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Introduction: Our aim was to investigate the insufficiently understood differences in the immune system between anti-citrullinated peptide antibody (ACPA)-positive (ACPA+) and ACPA-negative (ACPA-) early rheumatoid arthritis (eRA) patients. Methods: We performed multiple cytokine assays using sera from drug-naïve ACPA+ and ACPA- eRA patients. Additionally, we conducted single-cell RNA sequencing of CD45+ cells from peripheral blood samples to analyze and compare the distribution and functional characteristics of the cell subsets based on the ACPA status. Results: Serum concentrations of interferon-γ (IFN-γ) and interleukin (IL)-12 were higher in ACPA+ eRA than in ACPA- eRA. Single-cell transcriptome analysis of 37,318 cells identified 17 distinct cell types and revealed the expansion of IL1B+ proinflammatory monocytes, IL7R+ T cells, and CD8+ CCL4+ T cells in ACPA+ eRA. Furthermore, we observed an enrichment of IFN-γ response genes in nearly all monocytes and T cells of ACPA+ eRA subsets. Heightened interactions between IFN-γ and IFN-γ receptors were observed in ACPA+ eRA, particularly between monocytes and T cells. We examined IFITM2 and IFITM3 as potential key markers in ACPA+ eRA given their pronounced upregulation and association with the IFN response. Specifically, the expression of these genes was elevated in IL1B+ proinflammatory monocytes (likely M1 monocytes), correlating with serum IFN-γ levels. Discussion: Compared to ACPA- eRA, ACPA+ eRA showed higher serum IFN-γ and IL-12 levels, upregulated IFN-γ response genes, and enhanced IFN-γ-driven monocyte-T cell interactions. These distinct immune features of the peripheral circulation in ACPA+ eRA suggest a role for type 1 helper T cell-related immunity in its pathogenesis. [ABSTRACT FROM AUTHOR]

Background : Silver arowana (Osteoglossum bicirrhosum) is a basal fish species with sexual monomorphism, while its sex determination mechanism has been poorly understood, posing a significant challenge to its captive breeding efforts. Results: We constructed two high-quality chromosome-level genome assemblies for both female and male silver arowana, with scaffold N50 values over 10 Mb. Combining re-sequencing data of 109 individuals, we identified a female-specific region, which was localized in a non-coding region, i.e., around 26-kb upstream of foxl2 gene (encoding forkhead box L2). Its strong interaction with the neighboring foxl2 on the same chromosome suggests foxl2 as a candidate sex-related gene in silver arowana. We subsequently propose a complex gene network in the sex determination process of silver arowana, with foxl2 acting as the central contributor. Transcriptome sequencing of gonads support our hypothesis that the regulation of foxl2 can be influenced by the spatial proximity of the female-specific fragment, thereby promoting ovarian function or inhibiting testicular function to stimulate gonadal differentiation. Furthermore, we found the sex chromosomes to be homomorphic with a potentially recent origin, as a linkage disequilibrium analysis proved minor recombination suppression. Conclusions: These results taken together serve as a crucial foundation for conducting extensive investigations on the evolution and differentiation of sex-determining mechanisms, as well as the emergence and development of sex chromosomes in various fishes. [ABSTRACT FROM AUTHOR]

Background: Sleep is a conserved physiological phenomenon across species. It is mainly controlled by two processes: a circadian clock that regulates the timing of sleep and a homeostat that regulates the sleep drive. Even cnidarians, such as Hydra and jellyfish, which lack a brain, display sleep-like states. However, the manner in which environmental cues affect sleep-like states in these organisms remains unknown. In the present study, we investigated the effects of light and temperature cycles on the sleep-like state in Hydra vulgaris. Results: Our findings indicate that Hydra responds to temperature cycles with a difference of up to 5° C, resulting in decreased sleep duration under light conditions and increased sleep duration in dark conditions. Furthermore, our results reveal that Hydra prioritizes temperature changes over light as an environmental cue. Additionally, our body resection experiments show tissue-specific responsiveness in the generation ofthe sleep-like state under different environmental cues. Specifically, the upper body can generate the sleep-like state in response to a single environmental cue. In contrast, the lower body did not respond to 12-h light–dark cycles at a constant temperature. Conclusions: These findings indicate that both light and temperature influence the regulation of the sleep-like state in Hydra. Moreover, these observations highlight the existence of distinct regulatory mechanisms that govern patterns of the sleep-like state in brainless organisms, suggesting the potential involvement of specific regions for responsiveness of environmental cues for regulation of the sleep-like state. [ABSTRACT FROM AUTHOR]

Trichoderma spp. are among the most studied biocontrol agents. While extensive work has been done to understand Trichoderma antagonistic mechanisms, additional research is needed to fully understand how Trichoderma spp. recognize the pathogen-host and the intra-species variability i frequently observed upon interaction with a specific pathogen-host. This study focuses on elucidating the mechanisms underlying observed phenotypic differences among the T. afroharzianum isolates Th19A and Th4 during confrontation with Fusarium virguliforme by investigating differences in their transcriptome at different stages of interaction. In a dual plate assay, Th19A overgrows F. virguliforme, whereas Th4 forms an inhibition zone. Significant differences were observed in the F. virguliforme transcriptome upon interaction with Th19A compared to Th4 and across the different stages of interaction. GO molecular function categories enriched for F. virguliforme genes differed, indicating possible transcriptional plasticity upon interaction with Th19A versus Th4. Significant transcriptome changes were also observed in T. afroharzianum, with several differences in GO-enriched categories between isolates. Several differentially expressed genes-encoding secreted proteins, including CAZymes and CBM1-domain-containing proteins, were up-regulated in Th19A and Th4 upon interaction with the pathogen, even before physical contact, demonstrating possible volatile-mediated recognition of both isolates by F. virguliforme. This study contributes to a better understanding of the interaction between T. afroharzianum and F. virguliforme, which is crucial for developing efficient biological control programs. [ABSTRACT FROM AUTHOR]

Prostate-Specific Membrane Antigen (PSMA) is a highly expressed and structurally unique target specific to prostate cancer (PCa). Diagnostic and therapeutic approaches in nuclear medicine, coupling PSMA ligands with radionuclides, have shown significant clinical success. PSMA-PET/CT effectively identifies tumors and metastatic lymph nodes for imaging purposes, while 177Lu-PSMA-617 (Pluvicto) has received FDA approval for treating metastatic castration-resistant PCa (mCRPC). Despite their success, radionuclide-based diagnostic and therapeutic methods face limitations such as high costs and significant side effects. Recently, near-infrared (NIR) fluorescence imaging and phototherapy have advanced significantly in biomedical applications. It's benefits, such as deep tissue penetration, real-time precision, and minimal side effects, have driven broader clinical adoption, especially in fluorescence-guided surgery (FGS). This review suggests combining NIR dyes with PSMA ligands to enable targeted, high-resolution imaging with superior signal-to-background ratios, facilitating precise FGS. NIR techniques can also aid pathological diagnosis in ex vivo specimens. Furthermore, combining photosensitizers with PSMA ligands allows localized photothermal (PTT) or photodynamic therapy (PDT) under NIR irradiation, producing heat or reactive oxygen species (ROS) to treat PCa. This review aims to extend the clinical success of radionuclide-based PSMA targeting by exploring advances in NIR-based FGS and phototherapy, presenting a promising new diagnostic and therapeutic approach. [ABSTRACT FROM AUTHOR]

The mammalian nervous system controls complex functions through highly specialized and interacting structures. Single-cell sequencing can provide information on cell-type-specific chromatin structure and regulatory elements, revealing differences in chromatin organization between different cell types and their potential roles of these differences in brain function. Here, we generated a chromatin accessibility dataset through single-cell ATAC-seq of 174,593 high-quality nuclei from 16 adult rat brain regions. We identified cell subtypes of both neuronal and non-neuronal cells with highly specific distributions and characterized gene regulatory elements associated with cell type-specific regions. To further investigate the gene regulatory network involved in spinal cord regeneration, we integrated our scATAC-seq data with published single-nucleus RNA-seq data from the spinal cord, and we identified more detailed regeneration related elements by drawing GRNs centered on the transcription factor Jun in the OPC. We also performed similar integration analyses in the midbrain. Our findings provide a solid foundation for the comprehensive dissection of the molecular architecture of the mammalian nervous system. [ABSTRACT FROM AUTHOR]

Monocarpic plants flower only once and then produce seeds. Many monocarpic plants require a cold treatment known as vernalization before they flower. This requirement delays flowering until the plant senses warm temperatures in the spring. Exposure to high temperatures following vernalization causes devernalization, which cancels the vernalized state, inhibiting flowering and promoting vegetative growth. In this study, we screened over 16,000 chemical compounds and identified five small molecules (devernalizers; DVRs) that induce devernalization in Arabidopsis thaliana at room temperature without requiring a high-temperature treatment. Treatment with DVRs reactivated the expression of FLOWERING LOCUS C (FLC), a master repressor of flowering, by reducing the deposition of repressive histone modifications, thereby delaying flowering time. Three of the DVRs identified shared two structures: a hydantoin-like region and a spiro-like carbon. Treatment with DVR06, which has a simple chemical structure containing these domains, delayed flowering time and reduced the deposition of repressive histone modifications at FLC. RNA-seq and ChIP-seq analyses revealed both shared and specific transcriptomic and epigenetic effects between DVR06- and heat-induced devernalization. Overall, our extensive chemical screening indicated that hydantoin and spiro are key chemical signatures that reduce repressive histone modifications and promote devernalization in plants. Five compounds that delay flowering time are isolated by high-throughput chemical screening; these small molecules could potentially be used to increase crop and vegetable yields in agricultural fields. [ABSTRACT FROM AUTHOR]

Coronaviruses express their structural and accessory genes via a set of subgenomic RNAs, whose synthesis is directed by transcription regulatory sequences (TRSs) in the 5′ genomic leader and upstream of each body open reading frame. In SARS-CoV-2, the TRS has the consensus AAACGAAC; upon searching for emergence of this motif in the global SARS-CoV-2 sequences, we find that it evolves frequently, especially in the 3′ end of the genome. We show well-supported examples upstream of the Spike gene—within the nsp16 coding region of ORF1b—which is expressed during human infection, and upstream of the canonical Envelope gene TRS, both of which have evolved convergently in multiple lineages. The most frequent neo-TRS is within the coding region of the Nucleocapsid gene, and is present in virtually all viruses from the B.1.1 lineage, including the variants of concern Alpha, Gamma, Omicron and descendants thereof. Here, we demonstrate that this TRS leads to the expression of a novel subgenomic mRNA encoding a truncated C-terminal portion of Nucleocapsid, which is an antagonist of type I interferon production and contributes to viral fitness during infection. We observe distinct phenotypes when the Nucleocapsid coding sequence is mutated compared to when the TRS alone is ablated. Our findings demonstrate that SARS-CoV-2 is undergoing evolutionary changes at the functional RNA level in addition to the amino acid level. Coronaviruses can evolve novel transcription regulatory sequences, generating new RNA transcripts during human infection. This study identifies a prevalent subgenomic RNA in SARS-CoV-2, which produces a C-terminal nucleocapsid fragment that suppresses interferon signaling and enhances viral fitness. [ABSTRACT FROM AUTHOR]

Rosa laevigata is an excellent rose germplasm, highly resistant to aphid, and immune to both rose black spot and powdery mildew disease. It is also a well-known edible plant with a long history of medicinal use in China, having the effects of improving kidney function, inhibiting arteriosclerosis, and reducing inflammation. In this study, we assembled a high-quality chromosome-scale genome for R. laevigata by combining Illumina, PacBio, and Hi-C data, which has a length of approximately 494.2 Mb with a scaffold N50 of 68.6 Mb. A total of 493.2 Mb (99.8%) of the draft genome sequences were anchored on seven pseudochromosomes and two gapless pseudochromosomes were included in the final genome assembly. A total of 37,117 protein-coding genes were predicted, 34,047 of which were functionally annotated. Repeat annotation revealed 659,558 (285.6 Mb) repeat elements, accounting for 57.8% of the genome. The chromosome-scale genome provides valuable information to facilitate comparative genomic analysis of rose family and will accelerate genome-guided breeding and germplasm improvement of both R. laevigata itself and modern roses. [ABSTRACT FROM AUTHOR]

Background: Chickens and ducks are vital sources of animal protein for humans. Recent pangenome studies suggest that a single genome is insufficient to represent the genetic information of a species, highlighting the need for more comprehensive genomes. The bird genome has more than tens of microchromosomes, but comparative genomics, annotations, and the discovery of variations are hindered by inadequate telomere-to-telomere level assemblies. We aim to complete the chicken and duck genomes, recover missing genes, and reveal common and unique chromosomal features between birds. Results: The near telomere-to-telomere genomes of Silkie Gallus gallus and Mallard Anas platyrhynchos were successfully assembled via multiple high-coverage complementary technologies, with quality values of 36.65 and 44.17 for Silkie and Mallard, respectively; and BUSCO scores of 96.55% and 96.97% for Silkie and Mallard, respectively; the mapping rates reached over 99.52% for both assembled genomes, these evaluation results ensured high completeness and accuracy. We successfully annotated 20,253 and 19,621 protein-coding genes for Silkie and Mallard, respectively, and assembled gap-free sex chromosomes in Mallard for the first time. Comparative analysis revealed that microchromosomes differ from macrochromosomes in terms of GC content, repetitive sequence abundance, gene density, and levels of 5mC methylation. Different types of arrangements of centromeric repeat sequence centromeres exist in both Silkie and the Mallard genomes, with Mallard centromeres being invaded by CR1. The highly heterochromatic W chromosome, which serves as a refuge for ERVs, contains disproportionately long ERVs. Both Silkie and the Mallard genomes presented relatively high 5mC methylation levels on sex chromosomes and microchromosomes, and the telomeres and centromeres presented significantly higher 5mC methylation levels than the whole genome. Finally, we recovered 325 missing genes via our new genomes and annotated TNFA in Mallard for the first time, revealing conserved protein structures and tissue-specific expression. Conclusions: The near telomere-to-telomere assemblies in Mallard and Silkie, with the first gap-free sex chromosomes in ducks, significantly enhanced our understanding of genetic structures in birds, specifically highlighting the distinctive chromosome features between the chicken and duck genomes. This foundational work also provides a series of newly identified missing genes for further investigation. [ABSTRACT FROM AUTHOR]

Avian genomes exhibit compact organization and remarkable chromosomal stability. However, the extent and mechanisms by which structural variation in avian genomes differ from those in other vertebrate lineages are poorly explored. This study generated a diploid genome assembly for the golden pheasant (Chrysolophus pictus), a species distinguished by the vibrant plumage of males. Each haploid genome assembly included complete chromosomal models, incorporating all microchromosomes. Analysis revealed extensive tandem amplification of immune-related genes across the smallest microchromosomes (dot chromosomes), with an average copy number of 54. Structural variation between the haploid genomes was primarily shaped by large insertions and deletions (indels), with minimal contributions from inversions or duplications. Approximately 28% of these large indels were associated with recent insertions of transposable elements, despite their typically low activity in bird genomes. Evidence for significant effects of transposable elements on gene expression was minimal. Evolutionary strata on the sex chromosomes were identified, along with a drastic rearrangement of the W chromosome. These analyses of the high-quality diploid genome of the golden pheasant provide valuable insights into the evolutionary patterns of structural variation in avian genomes. [ABSTRACT FROM AUTHOR]

Recent studies in vertebrates and Caenorhabditis elegans have reshaped models of how the axon guidance cue UNC-6/Netrin functions in dorsal-ventral axon guidance, which was traditionally thought to form a ventral-to-dorsal concentration gradient that was actively sensed by growing axons. In the vertebrate spinal cord, floorplate Netrin1 was shown to be largely dispensable for ventral commissural growth. Rather, short range interactions with Netrin1 on the ventricular zone radial glial stem cells was shown to guide ventral commissural axon growth. In C. elegans, analysis of dorsally-migrating growth cones during outgrowth has shown that growth cone polarity of filopodial extension is separable from the extent of growth cone protrusion. Growth cones are first polarized by UNC-6/Netrin, and subsequent regulation of protrusion by UNC-6/Netrin is based on this earlier-established polarity (the Polarity/Protrusion model). In both cases, short-range or even haptotactic mechanisms are invoked: in vertebrate spinal cord, interactions of growth cones with radial glia expressing Netrin-1; and in C. elegans, a potential close-range interaction that polarizes the growth cone. To explore potential short-range and long-range functions of UNC-6/Netrin, a potentially membrane-anchored transmembrane UNC-6 (UNC-6(TM)) was generated by genome editing. unc-6(tm) was hypomorphic for dorsal VD/DD axon pathfinding, indicating that it retained some unc-6 function. Polarity of VD growth cone filopodial protrusion was initially established in unc-6(tm), but was lost as the growth cones migrated away from the unc-6(tm) source in the ventral nerve cord. In contrast, ventral guidance of the AVM and PVM axons was equally severe in unc-6(tm) and unc-6(null). Together, these results suggest that unc-6(tm) retains short-range functions but lacks long-range functions due to reduced secreted UNC-6. Ectopic unc-6(+) expression from non-ventral sources did not dramatically perturb dorsal VD growth cone polarity or axon outgrowth, suggesting that ectopic UNC-6 cannot redirect polarity once it is established in the VD/DD neurons. This is not what would be expected of a growth cone dynamically reading a gradient of UNC-6, but is consistent with the Polarity/protrusion model of growth cone guidance away from UNC-6/Netrin. Author summary: In the developing nervous system, neurons extend thin cellular processes called axons that make precise contacts with other neurons in elaborate networks. The structure of these axon networks underlies the function of the nervous system. Thus precise axon guidance is critical during development to produce a functional nervous system. Axons are guided by the growth cone, a dynamic structure at the distal tup of the axon that senses and responds to signals in the environment that instruct axon guidance. One such signal if UNC-6/Netrin, a conserved, laminin-like secreted molecule that guides many aspects of axon guidance. In the nematode Caenorhabditis elegans, UNC-6 controls dorsal-ventral axon guidance of motor neurons and sensory neurons. The canonical textbook model of UNC-6/Netrin function involved the formation of a gradient of secreted UNC-6/Netrin in the ventral-to-dorsal axis which growth cones dynamically sensed and responded by growing up or down the gradient. The gradient model does not explain recent studies in vertebrates and C. elegans that have shown close-range or contact-mediated events that more resemble cell polarity events than dynamic reading of a gradient. In C. elegans, the growth cone of the VD axon is polarized dorsally by UNC-6/Netrin. UNC-6/Netrin then maintains this polarity and regulates protrusion based upon this polarity, stimulating protrusion dorsally and inhibiting protrusion ventrally, resulting in net dorsal outgrowth. This Polarity/Protrusion model resembles events that take place in ventral growth of sensory axons, the Statistically-Oriented Asymmetric Localization (SOAL) model. The Polarity/Protrusion model predicts that a short-range UNC-6 signal polarizes the VD growth cone, and that secreted, diffusible UNC-6/Netrin maintains initial polarity and regulates protrusion. This model is critically tested here by the construction of a membrane-anchored UNC-6 genome edit predicted to not produce any diffusible UNC-6, thus blocking any long-range function. Data here show that VD growth cones are initially polarized corerectly in unc-6(tm), but that this polarity is lost as the growth cones move dorsally away from the UNC-6(TM) source. This is consistent with a short-range interaction that polarizes the growth cone, and a long-range function needed to maintain polarity. Furthermore, ectopic expression of diffusible UNC-6 from ectopic sources did not perturb VD growth cone polarity, suggesting that the initial polarity event cannot be altered by later diffusible UNC-6/Netrin, as would be expected with a gradient model. These results are consistent with the Polarity/Protrusion model of UNC-6 function. [ABSTRACT FROM AUTHOR]

Objectives: Emerging data have shown that local treatment could provide clinical benefit for non-small cell lung cancer (NSCLC) patients with oligometastasis. Liver metastases have the worst prognosis in advanced NSCLC, but the genomic characteristics of liver oligometastasis remain unclear. The aim of our study was to elucidate the molecular features of liver oligometastatic NSCLC. Methods: Paired liver metastatic tissue samples and peripheral blood from 32 liver oligometastatic NSCLC patients were concurrently collected for comprehensive genomic analysis using next-generation sequencing. Results: A total of 206 mutated genes in 32 patients were detected, with a median of 4 mutations per sample. The most frequent alterations (> 10%) in liver oligometastasis were TP53 (72%), EGFR (50%), RB1 (19%) and SMARCA4 (12%). The co-occurrence rate of TP53 and RB1 in our cohort was significantly higher than that in the TCGA-LUAD cohort. Age, APOBEC, homologous recombination deficiency (HRD) and deficient mismatch repair (dMMR) established the mutational signature of liver oligometastatic NSCLC. The median tumor mutation burden (TMB) was 4.8 mutations/Mb. A total of 78.12% patients harbored at least one potentially actionable molecular alteration that may guide further targeted therapy according to the OncoKB evidence. Conclusions: Our study comprehensively delineated the genomic characteristics of liver oligometastatic NSCLC - such findings were helpful to better understand the distinct clinic-biological features of oligometastasis and optimize personalized treatment of this population. [ABSTRACT FROM AUTHOR]

The remarkable diversity of insect pigmentation offers a captivating avenue for studying evolution and genetics. In tephritids, understanding the molecular basis of mutant traits is also crucial for applied entomology, enabling the creation of genetic sexing strains through genome editing, thus facilitating sex-sorting before sterile insect releases. Here, we present evidence from classical and modern genetics showing that the black pupae (bp) phenotype in the GUA10 strain of Anastrepha ludens is caused by a large deletion at the ebony locus, removing the gene’s entire coding region. Targeted knockout of ebony induced analogous bp phenotypes across six major tephritid agricultural pests, demonstrating that disruption of Ebony alone is sufficient to produce the mutant trait in distantly related species. This functional characterization further allowed a deeper exploration of Ebony’s role in pigmentation and development across life stages in diverse species. Our findings offer key insights for molecular engineering of sexing strains based on the bp marker and for future evolutionary developmental biology studies in tephritids.This study integrates classical and modern genetics to show that the ebony gene is linked to the black pupae phenotype in tephritids, offering new insights into Ebony’s role in the evolutionary developmental biology of this diverse group of flies [ABSTRACT FROM AUTHOR]

In early mammalian embryogenesis, a shift from non-canonical histone H3 lysine 4 trimethylation (H3K4me3) linked to transcriptional repression to canonical H3K4me3 indicating active promoters occurs during zygotic genome activation (ZGA). However, the mechanisms and roles of these H3K4me3 states in embryogenesis remain poorly understood. Our research reveals that the histone methyltransferase MLL2 is responsible for installing H3K4me3 (both non-canonical and canonical) in totipotent embryos, while a transition to SETD1A/B-deposited H3K4me3 occurs in pluripotent embryos. Interestingly, MLL2-mediated H3K4me3 operates independently of transcription, fostering a relaxed chromatin state conducive to totipotency rather than directly influencing transcription. Conversely, SETD1A/B-mediated H3K4me3, which depends on transcription, is crucial for facilitating expression of genes essential for pluripotency and pre-implantation development. Our findings highlight the role of the H3K4me3 transition, mediated by an MLL2-to-SETD1A/B relay mechanism, in the regulation of transition from totipotency to pluripotency during early embryogenesis. Synopsis: During mammalian zygotic genome activation, a shift occurs from non-canonical histone H3 lysine 4 trimethylation (H3K4me3) linked to transcriptional repression to canonical H3K4me3 indicating active promoters. This study reveals that early embryos undergo the transition from totipotency to pluripotency by employing a relay between histone methyltransferases MLL2 and SETD1A/B to deposit H3K4me3. MLL2-dependent canonical H3K4me3 deposition occurs in a transcription-independent manner. SETD1A/B maintained canonical H3K4me3 distribution in a transcription-dependent manner. Disruption of MLL2-mediated H3K4me3 deposition does not affect zygotic genome activation. Transcription-dependent H3K4me3 catalyzed by SETD1A/B is essential for the first cell fate decision. SETD1A/B-dependent histone H3 lysine 4 trimethylation at active promoters is required for the first cell fate transition in mouse early embryogenesis. [ABSTRACT FROM AUTHOR]

Background: The determinants of differences in host infectivity among Cryptosporidium species and subtypes are poorly understood. Results from recent comparative genomic studies suggest that gains and losses of multicopy subtelomeric genes encoding insulinase-like proteases (INS-19 and INS-20 in Cryptosporidium parvum and their orthologs in closely related species) may potentially contribute to these differences. Methodology/Principal findings: In this study, we investigated the expression and biological function of the INS-19 and INS-20 of C. parvum. CRISPR/Cas9 was used to endogenously tag both genes with the hemagglutinin epitope. Immunofluorescence analysis revealed that INS-19 and INS-20 are expressed at different developmental stages of the pathogen. Although knockout of either had no detectable effect on the in vitro growth of C. parvum, knockout of INS-20, deletion of its multiple domains, or mutation of the active motif in the functional domain reduced the intensity of C. parvum infection in IFN-γ knockout mice. Consistent with this, mice infected with the INS-20-deleted mutant had reduced intestinal damage and parasite burden. Conclusions/Significance: These results suggest that INS-19 and INS-20 have stage-specific expression with distinct biological functions, and that the presence of the INS-20 in zoonotic C. parvum contributes to its infectivity and fitness in mice. Author summary: Understanding why Cryptosporidium species and isolates differ in infectivity to their hosts is a complex puzzle. Recent comparative genomic analyses suggest that copy number variation of genes may be a contributing factor. In this research, we focused on two subtelomeric genes encoding the proteases INS-19 and INS-20 in C. parvum, the most important zoonotic Cryptosporidium species. We genetically tagged and engineered these genes to study their expression and function. Our results showed that INS-19 and INS-20 are expressed at different stages of the parasite life cycle. Interestingly, while knocking out each gene did not affect parasite growth in vitro, deleting or mutating INS-20 significantly reduced the severity of infection in mice. Mice infected with the C. parvum mutants had less intestinal damage and lower parasite loads. This study highlights the distinct roles of these two proteases and suggests that INS-20, in particular, plays a critical role in the parasite's ability to infect and cause disease. These findings may facilitate the development of new drugs for cryptosporidiosis. [ABSTRACT FROM AUTHOR]

The synaptonemal complex (SC) is a protein-rich structure essential for meiotic recombination and faithful chromosome segregation. Acting like a zipper to paired homologous chromosomes during early prophase I, the complex is a symmetrical structure where central elements are connected on two sides by the transverse filaments to the chromatin-anchoring lateral elements. Despite being found in most major eukaryotic taxa implying a deeply conserved evolutionary origin, several components of the complex exhibit unusually high rates of sequence turnover. This is puzzlingly exemplified by the SC of Drosophila, where the central elements and transverse filaments display no identifiable homologs outside of the genus. Here, we exhaustively examine the evolutionary history of the SC in Drosophila taking a comparative phylogenomic approach with high species density to circumvent obscured homology due to rapid sequence evolution. Contrasting starkly against other genes involved in meiotic chromosome pairing, SC genes show significantly elevated rates of coding evolution due to a combination of relaxed constraint and recurrent, widespread positive selection. In particular, the central element cona and transverse filament c(3) G have diversified through tandem and retro-duplications, repeatedly generating paralogs with novel germline activity. In a striking case of molecular convergence, c(3) G paralogs that independently arose in distant lineages evolved under positive selection to have convergent truncations to the protein termini and elevated testes expression. Surprisingly, the expression of SC genes in the germline is prone to change suggesting recurrent regulatory evolution which, in many species, resulted in high testes expression even though Drosophila males are achiasmic. Overall, our study recapitulates the poor conservation of SC components, and further uncovers that the lack of conservation extends to other modalities including copy number, genomic locale, and germline regulation. Considering the elevated testes expression in many Drosophila species and the common ancestor, we suggest that the activity of SC genes in the male germline, while still poorly understood, may be a prime target of constant evolutionary pressures driving repeated adaptations and innovations. Author summary: The synaptonemal complex (SC) is essential for meiotic recombination and faithful chromosome segregation across eukaryotes, yet components of the SC are often poorly conserved. Here, we show that across the Drosophila phylogeny, SC genes have evolved under recurrent positive selection, resulting in orthologs and paralogs often with barely recognizable amino acid sequences. This is partly driven by duplications repeatedly generating paralogs, many of which appear to have adopted novel germline expression patterns, often highly active in the testes. While most SC genes are thought to be dispensable in the male germline of Drosophila where meiotic recombination does not occur, elevated testes expression independently emerged in different lineages and appears to be the norm across the genus and likely the ancestral state. Unexpectedly, SC expression in ovaries is also poorly conserved, revealing recurrent regulatory turnover. We suggest that the evolutionary lability of SC genes in Drosophila is likely a repeated source of functional diversifications and innovations in the germline. [ABSTRACT FROM AUTHOR]

Intracellular electrophysiology is essential in neuroscience, cardiology, and pharmacology for studying cells' electrical properties. Traditional methods like patch-clamp are precise but low-throughput and invasive. Nanoelectrode Arrays (NEAs) offer a promising alternative by enabling simultaneous intracellular and extracellular action potential (iAP and eAP) recordings with high throughput. However, accessing intracellular potentials with NEAs remains challenging. This study presents an AI-supported technique that leverages thousands of synchronous eAP and iAP pairs from stem-cell-derived cardiomyocytes on NEAs. Our analysis revealed strong correlations between specific eAP and iAP features, such as amplitude and spiking velocity, indicating that extracellular signals could be reliable indicators of intracellular activity. We developed a physics-informed deep learning model to reconstruct iAP waveforms from extracellular recordings recorded from NEAs and Microelectrode arrays (MEAs), demonstrating its potential for non-invasive, long-term, high-throughput drug cardiotoxicity assessments. This AI-based model paves the way for future electrophysiology research across various cell types and drug interactions. Intracellular recording involves puncturing the cell membrane to gain access to the cell. In this work, the authors introduce a puncture-free intracellular recording approach that leverages a deep learning model to translate extracellular recordings into intracellular signals. [ABSTRACT FROM AUTHOR]

Infectious spleen and kidney necrosis virus (ISKNV) is a highly virulent and rapidly transmissible fish virus that poses threats to the aquaculture of a wide variety of freshwater and marine fish. N6-methyladenosine (m6A), recognized as a common epigenetic modification of RNA, plays an important regulatory role during viral infection. However, the impact of m6A RNA methylation on the pathogenicity of ISKNV remains unexplored. Here, methylated RNA immunoprecipitation sequencing (MeRIP-seq) coupled with RNA sequencing (RNA-seq) was used to systematically detect variations in m6A methylation and gene expression between ISKNV-infected and noninfected MFF-1 cells, followed by functional enrichment and co-expression joint analysis. The findings revealed that the m6A methylation peaks were located mainly in coding sequences (CDSs), with more than 90% of the transcripts containing 1–5 m6A peaks. Through MeRIP-seq, 4361 differentially m6A-methylated mRNAs were identified. Gene enrichment analysis revealed that m6A-related genes were enriched in biological processes and pathways such as gene expression, cellular structure, immune responses, and cell death. Co-expression analysis revealed that the genes differentially expressed at both the mRNA and m6A modification levels were enriched in pathways such as the hippo, ErbB, and JAK-STAT pathways. The m6A modification at the genome-wide transcription level of ISKNV was subsequently shown to be pronounced in several pivotal genes, such as putative vascular endothelial growth factor, ribonucleotide reductase small subunit, and E3 ubiquitin ligase. This study comprehensively describes the m6A expression profile in ISKNV- and ISKNV-infected MFF-1 cells, providing a basis for investigating the role of m6A modification during ISKNV infection. [ABSTRACT FROM AUTHOR]

Argali stands as the largest species among wild sheep in Central and East Asia, with a concerning rate of decline estimated at 30%. The intraspecific taxonomy of argali remains contentious due to limited genomic data and unclear geographic separation. In this study, we constructed a chromosome-level genome assembly and annotation for the Tibetan argali (O. a. hodgsoni), together with population genomic resequencing of 32 individuals representing four subspecies. The contig-level genome was 2.64 Gb in size, with a contig N50 length of 71.69 Mb and an estimated genomic completeness of 96.01%. Using Hi-C sequencing data scaffolding, 99.90% of initially assembled sequences were mapped and oriented onto 28 pseudo-chromosomes except the Y chromosome. Annotation uncovered 21,564 protein-coding genes and 46.38% repeat sequences. The average coverage of the population resequencing data was 23.74 with mean mapping ratio up to of 97.19%. The high-quality genome assembly and annotation of the Tibetan argali, coupled with the high-depth population genomic data, will serve as a valuable genetic resource for studies on the taxonomy and conservation of argali. [ABSTRACT FROM AUTHOR]

Background: A subset of developmental disorders (DD) is characterized by disease-specific genome-wide methylation changes. These episignatures inform on the underlying pathogenic mechanisms and can be used to assess the pathogenicity of genomic variants as well as confirm clinical diagnoses. Currently, the detection of these episignature requires the use of indirect methylation profiling methodologies. We hypothesized that long-read whole genome sequencing would not only enable the detection of single nucleotide variants and structural variants but also episignatures. Methods: Genome-wide nanopore sequencing was performed in 40 controls and 20 patients with confirmed or suspected episignature-associated DD, representing 13 distinct diseases. Following genomic variant and methylome calling, hierarchical clustering and dimensional reduction were used to determine the compatibility with microarray-based episignatures. Subsequently, we developed a support vector machine (SVM) for the detection of each DD. Results: Nanopore sequencing-based methylome patterns were concordant with microarray-based episignatures. Our SVM-based classifier identified the episignatures in 17/19 patients with a (likely) pathogenic variant and none of the controls. The remaining patients in which no episignature was identified were also classified as controls by a commercial microarray assay. In addition, we identified all underlying pathogenic single nucleotide and structural variants and showed haplotype-aware skewed X-inactivation evaluation directs clinical interpretation. Conclusion: This proof-of-concept study demonstrates nanopore sequencing enables episignature detection. In addition, concurrent haplotyped genomic and epigenomic analyses leverage simultaneous detection of single nucleotide/structural variants, X-inactivation, and imprinting, consolidating a multi-step sequential process into a single diagnostic assay. [ABSTRACT FROM AUTHOR]

High-grade soft tissue sarcomas (STS) are a heterogeneous and aggressive set of cancers. Failure to respond anthracycline chemotherapy, standard first-line treatment, is associated with poor outcomes. We investigated the contribution of STS cancer stem cells (STS-CSCs) to doxorubicin resistance. We identified a positive correlation between CSC abundance and doxorubicin IC50. Utilizing patient-derived samples from five sarcoma subtypes we investigated if a common genetic signature across STS-CSCs could be targeted. We identified Enhancer of Zeste homolog 2 (EZH2), a member of the polycomb repressive complex 2 (PRC2) responsible for H3K27 methylation as being enriched in CSCs. EZH2 activity and a shared epigenetic profile was observed across subtypes and targeting of EZH2 ablated the STS-CSC population. Treatment of doxorubicin-resistant cell lines with tazemetostat resulted in a decrease in the STS-CSC population. These data confirm the presence of shared genetic programs across distinct subtypes of CSC-STS that can be therapeutically targeted. [ABSTRACT FROM AUTHOR]

Members of the piggyBac superfamily of DNA transposons are widely distributed in host genomes ranging from insects to mammals. The human genome has retained five piggyBac-derived genes as domesticated elements although they are no longer mobile. Here, we have investigated the transposition properties of piggyBat from Myotis lucifugus, the only known active mammalian DNA transposon, and show that its low activity in human cells is due to subterminal inhibitory DNA sequences. Activity can be dramatically improved by their removal, suggesting the existence of a mechanism for the suppression of transposon activity. The cryo-electron microscopy structure of the piggyBat transposase pre-synaptic complex showed an unexpected mode of DNA binding and recognition using C-terminal domains that are topologically different from those of the piggyBac transposase. Here we show that structure-based rational re-engineering of the transposase through the removal of putative phosphorylation sites and a changed domain organization - in combination with truncated transposon ends - results in a transposition system that is at least 100-fold more active than wild-type piggyBat. piggyBat is the only known DNA transposon active in mammals. Here the authors used cellular assays and 3D structure to show that sequences within the transposon ends restrict activity, and also how transposition activity can be substantially increased. [ABSTRACT FROM AUTHOR]

Background: Congenital anomalies (CAs) encompass a wide spectrum of structural and functional abnormalities during fetal development, commonly presenting at birth. Identifying the cause of CA is essential for accurate diagnosis and treatment. Using a target-gene approach, genetic variants could be found in certain CA patients. However, some patients were genetically undiagnosed; therefore, it is imperative to identify the causative variants from whole genome sequence (WGS) data of these patients. Results: An in-house pipeline utilizing DRAGEN-GATK-Hail was established for trio-based WGS data analysis (n = 18 undiagnosed CA patients and their parents) and thirty-five candidate variants, including SNV/Indel, CNV, and SV were identified. Among them, 10 variants of seven coding genes were selected as possible causal variants by variant pathogenicity, genotype–phenotype analysis, and a multidisciplinary team. Finally, functional validation of six genes including RYR3, NRXN1, FREM2, CSMD1, RARS1, and NOTCH1, revealed various phenotypes in zebrafish models that aligned with those observed in each patient. In addition to the above findings, eleven diagnostic variants initially discovered in a targeted-gene analysis from a previous study were also identified as diagnostic variants and the in-house pipeline demonstrated a significant advantage in accurately and efficiently identifying de novo variants (DNVs), compound heterozygous (CH), and homozygous variants. Conclusions: Taken together, the in-house pipeline established in this study provides a highly valuable diagnostic tool for the identification of potential candidate variants in patients with CA. Further research into the molecular mechanisms related to the development of CAs could shed light on the functional aspects of these genetic variations and contribute to the development of therapeutic drugs. [ABSTRACT FROM AUTHOR]

Background: PSEN1, PSEN2, and APP mutations cause Alzheimer's disease (AD) with an early age at onset (AAO) and progressive cognitive decline. PSEN1 mutations are more common and generally have an earlier AAO; however, certain PSEN1 mutations cause a later AAO, similar to those observed in PSEN2 and APP. Methods: We examined whether common disease endotypes exist across these mutations with a later AAO (~ 55 years) using hiPSC-derived neurons from familial Alzheimer's disease (FAD) patients harboring mutations in PSEN1A79V, PSEN2N141I, and APPV717I and mechanistically characterized by integrating RNA-seq and ATAC-seq. Results: We identified common disease endotypes, such as dedifferentiation, dysregulation of synaptic signaling, repression of mitochondrial function and metabolism, and inflammation. We ascertained the master transcriptional regulators associated with these endotypes, including REST, ASCL1, and ZIC family members (activation), and NRF1 (repression). Conclusions: FAD mutations share common regulatory changes within endotypes with varying severity, resulting in reversion to a less-differentiated state. The regulatory mechanisms described offer potential targets for therapeutic interventions. [ABSTRACT FROM AUTHOR]

Background: The study is to evaluate the efficacy and safety of combined anlotinib and EGFR-tyrosine kinase inhibitors (TKIs) in patients with advanced non-small cell lung cancer (NSCLC) who had gradual, oligo, or potential progression after previous EGFR-TKIs treatment. Methods: We conducted an open-label, single-arm, multicenter, phase II trial in China. Eligible patients were 18–75 years old with histologically or cytologically confirmed NSCLC who were EGFR mutation positive and showed gradual, oligo, or potential progression after EGFR-TKIs. Anlotinib (12 mg/day) was administered orally for 2 weeks and then off 1 week in a 3-week cycle. EGFR-TKIs were continue used. The primary endpoint was progression-free survival (PFS). The secondary endpoints included 6- and 12-month PFS rate, objective response rate (ORR), disease control rate (DCR), overall survival (OS) and safety. Results: From July 2019 to December 2022, 120 patients were enrolled. The median PFS (mPFS) was 9.1 months (95% CI 6.8–11.7). The PFS rates at 6 and 12 months was 68.5% and 38.8% respectively. For 86 patients with first-line 1st /2nd generation EGFR-TKIs, the mPFS was 9.2 months (95% CI 6.7–12.6). For 32 patients with first-line 3rd generation EGFR-TKIs, the mPFS was 10.3 months (95% CI 6.1–13.3). Overall ORR and DCR were 6.7% (95% CI 2.9–12.7) and 87.5% (95% CI 80.2–92.8), respectively. 52.5% of patients had grade 3 or higher treatment-emergent adverse events (TEAEs). Conclusion: Anlotinib in combination with continuation of EGFR-TKIs prolonged the clinical benefit of EGFR-TKIs, demonstrating favorable survival outcomes and manageable toxicity in NSCLC treated with EGFR-TKIs and had specific progression modes, such as gradual progression. Trial registration: NCT04007835. [ABSTRACT FROM AUTHOR]

Malaria has been a leading cause of death in human populations for centuries and remains a major public health challenge in African countries, especially affecting children. Among the five Plasmodium species infecting humans, Plasmodium falciparum is the most lethal. Ancient DNA research has provided key insights into the origins, evolution, and virulence of pathogens that affect humans. However, extensive screening of ancient skeletal remains for Plasmodium DNA has shown that such genomic material is rare, with no studies so far addressing potential intra-individual variability. Consequently, the pool of ancient mitochondrial DNA (mtDNA) or genomic sequences for P. falciparum is extremely limited, with fewer than 20 ancient sequences available for genetic analysis, and no complete P. falciparum mtDNA from Classical antiquity published to date. To investigate intra-individual diversity and genetic origins of P. falciparum from the Roman period, we generated 39 sequencing libraries from multiple teeth and two from the femur of a Roman malaria-infected individual. The results revealed considerable variability in P. falciparum recovery across different dental samples within the individual, while the femur samples showed no preservation of Plasmodium DNA. The reconstructed 43-fold P. falciparum mtDNA genome supports the hypothesis of an Indian origin for European P. falciparum and suggests mtDNA continuity in Europe over the past 2000 years. [ABSTRACT FROM AUTHOR]

The knowledge of diffusion mechanisms in materials is crucial for predicting their high-temperature performance and stability, yet accurately capturing the underlying physics like thermal effects remains challenging. In particular, the origin of the experimentally observed non-Arrhenius diffusion behavior has remained elusive, largely due to the lack of effective computational tools. Here we propose an efficient ab initio framework to compute the Gibbs energy of the transition state in vacancy-mediated diffusion including the relevant thermal excitations at the density-functional-theory level. With the aid of a bespoke machine-learning interatomic potential, the temperature-dependent vacancy formation and migration Gibbs energies of the prototype system body-centered cubic (BCC) tungsten are shown to be strongly affected by anharmonicity. This finding explains the physical origin of the experimentally observed non-Arrhenius behavior of tungsten self-diffusion. A remarkable agreement between the calculated and experimental temperature-dependent self-diffusivity and, in particular, its curvature is revealed. The proposed computational framework is robust and broadly applicable, as evidenced by first tests for a hexagonal close-packed (HCP) multicomponent high-entropy alloy. The successful applications underscore the attainability of an accurate ab initio diffusion database. Modelling of high-temperature diffusion requires accurate treatment of thermal effects. Here, the authors introduce a method to calculate Gibbs energies of diffusion transition states to uncover the mechanisms of non-Arrhenius diffusion behavior. [ABSTRACT FROM AUTHOR]

With the rapid development of society and economy and the acceleration of population aging, the global disease spectrum has undergone profound changes. Neurological diseases, especially brain functional disorders, have become major threats to human health. Neuromodulation technique, as a critical means to address this challenge, has opened new avenues for the treatment of brain functional disorders and has become one of the three innovative technologies in modern neurosurgery. This paper systematically elaborates on the concept, classification, and technological evolution of neuromodulation, focusing on the clinical application progress of invasive neuromodulation technique. It analyzes their efficacy and potential in the fields of movement disorders, epilepsy, pain, cognitive impairment, and neurorehabilitation. Additionally, it explores the integration trend between neuromodulation and brain-computer interface (BCI), pointing out that closed-loop neuromodulation has become an important component of BCI, providing new approaches for precise treatment and individualized modulation. Finally, it proposes future development directions in the field of neuromodulation, including expanding new indications, innovating target exploration models, updating treatment concepts, and conducting high - quality evidence - based medical research. Looking ahead, Chinese neuromodulation endeavors should seize current opportunities, achieve a leap from following to leading through continuous exploration and innovation, and make greater contributions to global brain health. [ABSTRACT FROM AUTHOR]

BACKGROUND: Millions worldwide are exposed to elevated levels of arsenic that significantly increase their risk of developing atherosclerosis, a pathology primarily driven by immune cells. While the impact of arsenic on immune cell populations in atherosclerotic plaques has been broadly characterized, cellular heterogeneity is a substantial barrier to in-depth examinations of the cellular dynamics for varying immune cell populations. OBJECTIVES: This study aimed to conduct single-cell multi-omics profiling of atherosclerotic plaques in apolipoprotein E knockout (ApoE–/– ) mice to elucidate transcriptomic and epigenetic changes in immune cells induced by arsenic exposure. METHODS: The ApoE–/– mice were fed a high-fat diet and were exposed to either 200 ppb arsenic in drinking water or a tap water control, and single-cell multi-omics profiling was performed on atherosclerotic plaque-resident immune cells. Transcriptomic and epigenetic changes in immune cells were analyzed within the same cell to understand the effects of arsenic exposure. RESULTS: Our data revealed that the transcriptional profile of macrophages from arsenic-exposed mice were significantly different from that of control mice and that differences were subtype specific and associated with cell–cell interaction and cell fates. Additionally, our data suggest that differences in arsenic-mediated changes in chromosome accessibility in arsenic-exposed mice were statistically more likely to be due to factors other than random variation compared to their effects on the transcriptome, revealing markers of arsenic exposure and potential targets for intervention. DISCUSSION: These findings in mice provide insights into how arsenic exposure impacts immune cell types in atherosclerosis, highlighting the importance of considering cellular heterogeneity in studying such effects. The identification of subtype-specific differences and potential intervention targets underscores the significance of understanding the molecular mechanisms underlying arsenic-induced atherosclerosis. Further research is warranted to validate these findings and explore therapeutic interventions targeting immune cell dysfunction in arsenic-exposed individuals. [ABSTRACT FROM AUTHOR]

The genus Capsella serves as a model for understanding speciation, hybridization, and genome evolution in plants. Here, we present a chromosome-scale genome assembly of Capsella orientalis , the maternal progenitor of a cosmopolitan allotetraploid C. bursa-pastoris. Using nanopore sequencing and data on chromatin contacts (Hi-C), we assembled the genome into eight pseudo-chromosomes with high contiguity, evidenced by a benchmarking universal single-copy orthologs (BUSCO) completeness score of 99.3%. Comparative analysis with C. rubella and C. bursa-pastoris revealed overall synteny, except for 2 Mb inversion on chromosome 4 of C. rubella. Comparative genome analysis highlighted the conservation of gene content and structural integrity in the C. orientalis -derived subgenome of C. bursa-pastoris , with the exception of a 1.8 Mb region absent in O subgenome but present in C. orientalis. The genome annotation includes 27,675 protein-coding genes, with most exhibiting one-to-one orthology with Arabidopsis thaliana. Notably, 2,155 genes showed no similarity to A. thaliana ones. These results establish a robust genomic resource for C. orientalis , facilitating future studies on polyploid evolution, gene regulation, and species divergence within Capsella. [ABSTRACT FROM AUTHOR]

The polar regions host a diverse array of moss species that have evolved to thrive in extreme environments. These mosses exhibit remarkable adaptations, including tolerance to freezing temperatures, desiccation, and ultraviolet radiation. Despite their ecological significance, genomic data on these organisms are still limited, impeding our understanding of their evolutionary history and adaptive mechanisms in the context of climate change. In this study, we present the first chromosome-scale genome assembly and annotation of the Arctic moss Ptychostomum knowltonii. The assembled genome is 408.8 Mb in size, anchored to 12 pseudochromosomes, with a scaffold N50 of 32.61 Mb. Repetitive elements account for 56.24% of the genome. The genome contains 28,014 protein-coding genes, with a BUSCO completeness of 96.20%. This genomic resource will enable future comparative genomic studies, enhancing our understanding of how polar mosses may respond to a warming climate and shedding light on their evolutionary trajectories in persistently extreme environments. [ABSTRACT FROM AUTHOR]

Background: The field of single cell technologies has rapidly advanced our comprehension of the human immune system, offering unprecedented insights into cellular heterogeneity and immune function. While cryopreserved peripheral blood mononuclear cell (PBMC) samples enable deep characterization of immune cells, challenges in clinical isolation and preservation limit their application in underserved communities with limited access to research facilities. We present CryoSCAPE (Cryopreservation for Scalable Cellular And Proteomic Exploration), a scalable method for immune studies of human PBMC with multi-omic single cell assays using direct cryopreservation of whole blood. Results: Comparative analyses of matched human PBMC from cryopreserved whole blood and density gradient isolation demonstrate the efficacy of this methodology in capturing cell proportions and molecular features. The method was then optimized and verified for high sample throughput using fixed single cell RNA sequencing and liquid handling automation with a single batch of 60 cryopreserved whole blood samples. Additionally, cryopreserved whole blood was demonstrated to be compatible with functional assays, enabling this sample preservation method for clinical research. Conclusions: The CryoSCAPE method, optimized for scalability and cost-effectiveness, allows for high-throughput single cell RNA sequencing and functional assays while minimizing sample handling challenges. Utilization of this method in the clinic has the potential to democratize access to single-cell assays and enhance our understanding of immune function across diverse populations. [ABSTRACT FROM AUTHOR]

Background/Aim: Bile tract cancer (BTC) is a malignant tumor with a poor prognosis. Recent studies have reported the heterogeneity of the genomic background and gene alterations in BTC, but its genetic heterogeneity and molecular profiles remain poorly understood. Whole-genome sequencing may enable the identification of novel actionable gene mutations involved in BTC carcinogenesis, malignant progression, and treatment resistance. Patients and Methods: We performed whole-genome sequencing of six BTC samples to elucidate its genetic heterogeneity and identify novel actionable gene mutations. Somatic mutations, structural variations, copy number alterations, and their associations with clinical factors were analyzed. Results: The average number of somatic mutations detected in each case was 53,705, with SNVs accounting for most of these mutations (85.02%). None of the 331 mutations related to BTC in The Cancer Genome Atlas (TCGA) database were found in the mutations identified in our study. A higher prevalence of gene mutations was observed in samples without vascular invasion than in those with vascular invasion. Several genes with differences in mutation accumulation between groups were identified, including ADAMTS7, AHNAK2, and CAPN10. Conclusion: Our study provides novel insights into the genomic landscape of BTC and highlights the potential of whole-genome sequencing analysis to identify actionable gene mutations and understand the molecular mechanisms underlying this malignancy. The high mutational burden, structural variations, and copy number alterations observed in BTC samples in this study underscore the genetic complexity and heterogeneity of this disease. [ABSTRACT FROM AUTHOR]

Sexual compatibility in the Basidiomycota is governed by genetic identity at one or two loci, resulting in compatibility systems called bipolar and tetrapolar. The loci are known as HD and P/R, encoding homeodomain transcription factors and pheromone precursors and receptors, respectively. Bipolarity is known to evolve either by linkage of the two loci or by loss of mating-type determination of either the HD or the P/R locus. The ancestor to basidiomycete fungi is thought to have been tetrapolar, and many transitions to bipolarity have been described in different lineages. In the diverse genus Marasmius (Agaricales), both compatibility systems are found, and the system has been shown to follow the infrageneric sections of the genus, suggesting a single origin of bipolarity. Here, we tested this hypothesis using a comprehensive phylogenetic framework and investigated the mode by which bipolarity has evolved in this group. We utilized available genomic data and marker sequences to investigate evolution of sexual compatibility in Marasmius and allied genera. By generating a concatenated multilocus phylogeny, we found support for a single transition to known bipolarity within Marasmius. Furthermore, utilizing genomic data of the bipolar species Marasmius oreades, we found that the HD and P/R loci likely have remained unlinked through this transition. By comparing nucleotide diversity at the HD and P/R loci in Ma. oreades, we show that the HD locus has retained high diversity, and thus likely the function of determining sexual identity, as similarly in other bipolar mushroom-forming fungi. Finally, we describe the genomic architecture of the MAT loci of species of both sexual compatibility systems in Marasmiaceae and related families. [ABSTRACT FROM AUTHOR]

Retroviruses are among the most extensively studied viral families, both historically and in contemporary research. They are primarily investigated in the fields of viral oncogenesis, reverse transcription mechanisms, and other infection-specific aspects. These include the integration of endogenous retroviruses (ERVs) into host genomes, a process widely utilized in genetic engineering, and the ongoing search for HIV/AIDS treatment. G-quadruplexes (G4) have emerged as potential therapeutic targets in antiviral therapy and have been identified in important regulatory regions of viral genomes. In this study, we examine the presence of potential G-quadruplex-forming sequences (PQS) across all currently available unique retroviral genomes. Given that these retroviral genomes typically consist of single-stranded RNA (ssRNA) molecules, we also investigated whether the localization of PQSs is strand-dependent. This is particularly relevant since antisense transcripts have been detected in HIV, and ERV integration into the host genome involves reverse transcription from genomic positive strand ssRNA to double-stranded DNA (dsDNA), implicating both strands in this process. We show that in most mammalian retroviruses, including human retroviruses, PQSs are significantly more prevalent on the negative (antisense) strand, with some notable exceptions such as HIV-1. In sharp contrast, avian retroviruses exhibit a higher prevalence of PQSs on the positive (sense) strand. [ABSTRACT FROM AUTHOR]

In mammalian and vertebrate genomes, the promoter regions of the gene and their distal enhancers may be located millions of base-pairs from each other, while a promoter may not interact with the closest enhancer. Since base-pair proximity is not a good indicator of these interactions, there is a significant body of work to develop methods for understanding Enhancer-Promoter Interactions (EPI) from genetic and epigenomic marks. Over the last decade, several machine learning and deep learning methods have reported increasingly higher accuracies for predicting EPI. Typically, these approaches perform analysis by randomly splitting the dataset of Enhancer-Promoter (EP) pairs into training and testing subsets followed by model training. However, the aforementioned random splitting inadvertently causes information leakage by assigning EP pairs from the same genomic region to both testing and training sets. As a result, it has been pointed out in the literature that the performance of EPI prediction algorithms is overestimated because of genomic region overlap among the training and testing parts of the data. Building on that, in this paper we propose to use a more thorough training and testing paradigm i.e., Leave-one-chromosome-out (LOCO) cross-validation for EPI prediction. LOCO has been used in other bioinformatics contexts and ensures that there is no genomic overlap between training and testing sets enabling more fair estimation of performance. We demonstrate that a deep learning algorithm which gives higher accuracies when trained and tested on random-splitting setting, drops drastically in performance under LOCO setting, showing overestimation of performance in previous literature. We also propose a novel hybrid multi-branch neural network architecture for EPI prediction. In particular, our architecture has one branch consisting of a deep neural network, while the other branch extracts traditional k-mer features derived from the nucleotide sequence. The two branches are later merged and the neural network is trained jointly to force the network to learn feature representations which are already not covered by k-mer features. We show that the hybrid architecture performs significantly better in a realistic and fair LOCO testing paradigm, demonstrating it can learn more general aspects of EP interactions instead of overfitting to genomic regions. Through this paper we are also releasing the LOCO splitting-based EPI dataset to encourage other research groups to benchmark their EPI algorithms using a consistent LOCO paradigm. Research data is available in this public repository: [ABSTRACT FROM AUTHOR]

Recently we characterized polymyositis in the Dutch Kooiker dog. The familial occurrence of the disease were suggestive of an inherited cause. Here we report the results of our molecular genetic investigation. A genome-wide association study of 33 cases and 106 controls indicated the involvement of a region on chromosome CFA19 (p = 4.7*10−10). Haplotype analysis indicated that the cases shared a 2.9 Mb region in the homozygous or the heterozygous state. Next Generation Sequencing of genomic DNA implicated a deletion of a 39 kb DNA fragment, located 10 kb upstream of the neighbouring interleukin genes IL21 and IL2. The frequency of the deletion allele was 0.81 in the available cases and 0.25 in a random sample of the Kooiker dog breed. Leukocytes of affected, untreated dogs that were homozygous for the deletion overexpress IL21 and IL2 upon stimulation with mitogens. We suggest that elements located 10–49 kb upstream of the IL21/IL2 locus play an important role in the regulation of the canine genes and that deletion of these elements is a risk factor for polymyositis in Kooiker dogs. Postulating causality, the penetrance of the disease phenotype was estimated at 10–20% for homozygous dogs and 0.5–2% for dogs that were heterozygous for the deletion. Our results suggest that distant variants upstream of IL21 could also be important for human autoimmune diseases that have been found to be associated with the IL21/IL2 chromosome region. Author summary: Polymyositis in the breed of so-called Kooiker dogs is an autoimmune mediated inflammatory disease of skeletal muscles. The dogs are presented with swallowing and locomotion problems and in general the outcome is unfavorable. An inherited cause was suspected because affected dogs were often closely related. A genome-wide screen showed that the cases shared DNA variants in the region of the interleukin genes IL21 and IL2, wich are regulators of the cellular immune response. By analysis of the DNA sequence it became evident that a fragment of 39,000 base pairs had been deleted from the shared region. The deletion is located in front of the IL21 gene. All but two affected dogs had one or two copies of the deletion. Kooiker dogs with two copies have a risk of 10–20% to develop polymyositis in their lifetime; for dogs with one copy the risk is only 0.5–2%. We suggest that the deletion disturbs the normal regulation of IL21 and/or IL2 expression. Our results could be important for the research of human autoimmune diseases that have been associated with the same gene region. [ABSTRACT FROM AUTHOR]

Rehmannia glutinosa, an extensively utilized Chinese herbal medicine, is highly valued for its medicinal properties. In this study, the complete mitochondrial genome (mitogenome) of R. glutinosa was sequenced and assembled for the first time. The mitogenome is 547,032 bp in length, with an overall GC content of 44.97%. The mitogenome contains 67 unique genes, comprising 43 protein-coding, three rRNA, and 21 tRNA genes, with six protein-coding and nine tRNA genes being chloroplast-derived. The phylogenetic analysis, based on the maximum-likelihood criterion, demonstrated that R. glutinosa is closely related to Aeginetia indica and Castilleja paramensis within the family Orobanchaceae. [ABSTRACT FROM AUTHOR]

The complete mitochondrial genome of the Lutjanus ehrenbergii was sequenced by Sanger platform. The circular mitogenome of L. ehrenbergii (16,512 bp) encoded the typical 37 genes, and one non-coding regions. All of the protein-encoding genes were located on the H chain except ND6. The nucleotide composition was A (28.04%), T (24.84%), C (30.89%) and G (16.23%). Phylogenetic analysis based on the 13PCGs sequences showed that L. ehrenbergii was closely related to the sister species of Lutjanus russellii and Lutjanus carponotatus. These studies provided important mitochondrial genome data of L. ehrenbergii, phylogenetic tree analysis revealed the position of L. ehrenbergii in Lutjaniformes. [ABSTRACT FROM AUTHOR]

The assembly of repressive heterochromatin in eukaryotic genomes is crucial for silencing lineage-inappropriate genes and repetitive DNA elements. Paradoxically, transcription of repetitive elements within constitutive heterochromatin domains is required for RNA-based mechanisms, such as the RNAi pathway, to target heterochromatin assembly proteins. However, the mechanism by which heterochromatic repeats are transcribed has been unclear. Using fission yeast, we show that the conserved trimeric transcription factor (TF) PhpCNF-Y complex can infiltrate constitutive heterochromatin via its histone-fold domains to transcribe repeat elements. PhpCNF-Y collaborates with a Zn-finger containing TF to bind repeat promoter regions with CCAAT boxes. Mutating either the TFs or the CCAAT binding site disrupts the transcription of heterochromatic repeats. Although repeat elements are transcribed from both strands, PhpCNF-Y-dependent transcripts originate from only one strand. These TF-driven transcripts contain multiple cryptic introns which are required for the generation of small interfering RNAs (siRNAs) via a mechanism involving the spliceosome and RNAi machinery. Our analyses show that siRNA production by this TF-mediated transcription pathway is critical for heterochromatin nucleation at target repeat loci. This study reveals a mechanism by which heterochromatic repeats are transcribed, initiating their own silencing by triggering a primary cascade that produces siRNAs necessary for heterochromatin nucleation. This study shows that the PhpCNF-Y transcription factor infiltrates heterochromatin to produce transcripts with cryptic introns that initiate RNAi-mediated heterochromatin assembly, investigating how heterochromatic repeats initiate their own silencing. [ABSTRACT FROM AUTHOR]

Nucleosome is the basic structural unit of the genome. During processes like DNA replication and gene transcription, the conformation of nucleosomes undergoes dynamic changes, including DNA unwrapping and rewrapping, as well as histone disassembly and assembly. However, the wrapping characteristics of nucleosomes across the entire genome, including region-specificity and their correlation with higher-order chromatin organization, remains to be studied. In this study, we investigate the wrapping length of DNA on nucleosomes across the whole genome using wrapping-seq. We discover that the chromatin of mouse ES cells forms Nucleosome Wrapping Domains (NRDs), which can also be observed in yeast and fly genomes. We find that the degree of nucleosome wrapping decreases after DNA replication and is promoted by transcription. Furthermore, we observe that nucleosome wrapping domains delineate Hi-C compartments and replication timing domains. In conclusion, we have unveiled a previously unrecognized domainization principle of the chromatin, encoded by nucleosome wrapping states. Nucleosome is the basic structural unit of the genome. Here, the authors find that the genome is segregated into domains with distinct nucleosome wrapping features, which coincident with Hi-C compartments and replication timing domains. [ABSTRACT FROM AUTHOR]

The mammalian early embryo development requires translation of maternal mRNA inherited from the oocyte. While poly(A) tail length influences mRNA translation efficiency during the oocyte-to-embryo transition (OET), molecular mechanisms regulating maternal RNA poly(A) tail length are not fully understood. In this study, we identified MARTRE, a previously uncharacterized protein family (MARTRE1-MARTRE6), as regulators expressed during mouse OET that modulate poly(A) tail length. MARTRE inhibits deadenylation through the direct interaction with the deadenylase CCR4-NOT, and ectopic expression of Martre stabilized mRNA by attenuating poly(A) tail shortening. Deletion of the Martre gene locus results in shortened poly(A) tails and decreased translation efficiency of actively translated mRNAs in mouse zygotes, but does not affect maternal mRNA decay. MARTRE proteins thus fine-tune maternal mRNA translation by negatively regulating the deadenylating activity of CCR4-NOT. Moreover, Martre knockout embryos show delayed 2-cell stage progression and compromised preimplantation development. Together, our findings highlight protection of long poly(A) tails from active deadenylation as an important mechanism to coordinate translation of maternal mRNA. This study identifies a family of protein, MARTRE, as regulators of poly(A) tail length in early mouse embryos. MARTREs inhibit CCR4-NOT complex-mediated deadenylation, ensuring efficient translation of maternal mRNA for embryo development. [ABSTRACT FROM AUTHOR]