selection very dramatically. Conversely, noncanonical SR protein Serine-arginine repetitive matrix 2/3/4 (Srrm234) is a main determinant of exon inclusion in the Dscam exon 9 cluster. Since long-range base-pairings are absent in the exon 9 cluster, our data argue for a small complement of regulatory factors as main determinants of exon inclusion in the Dscam exon 9 cluster., (© 2019 Ustaoglu et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)

Objective: To determine the frequency of mutations known to cause autosomal dominant Parkinson disease (PD) in a series with more than 10% of Sweden's estimated number of PD patients., Methods: The Swedish Parkinson Disease Genetics Network was formed as a national multicenter consortium of clinical researchers who together have access to DNA from a total of 2,206 PD patients; 85.4% were from population-based studies. Samples were analyzed centrally for known pathogenic mutations in SNCA (duplications/triplications, p.Ala30Pro, p.Ala53Thr) and LRRK2 (p.Asn1437His, p.Arg1441His, p.Tyr1699Cys, p.Gly2019Ser, p.Ile2020Thr). We compared the frequency of these mutations in Swedish patients with published PD series and the gnomAD database., Results: A family history of PD in first- and/or second-degree relatives was reported by 21.6% of participants. Twelve patients (0.54%) carried LRRK2 p.(Gly2019Ser) mutations, one patient (0.045%) an SNCA duplication. The frequency of LRRK2 p.(Gly2019Ser) carriers was 0.11% in a matched Swedish control cohort and a similar 0.098% in total gnomAD, but there was a marked difference between ethnicities in gnomAD, with 42-fold higher frequency among Ashkenazi Jews than all others combined., Conclusions: In relative terms, the LRRK2 p.(Gly2019Ser) variant is the most frequent mutation among Swedish or international PD patients, and in gnomAD. SNCA duplications were the second most common of the mutations examined. In absolute terms, however, these known pathogenic variants in dominant PD genes are generally very rare and can only explain a minute fraction of familial aggregation of PD. Additional genetic and environmental mechanisms may explain the frequent co-occurrence of PD in close relatives., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The mechanisms by which entire programmes of gene regulation emerged during evolution are poorly understood. Neuronal microexons represent the most conserved class of alternative splicing in vertebrates, and are critical for proper brain development and function. Here, we discover neural microexon programmes in non-vertebrate species and trace their origin to bilaterian ancestors through the emergence of a previously uncharacterized 'enhancer of microexons' (eMIC) protein domain. The eMIC domain originated as an alternative, neural-enriched splice isoform of the pan-eukaryotic Srrm2/SRm300 splicing factor gene, and subsequently became fixed in the vertebrate and neuronal-specific splicing regulator Srrm4/nSR100 and its paralogue Srrm3. Remarkably, the eMIC domain is necessary and sufficient for microexon splicing, and functions by interacting with the earliest components required for exon recognition. The emergence of a novel domain with restricted expression in the nervous system thus resulted in the evolution of splicing programmes that qualitatively expanded the neuronal molecular complexity in bilaterians.

Alternative splicing is a key feature of human genes, yet studying its regulation is often complicated by large introns. The Down Syndrome Cell Adhesion Molecule (Dscam) gene from Drosophila is one of the most complex genes generating vast molecular diversity by mutually exclusive alternative splicing. To resolve how alternative splicing in Dscam is regulated, we first developed plasmid-based UAS reporter genes for the Dscam variable exon 4 cluster and show that its alternative splicing is recapitulated by GAL4-mediated expression in neurons. We then developed gap-repair recombineering to very efficiently manipulate these large reporter plasmids in Escherichia coli using restriction enzymes or sgRNA/Cas9 DNA scission to capitalize on the many benefits of plasmids in phiC31 integrase-mediated transgenesis. Using these novel tools, we show that inclusion of Dscam exon 4 variables differs little in development and individual flies, and is robustly determined by sequences harbored in variable exons. We further show that introns drive selection of both proximal and distal variable exons. Since exon 4 cluster introns lack conserved sequences that could mediate robust long-range base-pairing to bring exons into proximity for splicing, our data argue for a central role of introns in mutually exclusive alternative splicing of Dscam exon 4 cluster., (© The Author(s) 2018. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The central dogma of molecular biology introduced by Crick describes a linear flow of information from DNA to mRNA to protein. Since then it has become evident that RNA undergoes several maturation steps such as capping, splicing, 3'-end processing, and editing. Likewise, nucleotide modifications are common in mRNA and are present in all organisms impacting on the regulation of gene expression. The most abundant modification found in mRNA is N6-methyladenosine (m 6 A). Deposition of m 6 A is a nuclear process and is performed by a megadalton writer complex primarily on mRNAs, but also on microRNAs and lncRNAs. The m 6 A methylosome is composed of the enzymatic core components METTL3 and METTL14, and several auxiliary proteins necessary for its correct positioning and functioning, which are WTAP, VIRMA, FLACC, RBM15, and HAKAI. The m 6 A epimark is decoded by YTH domain-containing reader proteins YTHDC and YTHDF, but METTLs can act as "readers" as well. Eraser proteins, such as FTO and ALKBH5, can remove the methyl group. Here we review recent progress on the role of m 6 A in regulating gene expression in light of Crick's central dogma of molecular biology. In particular, we address the complexity of the writer complex from an evolutionary perspective to obtain insights into the mechanism of ancient m 6 A methylation and its regulation.

Extensive analyses of known monogenic causes of stroke by whole-exome/genome sequencing are technically possible today. We here aimed to compile a comprehensive panel of genes associated with monogenic causes of stroke for use in clinical and research situations. We systematically searched the publically available database Online Mendelian Inheritance in Man, and validated the entries against original peer-reviewed publications in PubMed. First, we selected known pathogenic or putatively pathogenic stroke genes reported in at least one person with stroke, and classified the stroke phenotype for each gene into eight subgroups: (1) large artery atherosclerotic, (2) large artery non-atherosclerotic (tortuosity, dolichoectasia, aneurysm, non-atherosclerotic dissection, occlusion), (3) cerebral small-vessel diseases, (4) cardioembolic (arrhythmia, heart defect, cardiomyopathy), (5) coagulation dysfunctions (venous thrombosis, arterial thrombosis, bleeding tendency), (6) intracerebral hemorrhage, (7) vascular malformations (cavernoma, arteriovenous malformations), and (8) metabolism disorders. Second, we selected other genes that may plausibly cause stroke through diseases related to stroke, but without any documented stroke patient description. A third section comprised SNPs associated with stroke in genome-wide association studies (GWAS). We identified in total 214 genes: 120 associated with stroke, 62 associated with diseases that may cause stroke, and 32 stroke-related genes from recent GWAS. We describe these 214 genes and the clinical stroke subtype(s) associated with each of them. The resulting gene panel can be used to interpret exome sequencing results regarding monogenic stroke. Based on the panel's clinical phenotype description, the pathogenicity of novel variants in these genes may be evaluated in specific situations.

Accumulation and aggregation of TDP-43 is a major pathological hallmark of amyotrophic lateral sclerosis and frontotemporal dementia. TDP-43 inclusions also characterize patients with GGGGCC (G4C2) hexanucleotide repeat expansion in C9orf72 that causes the most common genetic form of amyotrophic lateral sclerosis and frontotemporal dementia (C9ALS/FTD). Functional studies in cell and animal models have identified pathogenic mechanisms including repeat-induced RNA toxicity and accumulation of G4C2-derived dipeptide-repeat proteins. The role of TDP-43 dysfunction in C9ALS/FTD, however, remains elusive. We found G4C2-derived dipeptide-repeat protein but not G4C2-RNA accumulation caused TDP-43 proteinopathy that triggered onset and progression of disease in Drosophila models of C9ALS/FTD. Timing and extent of TDP-43 dysfunction was dependent on levels and identity of dipeptide-repeat proteins produced, with poly-GR causing early and poly-GA/poly-GP causing late onset of disease. Accumulating cytosolic, but not insoluble aggregated TDP-43 caused karyopherin-α2/4 (KPNA2/4) pathology, increased levels of dipeptide-repeat proteins and enhanced G4C2-related toxicity. Comparable KPNA4 pathology was observed in both sporadic frontotemporal dementia and C9ALS/FTD patient brains characterized by its nuclear depletion and cytosolic accumulation, irrespective of TDP-43 or dipeptide-repeat protein aggregates. These findings identify a vicious feedback cycle for dipeptide-repeat protein-mediated TDP-43 and subsequent KPNA pathology, which becomes self-sufficient of the initiating trigger and causes C9-related neurodegeneration.

In the mid-1960s egg production, fertility, and hatchability of broiler breeder females dropped precipitously. Due to disrupted follicle hierarchies and development of the erratic oviposition and defective eggs (EODES) syndrome. EODES was controlled by restricting feed. In the 1990s, another set of problems arose at entry of broiler breeders into lay and characterized by high mortality followed by lower peak lay and reduction in egg and chick production. These problems are induced by even slight over-feeding, and hence we termed it the "Over Feeding Complex" (OFC). We have speculated that OFC is a quasi-EODES condition, induced by the intense selection for increased breast proportion. To test this, we compared, under fast (FF) and slow (SF) release from feed restriction, body composition and reproductive performance of a broiler breeder from year 1980 (B1980) and kept without selection for performance traits since then, to a line hatched in 2000 (B2000). During the first 16 d of lay, feeding treatment had little effect on egg mass or Laying % for the B1980 birds, while for the B2000 birds, SF treatment resulted in significantly greater egg mass and Laying % compared to FF, showing that the OFC indeed manifested in this experiment. However, contrary to hypothesis, follicle hierarchies were normal for both lines under both feeding treatments. To gain further insight into the OFC syndrome, we here report levels of estradiol, testosterone, and progesterone for these line and treatment groups in the time period leading up to and into lay. A significant line × feeding treatment interaction effect was found for estradiol and testosterone, to a lesser extent for progesterone. For all 3 hormones, for B1980 levels 2 to 3 wk post entry into lay were similar and intermediate under FF and SF, but differed significantly for B2000, being much greater under SF than under FF. Thus, the hormonal effects were parallel and may explain the egg mass and Laying % effects of FF and SF in the 2 genetic types.

N -methyladenosine (m 6 A) is the most abundant mRNA modification in eukaryotes, playing crucial roles in multiple biological processes. m 6 A is catalyzed by the activity of methyltransferase-like 3 (Mettl3), which depends on additional proteins whose precise functions remain poorly understood. Here we identified Zc3h13 (zinc finger CCCH domain-containing protein 13)/Flacc [Fl(2)d-associated complex component] as a novel interactor of m 6 A is catalyzed by the activity of methyltransferase-like 3 (Mettl3), which depends on additional proteins whose precise functions remain poorly understood. Here we identified Zc3h13 (zinc finger CCCH domain-containing protein 13)/Flacc [Fl(2)d-associated complex component] as a novel interactor of m 6 and mice. Like other components of this complex, Flacc controls m Drosophila and mice. Like other components of this complex, Flacc controls m 6 We demonstrate that Flacc promotes m Drosophila A deposition by bridging Fl(2)d to the mRNA-binding factor Nito. Altogether, our work advances the molecular understanding of conservation and regulation of the m 6 A deposition by bridging Fl(2)d to the mRNA-binding factor Nito. Altogether, our work advances the molecular understanding of conservation and regulation of the m 6 A machinery., (© 2018 Knuckles et al.; Published by Cold Spring Harbor Laboratory Press.)

RNA modifications are an emerging layer of posttranscriptional gene regulation in eukaryotes. N 6 -methyladenosine (m 6 A) is among the most abundant modifications in mRNAs (mRNAs) that was shown to influence many physiological processes from yeast to mammals. Like DNA methylation, m 6 A in mRNA is dynamically regulated. A conserved methyltransferase complex catalyzes the deposition of the methyl group on adenosine, which can be removed by specific classes of demethylases. Furthermore, YTH-domain containing proteins can recognize this modification to mediate m 6 A-dependent activities. Here we review the functions and mechanisms of the main m 6 A players with a particular focus on Drosophila melanogaster.

Modifications in mRNA constitute ancient mechanisms to regulate gene expression post-transcriptionally. N 6 -methyladenosine (m 6 A) is the most prominent mRNA modification, and is installed by a large methyltransferase complex (the m 6 A 'writer'), not only specifically bound by RNA-binding proteins (the m 6 A 'readers'), but also removed by demethylases (the m 6 A 'erasers'). m 6 A mRNA modifications have been linked to regulation at multiple steps in mRNA processing. In analogy to the regulation of gene expression by miRNAs, we propose that the main function of m 6 A is post-transcriptional fine-tuning of gene expression. In contrast to miRNA regulation, which mostly reduces gene expression, we argue that m 6 A provides a fast mean to post-transcriptionally maximize gene expression. Additionally, m 6 A appears to have a second function during developmental transitions by targeting m 6 A-marked transcripts for degradation., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Background: Mitochondrial diseases due to defective respiratory chain complex III (CIII) are relatively uncommon. The assembly of the eleven-subunit CIII is completed by the insertion of the Rieske iron-sulfur protein, a process for which BCS1L protein is indispensable. Mutations in the BCS1L gene constitute the most common diagnosed cause of CIII deficiency, and the phenotypic spectrum arising from mutations in this gene is wide., Results: A case of CIII deficiency was investigated in depth to assess respiratory chain function and assembly, and brain, skeletal muscle and liver histology. Exome sequencing was performed to search for the causative mutation(s). The patient's platelets and muscle mitochondria showed respiration defects and defective assembly of CIII was detected in fibroblast mitochondria. The patient was compound heterozygous for two novel mutations in BCS1L, c.306A > T and c.399delA. In the cerebral cortex a specific pattern of astrogliosis and widespread loss of microglia was observed. Further analysis showed loss of Kupffer cells in the liver. These changes were not found in infants suffering from GRACILE syndrome, the most severe BCS1L-related disorder causing early postnatal mortality, but were partially corroborated in a knock-in mouse model of BCS1L deficiency., Conclusions: We describe two novel compound heterozygous mutations in BCS1L causing CIII deficiency. The pathogenicity of one of the mutations was unexpected and points to the importance of combining next generation sequencing with a biochemical approach when investigating these patients. We further show novel manifestations in brain, skeletal muscle and liver, including abnormality in specialized resident macrophages (microglia and Kupffer cells). These novel phenotypes forward our understanding of CIII deficiencies caused by BCS1L mutations.

Subtelomeric 1q43q44 microdeletions cause a syndrome associating intellectual disability, microcephaly, seizures and anomalies of the corpus callosum. Despite several previous studies assessing genotype-phenotype correlations, the contribution of genes located in this region to the specific features of this syndrome remains uncertain. Among those, three genes, AKT3, HNRNPU and ZBTB18 are highly expressed in the brain and point mutations in these genes have been recently identified in children with neurodevelopmental phenotypes. In this study, we report the clinical and molecular data from 17 patients with 1q43q44 microdeletions, four with ZBTB18 mutations and seven with HNRNPU mutations, and review additional data from 37 previously published patients with 1q43q44 microdeletions. We compare clinical data of patients with 1q43q44 microdeletions with those of patients with point mutations in HNRNPU and ZBTB18 to assess the contribution of each gene as well as the possibility of epistasis between genes. Our study demonstrates that AKT3 haploinsufficiency is the main driver for microcephaly, whereas HNRNPU alteration mostly drives epilepsy and determines the degree of intellectual disability. ZBTB18 deletions or mutations are associated with variable corpus callosum anomalies with an incomplete penetrance. ZBTB18 may also contribute to microcephaly and HNRNPU to thin corpus callosum, but with a lower penetrance. Co-deletion of contiguous genes has additive effects. Our results confirm and refine the complex genotype-phenotype correlations existing in the 1qter microdeletion syndrome and define more precisely the neurodevelopmental phenotypes associated with genetic alterations of AKT3, ZBTB18 and HNRNPU in humans.

N 6 -methyladenosine (m 6 A) is the most common internal modification of eukaryotic messenger RNA (mRNA) and is decoded by YTH domain proteins. The mammalian mRNA m 6 A methylosome is a complex of nuclear proteins that includes METTL3 (methyltransferase-like 3), METTL14, WTAP (Wilms tumour 1-associated protein) and KIAA1429. Drosophila has corresponding homologues named Ime4 and KAR4 (Inducer of meiosis 4 and Karyogamy protein 4), and Female-lethal (2)d (Fl(2)d) and Virilizer (Vir). In Drosophila, fl(2)d and vir are required for sex-dependent regulation of alternative splicing of the sex determination factor Sex lethal (Sxl). However, the functions of m 6 A in introns in the regulation of alternative splicing remain uncertain. Here we show that m 6 A is absent in the mRNA of Drosophila lacking Ime4. In contrast to mouse and plant knockout models, Drosophila Ime4-null mutants remain viable, though flightless, and show a sex bias towards maleness. This is because m 6 A is required for female-specific alternative splicing of Sxl, which determines female physiognomy, but also translationally represses male-specific lethal 2 (msl-2) to prevent dosage compensation in females. We further show that the m 6 A reader protein YT521-B decodes m 6 A in the sex-specifically spliced intron of Sxl, as its absence phenocopies Ime4 mutants. Loss of m 6 A also affects alternative splicing of additional genes, predominantly in the 5' untranslated region, and has global effects on the expression of metabolic genes. The requirement of m 6 A and its reader YT521-B for female-specific Sxl alternative splicing reveals that this hitherto enigmatic mRNA modification constitutes an ancient and specific mechanism to adjust levels of gene expression.

Modified nucleotides in messenger RNA (mRNA) have been discovered over 40 years ago, but until recently little was known about which transcripts contain them and what their function is. High-throughput sequencing approaches revealed a dynamic landscape of the 'Epitranscriptome' for many mRNA modifications in various organisms from yeast to humans. Meanwhile, also many genes encoding mRNA-modifying enzymes and auxiliary proteins have been identified yielding functional insights by reverse genetics into their role in development and disease., (© 2016 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.)

Objective: Clinical genetic testing is commercially available for rs61764370, an inherited variant residing in a KRAS 3' UTR microRNA binding site, based on suggested associations with increased ovarian and breast cancer risk as well as with survival time. However, prior studies, emphasizing particular subgroups, were relatively small. Therefore, we comprehensively evaluated ovarian and breast cancer risks as well as clinical outcome associated with rs61764370., Methods: Centralized genotyping and analysis were performed for 140,012 women enrolled in the Ovarian Cancer Association Consortium (15,357 ovarian cancer patients; 30,816 controls), the Breast Cancer Association Consortium (33,530 breast cancer patients; 37,640 controls), and the Consortium of Modifiers of BRCA1 and BRCA2 (14,765 BRCA1 and 7904 BRCA2 mutation carriers)., Results: We found no association with risk of ovarian cancer (OR=0.99, 95% CI 0.94-1.04, p=0.74) or breast cancer (OR=0.98, 95% CI 0.94-1.01, p=0.19) and results were consistent among mutation carriers (BRCA1, ovarian cancer HR=1.09, 95% CI 0.97-1.23, p=0.14, breast cancer HR=1.04, 95% CI 0.97-1.12, p=0.27; BRCA2, ovarian cancer HR=0.89, 95% CI 0.71-1.13, p=0.34, breast cancer HR=1.06, 95% CI 0.94-1.19, p=0.35). Null results were also obtained for associations with overall survival following ovarian cancer (HR=0.94, 95% CI 0.83-1.07, p=0.38), breast cancer (HR=0.96, 95% CI 0.87-1.06, p=0.38), and all other previously-reported associations., Conclusions: rs61764370 is not associated with risk of ovarian or breast cancer nor with clinical outcome for patients with these cancers. Therefore, genotyping this variant has no clinical utility related to the prediction or management of these cancers., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Bovine respiratory disease (BRD) is the leading cause of morbidity and mortality in feedlot cattle, caused by multiple pathogens that become more virulent in response to stress. As clinical signs often go undetected and various preventive strategies failed, identification of genes affecting BRD is essential for selection for resistance. Selective DNA pooling (SDP) was applied in a genome wide association study (GWAS) to map BRD QTLs in Israeli Holstein male calves. Kosher scoring of lung adhesions was used to allocate 122 and 62 animals to High (Glatt Kosher) and Low (Non-Kosher) resistant groups, respectively. Genotyping was performed using the Illumina BovineHD BeadChip according to the Infinium protocol. Moving average of -logP was used to map QTLs and Log drop was used to define their boundaries (QTLRs). The combined procedure was efficient for high resolution mapping. Nineteen QTLRs distributed over 13 autosomes were found, some overlapping previous studies. The QTLRs contain polymorphic functional and expression candidate genes to affect kosher status, with putative immunological and wound healing activities. Kosher phenotyping was shown to be a reliable means to map QTLs affecting BRD morbidity.

Somatic mosaicism, the presence of genetically distinct cells within an organism, has been increasingly associated with human morbidity, ranging from being a cause of rare syndromes to a risk factor for common disorders such as malignancy and cardiovascular disease. Previous studies interrogating the normal prevalence of somatic mosaicism have focused on adults. We here present an estimate of the baseline frequency of somatic mosaic copy number variation (CNV) at the time around birth, by sampling eight different organs from a total of five fetuses and newborns. Overall we find a significantly lower frequency of organ specific (i.e. mosaic) CNVs as compared to adults (p = 0.003; Mann-Whitney U-test). The rate of somatic CNV in adults has been estimated to around 2.2 CNV per organ assayed. In contrast, after stringent filtering, we found no organ-private CNVs in fetuses or newborns with exception of the thymus. This organ exhibited a specific genome profile in the form of deletions resulting from polyclonal T-cell receptor rearrangements. This implies that somatic non-immune related CNVs, if present at birth, are typically confined to very small cell populations within organs.

Background and Purpose: The known monogenic forms of stroke are rare. The aim of this study was to analyze pedigrees of young stroke patients regarding possible monogenic cerebrovascular disease and to evaluate the possibility of genetic stroke in these families. This may contribute to a better understanding of disease mechanism in stroke., Methods: Lund Stroke Register includes consecutive patients with first-ever stroke from a defined geographical area in southern Sweden. Early-onset (≤55 years) stroke patients were systematically screened with regard to family history (FHx), and families with stroke aggregation were compiled. Participants provided information in a questionnaire on occurrence of stroke or transient ischaemic attack (TIA) in their families. Information on cardiovascular risk factors (VRFs) and clinical stroke subtype was collected. FHx for stroke was considered positive when the patient reported either ≥1 first-degree relative with stroke/TIA, or no first-degree relative but ≥3 second- or third-degree relatives with stroke/TIA in a distribution compatible with monogenic inheritance., Results: Of 4103 stroke patients registered, 426 (10%) had first-ever stroke at ≤55 years and 338 (79%) of these answered the questionnaire. Of them, 159 (47%) reported a positive FHx. Twenty-eight (18%) of the probands with positive FHx had no known VRFs. Thirty-two families with ≥4 members with stroke were identified. In all these larger families the affected individuals with stroke were present in more than one generation., Conclusion: Aggregation of stroke in families of early-onset stroke patients is not uncommon. Genetic factors with impact on stroke risk, including monogenic causes, need to be evaluated in future stroke studies., (© 2015 EAN.)

Background: The prevalence of Type 2 Diabetes (T2D) mellitus in the past decades, has reached epidemic proportions. Several lines of evidence support the role of genetic variation in the pathogenesis of T2D and insulin resistance. Elucidating these factors could contribute to developing new medical treatments and tools to identify those most at risk. The aim of this study was to characterize the phenotypic response of the Collaborative Cross (CC) mouse genetic resource population to high-fat diet (HFD) induced T2D-like disease to evaluate its suitability for this purpose., Results: We studied 683 mice of 21 different lines of the CC population. Of these, 265 mice (149 males and 116 females) were challenged by HFD (42% fat); and 384 mice (239 males and 145 females) of 17 of the 21 lines were reared as control group on standard Chow diet (18% fat). Briefly, 8 week old mice were maintained on HFD until 20 weeks of age, and subsequently assessed by intraperitoneal glucose tolerance test (IPGTT). Biweekly body weight (BW), body length (BL), waist circumstance (WC), and body mass index (BMI) were measured. On statistical analysis, trait measurements taken at 20 weeks of age showed significant sex by diet interaction across the different lines and traits. Consequently, males and females were analyzed, separately. Differences among lines were analyzed by ANOVA and shown to be significant (P <0.05), for BW, WC, BMI, fasting blood glucose, and IPGTT-AUC. We use these data to infer broad sense heritability adjusted for number of mice tested in each line; coefficient of genetic variation; genetic correlations between the same trait in the two sexes, and phenotypic correlations between different traits in the same sex., Conclusions: These results are consistent with the hypothesis that host susceptibility to HFD-induced T2D is a complex trait and controlled by multiple genetic factors and sex, and that the CC population can be a powerful tool for genetic dissection of this trait.

Neuronally coexpressed ELAV/Hu proteins comprise a family of highly related RNA binding proteins which bind to very similar cognate sequences. How this redundancy is linked to in vivo function and how gene-specific regulation is achieved have not been clear. Analysis of mutants in Drosophila ELAV/Hu family proteins ELAV, FNE, and RBP9 and of genetic interactions among them indicates that they have mostly independent roles in neuronal development and function but have converging roles in the regulation of synaptic plasticity. Conversely, ELAV, FNE, RBP9, and human HuR bind ELAV target RNA in vitro with similar affinities. Likewise, all can regulate alternative splicing of ELAV target genes in nonneuronal wing disc cells and substitute for ELAV in eye development upon artificially increased expression; they can also substantially restore ELAV's biological functions when expressed under the control of the elav gene. Furthermore, ELAV-related Sex-lethal can regulate ELAV targets, and ELAV/Hu proteins can interfere with sexual differentiation. An ancient relationship to Sex-lethal is revealed by gonadal expression of RBP9, providing a maternal fail-safe for dosage compensation. Our results indicate that highly related ELAV/Hu RNA binding proteins select targets for mRNA processing through alteration of their expression levels and subcellular localization but only minimally by altered RNA binding specificity., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Objective: Women with Lynch syndrome (LS) have up to a 60% lifetime risk of endometrial cancer (EC) and up to a 24% risk of ovarian cancer (OC). Gynecological surveillance is recommended, but the benefit and how it should be performed remain unclear. The purpose of this study was to assess diagnostic modalities for gynecological screening of LS patients in Sweden and clinical outcome., Methods: A retrospective nationwide study of 170 women with molecularly confirmed LS. Data including gynecological LS screening history, biopsy results (if any), genetic records, number of screening visits, results from screening including transvaginal ultrasound (TVUS), endometrial biopsy (EB), blood test for tumor marker cancer antigen (CA) 125, prophylactic surgery including age at procedure, and setting from which screening data were obtained from medical records., Results: A total of 117 women were eligible for gynecological screening and of these, 86 patients attended screening visits. Of these, 41 underwent prophylactic hysterectomy and/or bilateral salpingo-oophorectomy. Two patients (4.9%) were diagnosed with EC and two (4.9%) with precancerous lesions in conjunction with prophylactic surgery. Total incidence of gynecological cancer in the surveillance group (45 women) was 20% EC, 4% OC. Five patients had endometrial cancer or complex hyperplasia with atypia (n=2) detected by endometrial biopsy. Four additional cases were detected due to interval bleeding. Both cases of ovarian cancer were detected by transvaginal ultrasound in patients with ovarian cysts under surveillance. The youngest woman with endometrial cancer was diagnosed at 35 years of age, before she was aware of her diagnosis of Lynch syndrome., Conclusions: Gynecological surveillance of women with Lynch syndrome may lead to earlier detection of precancerous lesions, which might have some impact on the morbidity from endometrial cancer although further studies are needed to prove this. Prophylactic hysterectomy with or without bilateral salpingo-oophorectomy reduces the cancer incidence. A practical approach to surveillance in Lynch syndrome women would be to offer annual surveillance beginning at age 30 years including probably both TVUS and EB in order to increase diagnostic yield with prospective data registry for follow-up studies. Prophylactic surgery could be performed at a suitable age after childbearing to obtain a balance between reducing the risk of cancer and minimizing long-term complications from premature menopause., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: High-resolution mapping of the loci (QTN) responsible for genetic variation in quantitative traits is essential for positional cloning of candidate genes, and for effective marker assisted selection. The confidence interval (QTL) flanking the point estimate of QTN-location is proportional to the number of individuals in the mapping population carrying chromosomes recombinant in the given interval. Consequently, many designs for high resolution QTN mapping are based on increasing the proportion of recombinants in the mapping population. The "Targeted Recombinant Progeny" (TRP) design is a new design for high resolution mapping of a target QTN in crosses between pure, or inbred lines. It is a three-generation procedure generating a large number of recombinant individuals within a QTL previously shown to contain a QTN. This is achieved by having individuals that carry chromosomes recombinant across the target QTL interval as parents of a large mapping population; most of whom will therefore carry recombinant chromosomes targeted to the given QTL. The TRP design is particularly useful for high resolution mapping of QTN that differentiate inbred or pure lines, and hence are not amenable to high resolution mapping by genome-wide association tests., Results: In the absence of residual polygenic variation, population sizes required for achieving given mapping resolution by the TRP-F2 design relative to a standard F2 design ranged from 0.289 for a QTN with standardized allele substitution effect = 0.2, mapped to an initial QTL of 0.2 Morgan to 0.041 for equivalent QTN mapped to an initial QTL of 0.02 M. In the presence of residual polygenic variation, the relative effectiveness of the TRP design ranges from 1.068 to 0.151 for the same initial QTL intervals and QTN effect. Thus even in the presence of polygenic variation, the TRP can still provide major savings. Simulation showed that mapping by TRP should be based on 30-50 markers spanning the initial interval; and on at least 50 or more G2 families representing this number of recombination points,., Conclusions: The TRP design can be an effective procedure for achieving high and ultra-high mapping resolution of a target QTN previously mapped to a known confidence interval (QTL).