Orthologous Genes Research Articles

Systematic ocular phenotyping of 8,707 knockout mouse lines identifies genes associated with abnormal corneal phenotypes

PurposeCorneal dysmorphologies (CDs) are typically classified as either regressive degenerative corneal dystrophies (CDtrs) or defective growth and differentiation-driven corneal dysplasias (CDyps). Both eye disorders have multifactorial etiologies. While previous work has elucidated many aspects of CDs, such as presenting symptoms, epidemiology, and pathophysiology, the genetic mechanisms remain incompletely understood. The purpose of this study was to analyze phenotype data from 8,707 knockout mouse lines to identify new genes associated with the development of CDs in humans.Methods8,707 knockout mouse lines phenotyped by the International Mouse Phenotyping Consortium were queried for genes associated with statistically significant (P < 0.0001) abnormal cornea morphology to identify candidate CD genes. Corneal abnormalities were investigated by histopathology. A literature search was used to determine the proportion of candidate genes previously associated with CDs in mice and humans. Phenotypes of human orthologues of mouse candidate genes were compared with known human CD genes to identify protein-protein interactions and molecular pathways using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), Protein Analysis Through Evolutionary Relationships (PANTHER), and Kyoto Encyclopedia of Genes and Genomes.ResultsAnalysis of data from 8,707 knockout mouse lines identified 213 candidate CD genes. Of these, 37 (17%) genes were previously known to be associated with CD, including 14 in the mouse, 16 in humans, and 7 in both. The remaining 176 (83%) genes have not been previously implicated in CD. We also searched publicly available RNAseq data and found that 131 of the total 213 (61.5%) were expressed in adult human corneal tissue. STRING analysis showed several interactions within and between candidate and established CD proteins. All cellular pathways of the established genes were found in the PANTHER analysis of the candidate genes. Several of the candidate genes were implicated in corneal disease, such as TGF-ß signaling. We also identified other possible underappreciated mechanisms relevant to the human cornea.ConclusionsWe identified 213 mouse genes that resulted in statistically significant abnormal corneal phenotypes in knockout mice, many of which have not previously been implicated in corneal pathology. Bioinformatic analyses implicated candidate genes in several signaling pathways which are potential therapeutic targets.

Comparative analysis of the WRKY gene family between Chimonanthus praecox and C. salicifolius.

Gene duplications provide evolutionary potentials for generating novel functions. Chimonanthus praecox and C. salicifolius are closely related species from Calycantaceae, Magnoliids. In this study, we compared the WRKY gene family from C. praecox and C. salicifolius, and predicted the potential gene function through gene expression patterns to explore the evolution of orthologous and paralogous gene pairs. A total of 73 and 85 WRKY genes were identified and analyzed from the whole genome sequencing of C. praecox and C. salicifolius. Based on the phylogenetic analysis, CpWRKY and CsWRKY genes were clustered into three groups (Group I、II、III) and 5 subgroups (Group IIa、IIb、IIc、IId、IIe). In C. praecox and C. salicifolius, we identified thirty-six and fifty-four pairs of WRKY segmental duplicated genes, respectively, along with two and three pairs of tandem duplicates, indicating that segmental duplication plays a crucial role in the evolution of Chimonanthus WRKY gene family. Most WRKY duplication gene pairs originated from segmental duplications before the first whole genome duplication (WGD), highlighting this period as a significant source of genetic diversity and functionality for the WRKY family. The analysis of WRKY gene expression levels suggests that CsWRKY18 and CsWRKY68 may promote the growth of the roots in C. salicifolius. Comparisons of expression profiles between species revealed that five orthologous gene pairs presented identical expression trends, indicating functional conservation and absence of neo-functionalization or sub-functionalization. However, most orthologous gene pairs exhibit differences in expression patterns, suggesting that they have undergone functional divergence. This functional differentiation may be due to the different selective pressures faced by C. praecox and C. salicifolius during their speciation processes. This study provided detailed information on the WRKY gene family from C. praecox and C. salicifolius, and a new insight for studying gene duplication and function evolution.

Evolutionary lineage-specific genomic imprinting at the ZNF791 locus.

Genomic imprinting is an epigenetic process that results in parent-of-origin effects on mammalian development and growth. Research on genomic imprinting in domesticated animals has lagged due to a primary focus on orthologs of mouse and human imprinted genes. This emphasis has limited the discovery of imprinted genes specific to livestock. To identify genomic imprinting in pigs, we generated parthenogenetic porcine embryos alongside biparental normal embryos, and then performed whole-genome bisulfite sequencing and RNA sequencing on these samples. In our analyses, we discovered a maternally methylated differentially methylated region within the orthologous ZNF791 locus in pigs. Additionally, we identified both a major imprinted isoform of the ZNF791-like gene and an unannotated antisense transcript that has not been previously annotated. Importantly, our comparative analyses of the orthologous ZNF791 gene in various eutherian mammals, including humans, non-human primates, rodents, artiodactyls, and dogs, revealed that this gene is subjected to genomic imprinting exclusively in domesticated animals, thereby highlighting lineage-specific imprinting. Furthermore, we explored the potential mechanisms behind the establishment of maternal DNA methylation imprints in porcine and bovine oocytes, supporting the notion that integration of transposable elements, active transcription, and histone modification may collectively contribute to the methylation of embedded intragenic CpG island promoters. Our findings convey fundamental insights into molecular and evolutionary aspects of livestock species-specific genomic imprinting and provide critical agricultural implications.

Adaptive evolution of stress response genes in parasites aligns with host niche diversity

BackgroundStress responses are key the survival of parasites and, consequently, also the evolutionary success of these organisms. Despite this importance, our understanding of the evolution of molecular pathways dealing with environmental stressors in parasitic animals remains limited. Here, we tested the link between adaptive evolution of parasite stress response genes and their ecological diversity and species richness. We comparatively investigated antioxidant, heat shock, osmoregulatory, and behaviour-related genes (foraging) in two model parasitic flatworm lineages with contrasting ecological diversity, Cichlidogyrus and Kapentagyrus (Platyhelminthes: Monopisthocotyla), through whole-genome sequencing of 11 species followed by in silico exon bait capture as well as phylogenetic and codon analyses.ResultsWe assembled the sequences of 48 stress-related genes and report the first foraging (For) gene orthologs in flatworms. We found duplications of heat shock (Hsp) and oxidative stress genes in Cichlidogyrus compared to Kapentagyrus. We also observed positive selection patterns in genes related to mitochondrial protein import (Hsp) and behaviour (For) in species of Cichlidogyrus infecting East African cichlids—a host lineage under adaptive radiation. These patterns are consistent with a potential adaptation linked to a co-radiation of these parasites and their hosts. Additionally, the absence of cytochrome P450 and kappa and sigma-class glutathione S-transferases in monogenean flatworms is reported, genes considered essential for metazoan life.ConclusionsThis study potentially identifies the first molecular function linked to a flatworm radiation. Furthermore, the observed gene duplications and positive selection indicate the potentially important role of stress responses for the ecological adaptation of parasite species.

Potential of emodepside for vector-borne disease control

BackgroundEmodepside is an anthelmintic used in veterinary medicine that is currently under investigation in human clinical trials for the treatment of soil-transmitted helminths and possibly Onchocerca volvulus. Emodepside targets the calcium-activated voltage-gated potassium slowpoke 1 (SLO-1) channels of presynaptic nerves of pharynx and body wall muscle cells of nematodes leading to paralysis, reduced locomotion and egg laying, starvation, and death. Emodepside also has activity against Drosophila melanogaster SLO-1 channels. Orthologous SLO-1 genes are present in Anopheles gambiae and Aedes aegypti, suggesting that emodepside may have activity against mosquitoes.MethodsBoth Anopheles dirus and Ae. aegypti were blood-fed emodepside across a range of concentrations (1–10,000 nM) and mosquito survival was monitored for 10 days. Co-feeding experiments were also performed with An. dirus blood fed ivermectin at the concentrations that kills 25% (LC25) and 50% (LC50) of mosquitoes with and without emodepside at clinical peak concentration in humans (Cmax) and five times the Cmax, and mosquito survival was monitored for 10 days.ResultsEmodepside had weak mosquito-lethal effects in An. dirus but none observed in Ae. aegypti at the concentrations evaluated. The An. dirus emodepside LC50 was 4,623 [4,159–5,066] ng/ml which is > 100-fold greater than the peak concentrations seen in human. The ivermectin and emodepside co-feed experiment with An. dirus did not indicate any altered effect of ivermectin on mosquito survival when emodepside co-fed at human Cmax or five times that of the human Cmax.ConclusionsEmodepside was not lethal to An. dirus at human-relevant concentrations and had no effect on Ae. aegypti survival. Thus, mass distribution of emodepside does not appear to be a potential tool for vector-borne disease control. Emodepside induced mortality in An. dirus does suggest that the SLO-1 channel could be a potential target for novel vector control and may warrant further investigation.

A sweeping view of avian mycoplasmas biology drawn from comparative genomic analyses

BackgroundAvian mycoplasmas are small bacteria associated with several pathogenic conditions in many wild and poultry bird species. Extensive genomic data are available for many avian mycoplasmas, yet no comparative studies focusing on this group of mycoplasmas have been undertaken so far.ResultsHere, based on the comparison of forty avian mycoplasma genomes belonging to ten different species, we provide insightful information on the phylogeny, pan/core genome, energetic metabolism, and virulence of these avian pathogens. Analyses disclosed considerable inter- and intra-species genomic variabilities, with genome sizes that can vary by twice as much. Phylogenetic analysis based on concatenated orthologous genes revealed that avian mycoplasmas fell into either Hominis or Pneumoniae groups within the Mollicutes and could split into various clusters. No host co-evolution of avian mycoplasmas can be inferred from the proposed phylogenetic scheme. With 3,237 different gene clusters, the avian mycoplasma group under study proved diverse enough to have an open pan genome. However, a set of 150 gene clusters was found to be shared between all avian mycoplasmas, which is likely encoding essential functions. Comparison of energy metabolism pathways showed that avian mycoplasmas rely on various sources of energy. Superposition between phylogenetic and energy metabolism groups revealed that the glycolytic mycoplasmas belong to two distinct phylogenetic groups (Hominis and Pneumoniae), while all the arginine-utilizing mycoplasmas belong only to Hominis group. This can stand for different evolutionary strategies followed by avian mycoplasmas and further emphasizes the diversity within this group. Virulence determinants survey showed that the involved gene arsenals vary significantly within and between species, and could even be found in species often reported apathogenic. Immunoglobulin-blocking proteins were detected in almost all avian mycoplasmas. Although these systems are not exclusive to this group, they seem to present some particular features making them unique among mycoplasmas.ConclusionThis comparative genomic study uncovered the significant variable nature of avian mycoplasmas, furthering our knowledge on their biological attributes and evoking new hallmarks.

Comparative Genomics Reveals Evidence of the Genome Reduction and Metabolic Potentials of Aliineobacillus hadale Isolated from Challenger Deep Sediment of the Mariana Trench.

Hadal zones account for the deepest 45% of oceanic depth range and play an important role in ocean biogeochemical cycles. As the least-explored aquatic habitat on earth, further investigation is still required to fully elucidate the microbial taxonomy, ecological significance, metabolic diversity, and adaptation in hadal environments. In this study, a novel strain Lsc_1132T was isolated from sediment of the Mariana Trench at 10,954 m in depth. Strain Lsc_1132T contains heterogenous 16S rRNA genes, exhibiting the highest sequence similarities to the type strains of Neobacillus drentensis LMG 21831T, Neobacillus dielmonensis, Neobacillus drentensis NBRC 102427T, Neobacillus rhizosphaerae, and Neobacillus soli NBRC 102451T, with a range of 98.60-99.10% identity. The highest average nucleotide identity (ANI), the highest digital DNA-DNA hybridization (DDH) values, and the average amino acid identity (AAI) with Neobacillus sp. PS3-40 reached 73.5%, 21.4%, and 75.54%, respectively. The major cellular fatty acids of strain Lsc_1132T included iso-C15:0, Summed Feature 3 (C16:1ω6c and/or C16:1ω7c), iso-C17:0, anteiso-C15:0, and iso-C17:1ω5c. The respiratory quinone of strains Lsc_1132T was MK-7. The G + C content of the genomic DNA was 40.9%. Based on the GTDB taxonomy and phenotypic data, strain Lsc_1132T could represent a novel species of a novel genus, proposed as Aliineobacillus hadale gen. nov. sp. nov. (type strain Lsc_1132T = MCCC 1K09620T). Metabolically, strain Lsc_1132T demonstrates a robust carbohydrate metabolism with many strain-specific sugar transporters. It also has a remarkable capacity for metabolizing amino acids and carboxylic acids. Genomic analysis reveals a streamlined genome in the organism, characterized by a significant loss of orthologous genes, including those involved in cytochrome c synthesis, aromatic compound degradation, and polyhydroxybutyrate (PHB) synthesis, which suggests its adaptation to low oxygen levels and oligotrophic conditions through alternative metabolic pathways. In addition, the reduced number of paralogous genes in strain Lsc_1132T, together with its high protein-coding gene density, may further contribute to streamlining its genome and enhancing its genomic efficiency. This research expands our knowledge of hadal microorganisms and their metabolic strategies for surviving in extreme deep-sea environments.

Fusion, fission, and scrambling of the bilaterian genome in Bryozoa.

Groups of orthologous genes are commonly found together on the same chromosome over vast evolutionary distances. This extensive physical gene linkage, known as macrosynteny, is seen between bilaterian phyla as divergent as Chordata, Echinodermata, Mollusca, and Nemertea. Here, we report a unique pattern of genome evolution in Bryozoa, an understudied phylum of colonial invertebrates. Using comparative genomics, we reconstruct the chromosomal evolutionary history of five bryozoans. Multiple ancient chromosome fusions followed by gene mixing led to the near-complete loss of bilaterian linkage groups in the ancestor of extant bryozoans. A second wave of rearrangements, including chromosome fission, then occurred independently in two bryozoan classes, further scrambling bryozoan genomes. We also discover at least five derived chromosomal fusion events shared between bryozoans and brachiopods, supporting the traditional but highly debated Lophophorata hypothesis and suggesting macrosynteny to be a potentially powerful source of phylogenetic information. Finally, we show that genome rearrangements led to the dispersion of genes from bryozoan Hox clusters onto multiple chromosomes. Our findings demonstrate that the canonical bilaterian genome structure has been lost across all studied representatives of an entire phylum, and reveal that linkage group fission can occur very frequently in specific lineages.

A conserved fungal Knr4/Smi1 protein is crucial for maintaining cell wall stress tolerance and host plant pathogenesis.

Filamentous plant pathogenic fungi pose significant threats to global food security, particularly through diseases like Fusarium Head Blight (FHB) and Septoria Tritici Blotch (STB) which affects cereals. With mounting challenges in fungal control and increasing restrictions on fungicide use due to environmental concerns, there is an urgent need for innovative control strategies. Here, we present a comprehensive analysis of the stage-specific infection process of Fusarium graminearum in wheat spikes by generating a dual weighted gene co-expression network (WGCN). Notably, the network contained a mycotoxin-enriched fungal module (F12) that exhibited a significant correlation with a detoxification gene-enriched wheat module (W12). This correlation in gene expression was validated through quantitative PCR. By examining a fungal module with genes highly expressed during early symptomless infection that was correlated to a wheat module enriched in oxidative stress genes, we identified a gene encoding FgKnr4, a protein containing a Knr4/Smi1 disordered domain. Through comprehensive analysis, we confirmed the pivotal role of FgKnr4 in various biological processes, including oxidative stress tolerance, cell cycle stress tolerance, morphogenesis, growth, and pathogenicity. Further studies confirmed the observed phenotypes are partially due to the involvement of FgKnr4 in regulating the fungal cell wall integrity pathway by modulating the phosphorylation of the MAP-kinase MGV1. Orthologues of the FgKnr4 gene are widespread across the fungal kingdom but are absent in other Eukaryotes, suggesting the protein has potential as a promising intervention target. Encouragingly, the restricted growth and highly reduced virulence phenotypes observed for ΔFgknr4 were replicated upon deletion of the orthologous gene in the wheat fungal pathogen Zymoseptoria tritici. Overall, this study demonstrates the utility of an integrated network-level analytical approach to pinpoint genes of high interest to pathogenesis and disease control.

Genome sequencing of Plasmodium malariae identifies continental segregation and mutations associated with reduced pyrimethamine susceptibility

AbstractPlasmodium malariae parasites are widely observed across the tropics and sub-tropics. This slow-growing species, known to maintain chronic asymptomatic infections, has been associated with reduced antimalarial susceptibility. We analyse 251 P. malariae genomes from 28 countries, and leveraging 131,601 high-quality SNPs, demonstrate segregation of African and Asian isolates. Signals of recent evolutionary selection were identified in genes encoding putative surface proteins (pmmsp1) and putative erythrocyte invasion proteins (pmdpap3, pmrbp2, pmnif4). Amino acid substitutions were identified in orthologs of genes associated with antimalarial susceptibility including 2 amino acid substitutions in pmdhfr aligning with pyrimethamine resistance mutations in P. falciparum. Additionally, we characterise pmdhfr mutation F57L and demonstrate its involvement in reduced susceptibility to pyrimethamine in an in vitro parasite assay. We validate CRISPR-Cas9 mediated ortholog replacement in P. knowlesi parasites to determine the function of pmdhfr mutations and demonstrate that circulating pmdhfr genotypes are less susceptible to pyrimethamine.

Transcriptome-Wide Identification of miRNAs and Their Targets During Riboflavin-Promoted Dormancy Release in Lilium ‘Siberia’

Dormancy release is an important process for improving the quality of cut-flower lily production and promoting the factory production of lily bulbs. However, the regulatory mechanisms of microRNAs (miRNAs) and their target genes during the dormancy release of lily remain elusive. Anatomy, transcriptomic, molecular biology, and transient transformation techniques involving subcellular localization were applied in our study. There were significant results showing that 0.1 mM riboflavin promoted dormancy release and floral bud differentiation and influenced the flowering time of the Lilium ‘Siberia’. Moreover, some differentially expressed miRNAs and their targets (miR395-y: LoAPS1, miR529-z: LoSPL14, miR396-y: LoCFDP1, miR1863-z: LoFBA3, miR399-y: LoDIT1, and miR11525-z: Lopgm) were identified and predicted. Exogenous riboflavin may activate primary metabolic processes and promote dormancy release in Lilium ‘Siberia’ bulbs. Furthermore, riboflavin upregulated genes related to the riboflavin pathway, H3K4me3 methylation, dormancy control, and the flowering pathway and downregulated dormancy maintenance genes. Moreover, riboflavin promoted endogenous riboflavin and acetyl-CoA accumulation. LoPurple acid phosphatase17 (LoPAP17), a pivotal gene of the riboflavin metabolism pathway, was subsequently cloned. LoPAP17 was most closely related to the orthologous genes of Acorus calamus, Asparagus officinalis, and Musa acuminata. The LoPAP17 protein was subcellularly located in the nucleus. Our study revealed that miRNAs and their target genes might regulate the primary metabolic pathway, promote the accumulation of endogenous riboflavin and acetyl-CoA, and affect protein acetylation during the riboflavin-promoted release of dormancy and flower bud differentiation in the Lilium Oriental hybrid ‘Siberia’.

Model Organisms in Aging Research: Evolution of Database Annotation and Ortholog Discovery.

This study aims to analyze the exploration degree of popular model organisms by utilizing annotations from the UniProtKB (Swiss-Prot) knowledge base. The research focuses on understanding the genomic and post-genomic data of various organisms, particularly in relation to aging as an integral model for studying the molecular mechanisms underlying pathological processes and physiological states. Having characterized the organisms by selected parameters (numbers of gene splice variants, post-translational modifications, etc.) using previously developed information models, we calculated proteome sizes: the number of possible proteoforms for each species. Our analysis also involved searching for orthologs of human aging genes within these model species. Our findings indicate that genomic and post-genomic data for more primitive species, such as bacteria and fungi, are more comprehensively characterized compared to other organisms. This is attributed to their experimental accessibility and simplicity. Additionally, we discovered that the genomes of the most studied model organisms allow for a detailed analysis of the aging process, revealing a greater number of orthologous genes related to aging. The results highlight the importance of annotating the genomes of less-studied species to identify orthologs of marker genes associated with complex physiological processes, including aging. Species that potentially possess unique traits associated with longevity and resilience to age-related changes require comprehensive genomic studies.

Unraveling the Mysteries of Sleep: Exploring Phylogenomic Sleep Signals in the Recently Characterized Archaeal Phylum Lokiarchaeota near Loki's Castle.

Sleep is a universally conserved behavior whose origin and evolutionary purpose are uncertain. Using phylogenomics, this article investigates the evolutionary foundations of sleep from a never before used perspective. More specifically, it identifies orthologs of human sleep-related genes in the Lokiarchaeota of the Asgard superphylum and examines their functional role. Our findings indicate that a conserved suite of genes associated with energy metabolism and cellular repair is involved, thus suggesting that sleep plays a primordial role in cellular maintenance. The data cited lend credence to the idea that sleep improves organismal fitness across evolutionary time by acting as a restorative process. Notably, our approach demonstrates that phylogenomics is more useful than standard phylogenetics for clarifying common evolutionary traits. By offering insight into the evolutionary history of sleep and putting forth a novel model framework for sleep research across taxa, these findings contribute to our growing understanding of the molecular foundation of sleep. This study lays the groundwork for further investigations into the importance of sleep in various organisms. Such investigations could have consequences for improving human health and more generally could provide a deeper comprehension of the fundamental processes of life.

Transcriptional Reprogramming and Allelic Variation in Pleiotropic QTL Regulates Days to Flowering and Growth Habit in Pigeonpea.

The present study investigated the linkage between days to flowering (DTF) and growth habit (GH) in pigeonpea using QTL mapping, QTL-seq, and GWAS approaches. The linkage map developed here is the largest to date, spanning 1825.56 cM with 7987 SNP markers. In total, eight and four QTLs were mapped for DTF and GH, respectively, harbouring 78 pigeonpea orthologs of Arabidopsis flowering time genes. Corroboratively, QTL-seq analysis identified a single linked QTL for both traits on chromosome 3, possessing 15 genes bearing genic variants. Together, these 91 genes were clustered primarily into autonomous, photoperiod, and epigenetic pathways. Further, we identified 39 associations for DTF and 111 associations for GH through GWAS in the QTL regions. Of these, nine associations were consistent and constituted nine haplotypes (five late, two early, one each for super-early and medium duration). The involvement of multiple genes explained the range of allelic effects and the presence of multiple LD blocks. Further, the linked QTL on chromosome 3 was fine-mapped to the 0.24-Mb region with an LOD score of 8.56, explaining 36.47% of the phenotypic variance. We identified a 10-bp deletion in the first exon of TFL1 gene of the ICPL 20338 variety, which may affect its interaction with the Apetala1 and Leafy genes, resulting in determinate GH and early flowering. Further, the genic marker developed for the deletion in the TFL1 gene could be utilized as a foreground marker in marker-assisted breeding programmes to develop early-flowering pigeonpea varieties.

Gone with the Species: From Gene Loss to Gene Extinction.

Vertebrae protein-coding genes exhibit remarkable diversity and are organized into many gene families. These gene families have emerged through various gene duplication events, the most prominent being the two rounds of whole-genome duplication (WGD). The current research project analyzed a unique class of genes called "singletons". Notably, we introduce the concept of "super-singletons": genes that stand as the last representatives of their ancestral families and the sole representatives of their genetic makeup with no ortholog in any other species. We used the Ensembl/Biomart pipeline to identify duplicated and unduplicated protein-coding genes in different vertebrate species and found orthologs of human genes. We showed the frequency of duplicated genes and singletons, demonstrating that singletons are more vulnerable to evolutionary loss than duplicated genes. Additionally, we found that contractions in vertebrate gene families are more prevalent than expansion. Our study provides insight into the evolution of gene families and presents a novel scenario where the extinction of species would lead to the extinction of a gene, ultimately shifting the narrative from the impact of genetics on species extinction to the extinction of genes.

Girdin deficiency causes developmental and epileptic encephalopathy with hippocampal sclerosis and interneuronopathy.

Loss-of-function mutations in the GIRDIN/CCDC88A gene cause developmental epileptic encephalopathy (DEE) in humans. However, its pathogenesis is largely unknown. Global knockout mice of the corresponding orthologous gene (gKOs) have a preweaning lethal phenotype with growth failure, preventing longitudinal analysis. We aimed to overcome this lethality and elucidate DEE pathogenesis. We developed a novel lifelong feeding regimen (NLFR), which consists of providing mash food from postnatal day 14 (P14) until weaning (P28), followed by agar-bound food exclusively after weaning. Videography, electroencephalography (EEG), and histological analyses were performed. Conditional Girdin/Ccdc88a knockout mice (cKOs) of variable lineages (Nestin, Emx1, or Nkx2-1) were generated to identify the region responsible for epilepsy. Under the NLFR, gKOs survived beyond 1 year and displayed fully penetrant, robust epileptic phenotypes, including early-onset (P22.3 in average) generalized tonic-clonic seizures (GTCSs) (averaging eight per day), which were completely synchronized with fast rhythms on EEG, frequent interictal electroencephalographic spikes (averaging 430 per hour), and progressive deformation of visceral organs. In addition, gKOs had absence seizures, which were not always time-locked to frequent spike waves on EEG. The frequent GTCSs and interictal spikes in gKOs were suppressed by known antiepileptic drugs. Histologically, bilateral hippocampi in gKOs exhibited congenital cornu-ammonis splitting, granule cell dispersion, and astrogliosis. Furthermore, analysis of conditional knockouts using multiple Cre-deleters identified a defect in the delivery of interneuron precursors from the medial ganglionic eminence into the hippocampal primordium during embryogenesis as a major cause of epileptogenesis. These findings give rise to a new approach of lifelong caregiving to overcome the problem of preweaning lethality in animal models. We propose a useful model for studying DEE with hippocampal sclerosis and interneuronopathy. gKOs with NLFR combine the contradictory properties of robust epileptic phenotypes and long-term survivability, which can be used to investigate spontaneous epileptic wave propagation and therapeutic intervention in hippocampal sclerosis.

Identification of dehydrin family genes in three Medicago species and insights into their tolerant mechanism to salt stress.

All ten dehydrin genes from three Medicago species are responsive to different kinds of abiotic stress, and CAS31 confers transgenic plants salt tolerance by down-regulating HKT1 expression. Dehydrins are protective proteins playing crucial roles in the tolerance of plants to abiotic stresses. However, a full-scale and systemic analysis of total dehydrin genes in Medicago at the genome level is still lacking. In this study, we identified ten dehydrin genes from three Medicago species (M. truncatula, M. ruthenica, and M. sativa), categorizing the coding proteins into four types. Genome collinearity analysis among the three Medicago species revealed six orthologous gene pairs. Promoter regions of dehydrin genes contained various phytohormone- and stress-related cis-elements, and transcriptome analysis showed up-regulation of all ten dehydrin genes under different stress conditions. Transformation of dehydrin gene CAS31 increased the tolerance of transgenic seedlings compared with wild-type seedlings under salt stress. Our study demonstrated that transgenic seedlings maintained the more chlorophyll, accumulated more proline and less hydrogen peroxide and malondialdehyde than wild-type seedlings under salt stress. Further study revealed that CAS31 reduced Na+ accumulation by down-regulating HKT1 expression under salt stress. These findings enhance our understanding of the dehydrin gene family in three Medicago species and provide insights into their mechanisms of tolerance.

Large-scale CRISPR/Cas9 deletions within the WFDC gene cluster uncover gene functionality and critical roles in mammalian reproduction

Despite 96 million years of evolution separating humans and rodents, 11 closely related reproductive tract-specific genes in humans-SPINT3, WFDC6, EPPIN, WFDC8, WFDC9, WFDC10A, WFDC11, WFDC10B, WFDC13, SPINT4, and WFDC3-and the 13 reproductive tract-specific orthologous genes in mice, form highly conserved syntenic gene clusters indicative of conserved, combined critical functions. Further, despite significant progress toward a nonhormonal male contraceptive targeting the protein encoded by one of these genes, epididymal peptidase inhibitor (EPPIN), and associations found between mutations in EPPIN and an increased risk of male infertility, neither EPPIN nor any closely related whey acidic protein four-disulfide core (WFDC) gene have been explored functionally. To clarify the involvement of WFDC genes in male fertility, we strategically used CRISPR/Cas9 to generate mice lacking 13, 10, 5, or 4 genes within the cluster and demonstrated that males with deletions of 13, 10, or 4 genes (Wfdc6a, Eppin, Wfdc8, and Wfdc6a) were sterile due to an arrest in spermatogenesis, preventing formation beyond round spermatids. In contrast, the five gene knockout (KO) males (lacking Wfdc16, Wfdc9, Wfdc10, Wfdc11, and Wfdc13), despite normal spermatogenesis and sperm counts, were infertile due to defects in sperm motility and increased sperm death. Similarly to our previously reported Spint3 single gene KO, Wfdc3 single KO mice were fertile with no obvious reproductive phenotype. Our KO mouse studies to explore the entire WFDC locus of closely related genes have clarified the functional requirements of WFDC locus genes in different aspects of male fertility. Our research has implications for improving clinical diagnoses of male infertility and identifying additional targets for nonhormonal male contraception.

Orthrus: Towards Evolutionary and Functional RNA Foundation Models.

In the face of rapidly accumulating genomic data, our ability to accurately pre-dict key mature RNA properties that underlie transcript function and regulation remains limited. Pre-trained genomic foundation models offer an avenue to adapt learned RNA representations to biological prediction tasks. However, existing genomic foundation models are trained using strategies borrowed from textual or visual domains that do not leverage biological domain knowledge. Here, we intro-duce Orthrus, a Mamba-based mature RNA foundation model pre-trained using a novel self-supervised contrastive learning objective with biological augmentations. Orthrus is trained by maximizing embedding similarity between curated pairs of RNA transcripts, where pairs are formed from splice isoforms of 10 model organ-isms and transcripts from orthologous genes in 400+ mammalian species from the Zoonomia Project. This training objective results in a latent representation that clusters RNA sequences with functional and evolutionary similarities. We find that the generalized mature RNA isoform representations learned by Orthrus significantly outperform existing genomic foundation models on five mRNA prop-erty prediction tasks, and requires only a fraction of fine-tuning data to do so. Finally, we show that Orthrus is capable of capturing divergent biological function of individual transcript isoforms.

Identification and Evolution Analysis of the Genes Involved in the 20-Hydroxyecdysone Metabolism in the Mud Crab, Scylla paramamosain: A Preliminary Study.

20-Hydroxyecdysone (20E) is the most ubiquitous ecdysteroid (Ecd) and plays critical roles during the life cycle of arthropods. To elucidate the metabolism pathway of 20E in the economically important species, Scylla paramamosain, we conducted a comprehensive exploration of the genes involved in the 20E metabolism pathway. A comprehensive exploration of genes involved in the 20E metabolism pathway was conducted, including gene annotation, local blast using the Drosophila ortholog as query, and TreeFam ortholog genes identification. Bioinformatics and expression profiling of the identified genes were performed to assess their roles in the 20E metabolism of green mud crabs. This experiment indicated that, except for CYP306a1 and CYP314a1, all other ortholog genes involved in the Drosophila 20E metabolism can be found in the mud crab, suggesting that the function of these two genes might be replaced by other CYP genes or the "active" Ecd in mud crabs was not the 20E. All genes had the typical features of each gene family, clustered with the specific clade in the phylogenetic trees. In addition, all the identified genes had the highest expression level in the Y-organ, and sex-biased gene expression was observed in these genes. This study provided some valuable insights into the metabolism and diversity of ecdysteroids in crustaceans.