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Utilizing evolutionary conservation to detect deleterious mutations and improve genomic prediction in cassava.

INTRODUCTION: Cassava (Manihot esculenta) is an annual root crop which provides the major source of calories for over half a billion people around the world. Since its domestication ~10,000 years ago, cassava has been largely clonally propagated through stem cuttings. Minimal sexual recombination has led to an accumulation of deleterious mutations made evident by heavy inbreeding depression. METHODS: To locate and characterize these deleterious mutations, and to measure selection pressure across the cassava genome, we aligned 52 related Euphorbiaceae and other related species representing millions of years of evolution. With single base-pair resolution of genetic conservation, we used protein structure models, amino acid impact, and evolutionary conservation across the Euphorbiaceae to estimate evolutionary constraint. With known deleterious mutations, we aimed to improve genomic evaluations of plant performance through genomic prediction. We first tested this hypothesis through simulation utilizing multi-kernel GBLUP to predict simulated phenotypes across separate populations of cassava. RESULTS: Simulations showed a sizable increase of prediction accuracy when incorporating functional variants in the model when the trait was determined by<100 quantitative trait loci (QTL). Utilizing deleterious mutations and functional weights informed through evolutionary conservation, we saw improvements in genomic prediction accuracy that were dependent on trait and prediction. CONCLUSION: We showed the potential for using evolutionary information to track functional variation across the genome, in order to improve whole genome trait prediction. We anticipate that continued work to improve genotype accuracy and deleterious mutation assessment will lead to improved genomic assessments of cassava clones.

cassava (Manihot esculenta)

Evolutionary conservation of heat shock proteins in Blattodea and their roles in wing morphogenesis and ovarian development of Blattella germanica.

Heat shock proteins (Hsps) are essential molecular chaperones for protein homeostasis and stress responses. However, the Hsp repertoires and functions in Blattodea remain underexplored. Our genome-scale survey of nine Blattodea species revealed 37-46 conserved Hsp90, Hsp70, and DNAJ (Hsp40) genes, with DNAJ the most abundant and Hsp90 the least. Phylogenetic analysis confirmed the evolutionary conservation of three Hsp90, seven Hsp70, and 29 DNAJ subclades in Blattodea. Selection pressure analysis revealed predominant purifying selection (dN/dS&#xa0;&#x226a;&#xa0;1) across lineages, strongest in DNAJ and highest in Hsp90 conservation. In Blattella germanica, expression of six representative BgHsp genes progressively increased during development, peaking in fifth-instar nymphs. Tissue expression profiling revealed that BgHspA1-2/3/4 were predominantly expressed in legs, BgDNAJB5 and BgHsp90AB1-2 were enriched in the fat body, and BgHsp90AB1 was highly expressed in the head. dsRNA injection targeting conserved Hsp gene regions achieved 61.9-94.1% knockdown of all six target genes. RNAi knockdown of six BgHsp genes disrupted wing morphogenesis, causing distinct phenotypes: wing whitening (56.7%, dsBgHspA1-4), unequal length (66.7%, dsBgHspA1-3; 76.7%, dsBgDNAJB5), and wing wrinkling (70%, dsBgHspA1-2; 63.3%, dsBgHsp90AB1; 76.7%, dsBgHsp90AB1-2). During ovarian formation, the developmental delay was most severe in the dsBgHsp90AB1 group, moderate in the dsBgHsp90AB1-2 and dsBgHspA1-2/3/4 groups, and weakest in the dsBgDNAJB5 group. Besides, knockdown significantly downregulated key developmental genes (apterous-a, nubbin, scalloped, ultrabithorax, wingless, and vitellogenin). These findings provide a reference for understanding the evolutionary patterns of Hsps in Blattodea, and offer mechanistic insights into the developmental regulation mediated by Hsps in this important public-health pest.

Animals

Evolutionary Conservation and Reproductive Expression of ABC Transporter Genes in Two Sphenomorphus Skinks.

ATP-binding cassette (ABC) transporters represent one of the largest membrane protein superfamilies in vertebrates, playing essential roles in translocating diverse substrates across membranes. However, knowledge of ABC transporter genes in reptiles remains limited. In this study, we conducted a comprehensive genome-wide identification and characterization of the ABC gene family in oviparous Sphenomorphus incognitus and viviparous Sphenomorphus indicus. A total of 45 ABC genes were identified in each species and classified into seven subfamilies (ABCA-ABCG). Comparative and phylogenetic analyses revealed a generally conserved gene repertoire, with limited duplication events observed mainly in the ABCA and ABCG subfamilies, whereas other subfamilies (e.g., ABCE, ABCF, and ABCD) remained highly conserved. Interestingly, a lineage-specific duplication of ABCC2 was identified in lizards. Oviductal expression profiling revealed distinct temporal patterns of ABC gene expression across reproductive stages. Several genes, including ABCG1, ABCC3, and ABCD4, exhibited conserved expression trajectories across both species, suggesting shared transcriptional regulation. In contrast, ABCA1, ABCB1, and ABCG2 showed species-specific expression patterns, indicating regulatory divergence between the two lizard species. Overall, ABC gene expression was more dynamic in S. incognitus than in S. indicus. In summary, although the ABC transporter family is structurally conserved, it exhibits lineage-specific evolutionary changes and divergent transcriptional regulation in lizard oviducts. This study provides a foundation for understanding the diversity and regulation of ABC transporter genes in reptiles.

Animals

Evolutionary conservation and adaptability of cholecystokinin neuropeptide signaling in the sea cucumber Apostichopus japonicus.

BACKGROUND: Food ingestion is fundamental for animal survival and growth, with the cessation of feeding upon nutrient fulfillment being tightly regulated by a variety of satiety factors. Notably, sulfakinin/cholecystokinin (SK/CCK)-type neuropeptide signaling has been identified as an inhibitory regulator of food intake across the animal kingdom. However, its regulatory mechanism in feeding in deuterostome invertebrates remains unclear. Here, we characterized SK/CCK-type signaling in a deuterostome invertebrate, the sea cucumber Apostichopus japonicus (phylum Echinodermata). RESULTS: A single SK/CCK-type precursor in A. japonicus generates two mature peptides (AjSK/CCK1, AjSK/CCK2) that activate a shared receptor (AjSK/CCKR), triggering Ca2+ mobilization via the G&#x3b1;q-dependent pathway and extracellular signal regulated kinase 1/2 (ERK1/2) phosphorylation. Both peptides induce dose-dependent contraction of longitudinal muscles, while AjSK/CCK2 additionally elicits sustained contraction of the posterior intestine, an effect absent in other gut regions. Long-term injection of both peptides reduces food intake and significantly downregulates orexin-type neuropeptide genes (AjOrexin1P, AjOrexin2P) in the circumoral nerve ring (CNR) and intestine. CONCLUSIONS: Unlike mammals, where CCK inhibits feeding by contracting the pyloric sphincter to delay gastric emptying, SK/CCK-type peptides in sea cucumbers exert their anorexic effect in part by selectively contracting the posterior intestine, thereby inhibiting intestinal emptying. This divergence in action sites highlights the evolutionary adaptability of SK/CCK-type signaling as a conserved inhibitory regulator of feeding across bilaterian animals. Elucidating these mechanisms in the economically important A. japonicus may inform development of appetite-promoting agents for sustainable aquaculture.

Animals

Fishing for a reelGene: evaluating gene models with evolution and machine learning.

Assembled genomes and their associated annotations have transformed our study of gene function. However, each new annotated assembly generates new gene models. Inconsistencies between annotations likely arise from biological and technical causes, including pseudogene misclassification, transposon activity, and intron retention from sequencing of unspliced transcripts. To evaluate gene model predictions, we developed reelGene, a pipeline of machine learning models focused on (1) transcription boundaries, (2) mRNA integrity, and (3) protein structure. The first two models leverage sequence characteristics and evolutionary conservation across related taxa to learn the grammar of conserved transcription boundaries and mRNA sequences, while the third uses the conserved evolutionary grammar of protein sequences to predict whether a gene can produce a protein. Evaluating 1.8 million transcript models in Zea mays ssp. mays (maize), reelGene classified 28% as incorrectly annotated or non-functional. We find that reelGene classifies 92.2% of genes in the maize proteome and 99.2% of genes within the maize classical gene list as functional. reelGene also provides a way to further investigate genome biology- for instance, reelGene indicates that 10.3% of dispensable genes in B73 are functional, and within retained duplicate genes, reelGene identifies a 30% bias toward the retention of the M1 subgenome when one copy is functional and the other is non-functional. As an annotation-evaluating tool, reelGene is directly applicable to species of the Andropogoneae tribe, including other important crops like sorghum and miscanthus. As a community resource, reelGene has been integrated onto MaizeGDB both as a browser track and as an individual Shiny App, allowing researchers to evaluate gene model accuracy and further investigate genome biology.

Machine Learning

Chromosome-level genome assembly of Triplophysa scleroptera.

Triplophysa scleroptera is an endemic fish species in Qinghai Lake and the upper reaches of the Yellow River. However, studies on conservation and evolutionary genetics were seriously impeded by the absence of a reference genome. Here, by using PacBio HiFi sequencing and Hi-C assembly technology, we assembled a chromosome-level genome of T. scleroptera, with a total length of 660.22&#x2009;Mb and 99.82% of the sequence anchored to 25 chromosomes. The contig N50 and scaffold N50 were 9.09&#x2009;Mb and 24.38&#x2009;Mb, respectively. The evaluation using BUSCO indicated the genome assembly to be 96.40% complete. About 33.41% of the genome consists of repeat elements. We predicted 26,168 protein-coding genes in the genome, and 99.02% of them were functionally annotated. This high-quality reference genome would serve as a valuable genomic resource for advancing evolutionary conservation genetics studies in this species.

Animals

Molting in Pancrustacea Is Characterized by Both Deeply Conserved and Recently Evolved Gene Modules.

Arthropods such as insects and crustaceans, which together form the monophyletic group Pancrustacea, possess a rigid chitinous exoskeleton that must be periodically shed through molting to allow growth and morphological change. Although molting is a deeply conserved developmental process across Arthropoda, our understanding of its molecular mechanisms is still largely derived from insect model species. Lineage-specific innovations and losses of molting-related genes raise fundamental questions about the extent of its conservation outside noninsect arthropods. Here, we investigate the evolutionary conservation of molting gene expression across five representative pancrustacean species using publicly available transcriptomic datasets. Changes in gene expression during molting are characterized by both deeply conserved and lineage-specific gene modules. Temporal gene expression analyses reveal that these lineage-specific signatures are not uniformly distributed across the molting process: the middle transitional phase is more lineage-specific, thereby exhibiting an inverse hourglass pattern. This is likely due to life-history-specific processes, development of the cuticle, and specialized structures of the exoskeleton. Overall, this study provides evidence for both the evolutionary conservation and divergence of this key postembryonic developmental process and highlights the modular architecture of the molting program.

Animals

Genome-Wide Identification of the R2R3-MYB Gene Family in Solanum americanum and Functional Analysis of Its Role in Fruit Coloration.

Anthocyanins are key secondary metabolites responsible for fruit coloration in plants, and their biosynthesis is largely regulated by R2R3-MYB transcription factors. However, the R2R3-MYB regulators controlling fruit anthocyanin accumulation in wild Solanum species remain poorly understood. Here, Solanum americanum was used to identify candidate R2R3-MYB genes associated with fruit coloration through genome-wide identification, phylogenetic analysis, synteny analysis, expression profiling, and virus-induced gene silencing (VIGS). A total of 122 SaMYB genes were identified, and phylogenetic analysis revealed that SaMYB proteins clustered with Arabidopsis thaliana R2R3-MYB members in conserved subgroups, suggesting evolutionary conservation of this family. Synteny analysis identified 37 syntenic gene pairs among SaMYB genes, and the Ka/Ks values of all analyzable gene pairs were below 1, indicating that these duplicated genes are subject to functional constraint. Integrated analysis of phylogenetic relationships, protein structures, promoter cis-elements, and fruit developmental expression patterns identified SaMYB59 and SaMYB106 as candidate regulators of anthocyanin accumulation. VIGS analysis demonstrated that silencing SaMYB106 reduced purple coloration, decreased anthocyanin content, and downregulated the expression of the structural gene DFR. These results indicate that SaMYB106 functions as a positive regulator of fruit anthocyanin accumulation in S. americanum. This study provides insights into the molecular basis of fruit coloration in wild Solanum species.

Solanum americanum

Identification and differential regulation of proteasome &#x3b2; family genes by viral infection and cytokines in grass carp (Ctenopharyngodon idella).

Proteasome &#x3b2; (PSMB) subunits are essential components of the proteasome complex and play important roles in antigen processing and immune regulation. In this study, we identified 14 Psmb genes in grass carp (Ctenopharyngodon idella), including seven constitutive Psmbs (Psmb1-7), three immunoproteasome genes (Psmb8-10), two thymoproteasome-related paralogs (Psmb11a and Psmb11b), and two telelost-specific members (Psmb12 and Psmb13). Comparative genomic analyses showed that grass carp Psmb genes are highly conserved in genomic organization, gene synteny, and predicted &#x3b2;-subunit-like protein structures, supporting the evolutionary conservation of the proteasome &#x3b2;-subunit family in fish. Phylogenetic and syntenic analyses further revealed lineage-specific expansion of immunoproteasome-related Psmb genes in teleost fish, with Psmb12 and Psmb13 likely derived from duplications of Psmb9 and Psmb10, respectively. Tissue expression analysis suggested functional divergence among duplicated Psmb members, as constitutive Psmbs were relatively enriched in the brain, whereas immunoproteasome-related and teleost-specific Psmbs were highly expressed in immune- and mucosa-associated tissues. Moreover, GCRV-I infection rapidly induced Psmb8-10 and Psmb11b expression in CIK cells. IFN-&#x3b3; induced a broader set of Psmb genes than IFNa, whereas IL-10 selectively suppressed several Psmbs. Together, these findings highlight the evolutionary conservation, expansion, and immune-related diversification of the Psmb family in teleost fish.

Animals

Generation of Hoxa11-3XFLAG and Hoxd11-3XFLAG alleles to investigate Hox11 genome-wide binding.

Hox genes encode for evolutionary conserved transcription factors that direct the proper development of the body plan. Despite decades of research, little is known regarding their downstream target genes, especially in vertebrates. The strong evolutionary conservation of their DNA-binding homeodomain, their generic AT-rich binding sites, and the lack of specific antibodies has precluded rigorous examination. To circumvent these limitations, we have generated two mouse models in which a 3XFLAG epitope tag has been inserted into the 5' end of the coding sequence of both Hoxa11 and Hoxd11 loci via Cas9/CRISPR. The alleles have been validated by sequencing, PCR genotyping, western blotting, and protein expression analyses, demonstrating proper targeting and expression. Breeding these alleles in combination produces viable and fertile Hoxa11FLAG/FLAG; Hoxd11FLAG/FLAG animals, with no overt patterning defects unlike Hoxa11/Hoxd11 mutants that are infertile and have severe kidney and limb defects. By performing CUT&RUN and CUT&Tag analyses, we have confirmed DNA binding to a known Six2 enhancer in the developing kidney. These novel alleles will allow characterization of the genome-wide binding profile of Hox11 proteins in vivo.

Animals

Single-organ proteomics in Drosophila melanogaster larva.

The combination of genetic accessibility, organ complexity, evolutionary conservation, and cost-efficiency makes Drosophila melanogaster (Dm) a well-known model system for biomedical and fundamental biological research. Proteomic analysis of single organs enables the identification and quantification of proteins expressed in specific organs. This will help to uncover specific biological functions and unique protein profiles that are not detectable in whole-organism analyses. In this study we have isolated single organs form Dm larvae, and we have performed a deep proteomics mapping by following a minimal manipulation preparation procedure. The combined dataset across all organs comprised 9132 identified proteins. As anticipated, principal component analysis (PCA) revealed clear separation between the proteomes of most organs, confirming distinct protein profiles. These findings demonstrate the applicability of the sample preparation strategy for high-resolution proteomic characterization of individual organs in Drosophila. Given the extensive genetic tools available for this model organism, our approach has the potential to open new avenues for proteomic studies in Drosophila melanogaster and any other biological systems where the sample amount is limiting. SIGNIFICANCE STATEMENT: Drosophila melanogaster is a well-known model system for biomedical and fundamental biological research that serves as a valuable in vivo model organism due to its high degree of evolutionary conservation with higher vertebrates, tractable genetics, and logistical efficiency. However, the proteome of Drosophila at single organ level has been elusive to date, due to several factors like low sensitivity of previous generation mass spectrometers and sample preparation procedures, difficult isolation of some organs. In this study we have applied a compilation of advanced methods including minimal sample manipulation together with simple, straightforward and efficient protein extraction and digestion methods. Obtained peptides were minimally handled to be analyzed by applying specific and sensitive nLC methods coupled on-line to state-of-the-art MS/MS system. Altogether, the applied strategy allowed us to get the first single organ study to date for this animal. These datasets represent a significative resource for future genomic, transcriptomic and proteomic studies in Drosophila, as multi-omic integration requires deep proteomics to translate data into functional biochemistry, and serves as a critical bridge and an indispensable standalone resource across the genomic, transcriptomic, and proteomic landscapes.

Animals

Genome-wide identification of the HSP70 superfamily in tropical sea cucumber Stichopus monotuberculatus and their expression analysis under low-salinity stress.

Heat shock proteins (HSPs) are a group of evolutionarily conserved molecular chaperones that serve as indispensable core regulators in preserving cellular homeostasis and orchestrating organismal stress responses. The tropical sea cucumber Stichopus monotuberculatus, a high-value aquaculture species, is sensitive to fluctuations in environmental salinity-a challenge that has emerged as a critical bottleneck limiting its large-scale commercial cultivation. However, no systematic investigation has been conducted to characterize the HSP70 superfamily in S. monotuberculatus and elucidate its functional roles in salinity adaptation. In the present study, we performed a comprehensive genome-wide scan and identified 19 HSP70 superfamily genes in the S. monotuberculatus genome, with the HSP70IV subfamily showing remarkable gene expansion, containing 8 distinct copies. Phylogenetic analysis, conserved motif identification, and gene structure characterization demonstrated high evolutionary conservation within each HSP subfamily. These genes were unevenly distributed across the chromosomes of S. monotuberculatus, and prediction of cis-acting elements revealed that their upstream regulatory regions were enriched with numerous functional elements associated with stress response and immune regulation. Salinity stress experiments revealed that under severe low-salinity conditions (18&#x2030;), the expression levels of SmHSPA14L and multiple HSP70IV subfamily members were significantly elevated, while SmHYOU1D was significantly downregulated; in contrast, only subtle changes were detected in the expression of most HSP70 genes under moderate low-salinity stress (24&#x2030;). These findings strongly suggest that HSP70 genes, particularly the expanded HSP70IV subfamily, may act as key modulators in the low-salinity stress response. This work provides valuable insight into the molecular mechanisms underlying salinity adaptation in tropical sea cucumbers.

Animals

Genomic identification and functional characterization of the nuclear receptor gene family in relation to sex determination and gonad development in the Pacific oyster (Crassostrea gigas).

Nuclear receptors (NRs) are a large superfamily of transcription factors that control a wide range of physiological processes by modulating the expression of downstream target genes. Numerous studies have confirmed that NR family members play critical and conserved roles in sex determination and gonadal development across metazoans. However, in mollusks, systematic characterization of NRs and their potential functions in gonadal regulation remain largely unexplored. In this study, 46 NR gene family members in the Pacific oyster (Crassostrea gigas) were identified and assigned to eight subfamilies. All NR family members contain at least one of the two core domains (DNA-binding domain, DBD; ligand-binding domain, LBD), and conserved exon-intron structures were observed within the same subgroup, indicating their evolutionary conservation. Furthermore, expression profiling revealed high expression of CgNR2F, CgNR5A1-1, and CgNR0B1 in undifferentiated gonads, suggesting their potential involvement in sex determination. CgNR1A and CgNR2E5 were specifically expressed in female gonads and exhibited female-biased expression patterns, indicating a putative role in ovarian development. Moreover, CgNR3A and CgNR3B showed high expression levels during the undifferentiated stage and early male development stage, implying their possible participation in male gonadal development and gametogenesis. These results expand the understanding of the NR gene family in C. gigas and help elucidate the potential functions of NR genes in sex determination and gonadal development.

Animals

Molecular and Physiological Insights into CAT- and SOD-Associated Redox Homeostasis Under Salt Stress in Artemisia argyi.

Soil salinity disrupts redox homeostasis and limits plant growth and development. Although catalase (CAT) and superoxide dismutase (SOD) are key enzymatic antioxidants, the CAT and SOD gene families have not been characterized in Artemisia argyi (A. argyi), a species of medicinal and ecological importance. While SOD and CAT serve as the primary enzymatic scavengers for reactive oxygen species (ROS) detoxification, their genomic architecture and stress-responsive regulatory networks in A. argyi have remained uncharacterized. In this study, we conducted the first comprehensive genome-wide analysis of these gene families in A. argyi, identifying 22 structurally conserved members (8 AarCATs and 14 AarSODs). Collinearity and synteny analyses revealed strict lineage-specific evolutionary conservation, while tertiary protein modeling and subcellular localization illustrated a highly organized multi-organelle defense compartmentalization. High salinity (up to 200 mM NaCl) reduced the stomatal conductance and net photosynthetic rate. Salt stress reduced growth and increased osmoprotectant and antioxidant accumulation in A. argyi. Furthermore, histochemical staining using nitroblue tetrazolium (NBT) and 3,3'-Diaminobenzidine (DAB) provided comprehensive evidence of significant accumulation of ROS in leaves, which indicates the intense oxidative stress triggered by ionic stress. Tissue-specific analysis revealed that AarCAT1, AarCSD1, and AarFSD2 were 3.9-, 7.9-, and 12.7-fold higher in leaves than in roots, respectively. Under stress, AarCAT6 and AarCSD1 were strongly repressed in leaves by ~50% and ~46-70%, respectively, whereas AarMSD2 and AarMSD3 were significantly induced in roots by ~2.2- and ~1.8-fold. These distinct expression patterns suggest their potential involvement in tissue-specific stress adaptation and ROS homeostasis. These findings uncover the evolutionary and physiological basis of salt tolerance in A. argyi, providing genetic targets for climate-resilient breeding.

Artemisia

Recurrent Evolutionary Innovations in Rodent and Primate Schlafen Genes.

SCHLAFEN proteins are a large family of RNase-related enzymes carrying essential immune and developmental functions. Despite these important roles, Schlafen genes display varying degrees of evolutionary conservation in mammals. While this appears to influence their molecular activities, a detailed understanding of these evolutionary innovations is still lacking. Here, we used in-depth phylogenomic approaches to characterize the evolutionary trajectories and selective forces shaping mammalian Schlafen genes. We traced lineage-specific Schlafen amplifications and found that recent duplicates evolved under distinct selective forces, supporting repeated subfunctionalization cycles. Codon-level natural selection analyses in primates and rodents identified recurrent positive selection over Schlafen protein domains engaged in viral interactions. Combining known crystal structures and predicted protein structures, we discovered a novel class of rapidly evolving residues enriched at the contact interface of SCHLAFEN protein dimers. Our results suggest that inter-SCHLAFEN compatibilities are under strong selective pressures and are likely to impact their molecular functions. We posit that cycles of genetic conflicts with pathogens and between paralogs drove Schlafens' recurrent evolutionary innovations in mammals.

Animals

Genome-wide characterization of the bZIP gene family in Rattus norvegicus and expression profiling analysis during brain development.

BACKGROUND: The brown rat (Rattus norvegicus) serves as a cornerstone model organism in biomedical research, particularly for understanding physiological homeostasis and stress responses. The basic leucine zipper (bZIP) transcription factor family is a pivotal regulatory network involved in growth, organogenesis, and neurodevelopment. Despite its importance, a systematic characterization of the bZIP gene family in rats has remained elusive. RESULTS: In this study, we performed a genome-wide identification of 61 RnbZIP genes, which were categorized into 10 distinct subfamilies based on phylogenetic relationships and chromosomal localization. Structural analysis revealed conserved motif arrangements within subfamilies, while collinearity analysis identified significant gene duplication events-predominantly tandem and segmental duplications-that have driven the evolutionary expansion of the RnbZIP family. Quantitative analysis showed that members within the same subfamily shared 45%-92% sequence similarity (calculated using the BLOSUM62 scoring matrix), and all duplicated gene pairs underwent strong purifying selection (Ka/Ks&#x2009;<&#x2009;1). Comparative genomics across seven rodent species further underscored the evolutionary conservation and divergence of these factors. Expression profiling across diverse organs and brain developmental stages indicated that RnbZIP genes exhibit high tissue specificity. Notably, 10 candidate genes, including RnbZIP01, RnbZIP02, and RnbZIP08, demonstrated dynamic expression patterns during brain maturation, suggesting their essential roles in neurodevelopmental processes. CONCLUSIONS: Our findings provide a comprehensive structural and evolutionary framework for the RnbZIP gene family, highlighting their potential regulatory functions in rat organogenesis and brain development. This study establishes a valuable resource for further functional characterization of specific bZIP members in mammalian neurological systems.

Animals

From pan-life phase insights to PhaseHub: Analyzing protein condensate complexity.

Intracellular biomolecular condensation forms multicomponent signaling hubs that regulate development, stress responses, and environmental adaptation. While the molecular grammar encoded within scaffold proteins defines the basal associative features driving condensation, heterotypic condensates are intrinsically dynamic, multicomponent, and far-from-equilibrium systems. Consequently, how condensates organize component composition, stoichiometry, and functional specificity in space and time under physiological conditions remains poorly understood. Addressing this challenge requires integrative frameworks that combine predictive biophysical features with experimental information on protein abundance, interaction networks, subcellular localization, and evolutionary conservation. In this study, we first analyzed phase separation (PS) proteins across the tree of life in 1106 species, revealing a stark contrast in computationally predicted PS propensity between eukaryotes and prokaryotes, with genome size as a key determinant. Through a broad analysis of amino acid homorepeat-containing proteins (HRPs) across all species, we uncovered how PS evolves via a balance between functional condensation and avoidance of harmful, aggregation-prone sequences. We further identified potential signaling hubs and components across kingdoms by integrating PS-positive proteins with experimentally derived abundance and interactome data from four model eukaryotic species. Using Arabidopsis as a model, we dissected the relationships among PS propensity, condensation hub prediction, HRPs, subcellular localization, and structural conservation. Finally, we developed PhaseHub, a user-friendly interface for exploring scaffold-client dynamics, PS components, sequence signatures within each PS protein, and hubs. Collectively, our work provides an evolutionary framework for understanding multicomponent PS hubs by integrating molecular grammar with physiological context, thereby facilitating hypothesis generation and rational design.

Phase Separation

Conserved HSFA1-dependent chromatin dynamics drive heat stress responses in plants.

Eukaryotic organisms remodel chromatin landscapes to regulate gene expression in response to environmental stress. In plants, heat stress (HS) induces widespread chromatin changes, yet the role of heat shock transcription factors (HSFs) in chromatin remodeling and their evolutionary conservation remains unclear. Using Marchantia polymorpha Mphsf mutants and Arabidopsis thaliana Athsfa1s mutants, we identify HSFA1 as a key regulator of HS-induced cis-regulatory element (CRE) accessibility, a mechanism conserved across land plants, mice, and humans. Gene regulatory network modeling reveals parallel transcription factor subnetworks, with MpWRKY10 and MpABI5B acting as indirect and negative HS regulators. We further showed that ABA modulates gene expression in an HSFA1-dependent manner without inducing chromatin remodeling. Finally, we develop a machine learning framework integrating chromatin accessibility and CRE information to predict gene expression across species, revealing stress-responsive regulatory logic at the transcriptional level. These findings provide insights into how TFs coordinate chromatin architecture to drive stress adaptation.

Heat-Shock Response