Multiple functions and regulatory networks of WRKY33 and its orthologs.

Arabidopsis thaliana WRKY33 is currently one of the most studied members of the Group I WRKY transcription factor family. Research has confirmed that WRKY33 is involved in the regulation of various biological and abiotic stresses and occupies a central position in the regulatory network. The functional studies of orthologous genes of WRKY33 from other species are also receiving increasing attention. In this article, we summarized thirty-eight orthologous genes of AtWKRY33 from twenty-five different species. Their phylogenetic relationship and conserved WRKY domain were analyzed and compared. Similar to AtWKRY33, the well-studied orthologous gene members from rice and tomato also have multiple functions. In addition to playing important regulatory roles in responding to their specific pathogens, they are also involved in regulating various abiotic stresses and development. AtWKRY33 exerts its multiple functions through a complex regulatory network. Upstream transcription factors or other regulatory factors activate or inhibit the expression of AtWKRY33 at the chromatin and transcriptional levels. Interacting proteins affect the transcriptional activity of AtWKRY33 through phosphorylation, ubiquitination, SUMOylation, competition, or cooperation. The downstream genes are diverse and include three major categories: transcription factors, synthesis, metabolism, and signal transduction of various hormones, and disease resistance genes. In the regulatory network of AtWRKY33 orthologs, many conserved regulatory characteristics have been discovered, such as self-activation and phosphorylation by MAP kinases. This can provide a comparative reference for further studying the functions of other orthologous genes of AtWKRY33.

Depletion of lipid storage droplet-1 delays endoreplication progression and induces cell death in Drosophila salivary gland.

Perilipins are evolutionarily conserved from insects to mammals. Drosophila lipid storage droplet-1 (LSD-1) is a lipid storage droplet membrane surface-binding protein family member and a counterpart to mammalian perilipin 1 and is known to play a role in lipolysis. However, the function of LSD-1 during specific tissue development remains under investigation. This study demonstrated the role of LSD-1 in salivary gland development. Knockdown of Lsd-1 in the salivary gland was established using the GAL4/UAS system. The third-instar larvae of knockdown flies had small salivary glands containing cells with smaller nuclei. The null mutant Drosophila also showed the same phenotype. The depletion of LSD-1 expression induced a delay of endoreplication due to decreasing CycE expression and increasing DNA damage. Lsd-1 genetically interacted with Myc in the third-instar larvae. These results demonstrate that LSD-1 is involved in cell cycle and cell death programs in the salivary gland, providing novel insight into the effects of LSD-1 in regulating salivary gland development and the interaction between LSD-1 and Myc.

OrthoRefine: automated enhancement of prior ortholog identification via synteny.

BACKGROUND: Identifying orthologs continues to be an early and imperative step in genome analysis but remains a challenging problem. While synteny (conservation of gene order) has previously been used independently and in combination with other methods to identify orthologs, applying synteny in ortholog identification has yet to be automated in a user-friendly manner. This desire for automation and ease-of-use led us to develop OrthoRefine, a standalone program that uses synteny to refine ortholog identification. RESULTS: We developed OrthoRefine to improve the detection of orthologous genes by implementing a look-around window approach to detect synteny. We tested OrthoRefine in tandem with OrthoFinder, one of the most used software for identification of orthologs in recent years. We evaluated improvements provided by OrthoRefine in several bacterial and a eukaryotic dataset. OrthoRefine efficiently eliminates paralogs from orthologous groups detected by OrthoFinder. Using synteny increased specificity and functional ortholog identification; additionally, analysis of BLAST e-value, phylogenetics, and operon occurrence further supported using synteny for ortholog identification. A comparison of several window sizes suggested that smaller window sizes (eight genes) were generally the most suitable for identifying orthologs via synteny. However, larger windows (30 genes) performed better in datasets containing less closely related genomes. A typical run of OrthoRefine with ~ 10 bacterial genomes can be completed in a few minutes on a regular desktop PC. CONCLUSION: OrthoRefine is a simple-to-use, standalone tool that automates the application of synteny to improve ortholog detection. OrthoRefine is particularly efficient in eliminating paralogs from orthologous groups delineated by standard methods.

Impact of precursor-derived peracetic acid on post-weaning diarrhea, intestinal microbiota, and predicted microbial functional genes in weaned pigs.

Post-weaning diarrhea affects piglets in the nursery phase of production, leading to a substantial impact both at the farm and financial levels. The multifactorial etiology of this disease includes housing conditions, pig genetics, microbial composition, and metagenomic assets. Among the common therapeutic approaches, the widely used zinc oxide underwent a European Union ban in 2022 due to its negative environmental impact and correlation to increased antimicrobial resistance. During this study, we have tested two levels of inclusion of the potential antimicrobial alternative peracetic acid, delivered in water via the hydrolysis of the precursors sodium percarbonate and tetraacetylethylenediamine, in comparison to zinc oxide and an untreated control during a 2-week animal study. We assessed the microbial composition and predicted the metagenome, together with performance and physiological parameters, in order to describe the microbial functional role in etiopathology. Both zinc oxide and peracetic acid resulted in amelioration of the diarrheal status by the end of the trial period, with noticeable zinc oxide effects visible from the first week. This was accompanied by improved performance when compared to the first-week figures and a decreased stomach pH in both peracetic acid levels. A significant reduction in both stomach and caecal Proteobacteria was recorded in the zinc oxide group, and a significant reduction of Campylobacter in the stomach was reported for both zinc oxide and one of the peracetic acid concentrations. Among other functional differences, we found that the predicted ortholog for the zonula occludens toxin, a virulence factor present in pathogens like Escherichia coli and Campylobacter jejuni, was less abundant in the stomach of treated pigs compared to the control group. In water, peracetic acid delivered via precursor hydrolysis has the potential to be a valid intervention, an alternative to antimicrobial, to assist the weaning of piglets. Our findings support the view that post-weaning diarrhea is a complex multifactorial disease with an important metagenomic component characterized by the differential abundance of specific predicted orthologs and microbial genera in the stomach and caecum of pigs.

Phylogenetic Inference of Homologous/Orthologous Genes Among Distantly Related Plants.

The recent surge in plant genomic and transcriptomic data has laid a foundation for reconstructing evolutionary scenarios and inferring potential functions of key genes related to plants' development and stress responses. The classical scheme for identifying homologous genes is sequence similarity-based searching, under the crucial assumption that homologous sequences are more similar to each other than they are to any other non-homologous sequences. Advances in plant phylogenomics and computational algorithms have enabled us to systemically identify homologs/orthologs and reconstruct their evolutionary histories among distantly related lineages. Here, we present a comprehensive pipeline for homologous sequences identification, phylogenetic relationship inference, and potential functional profiling of genes in plants. Key features • Identification of orthologs using large-scale genomic and transcriptomic data. • This protocol is generalized for analyzing the evolution of plant genes.

The Interactive Complex between Cytomegalovirus Kinase vCDK/pUL97 and Host Factors CDK7-Cyclin H Determines Individual Patterns of Transcription in Infected Cells.

The infection of human cytomegalovirus (HCMV) is strongly determined by the host-cell interaction in a way that the efficiency of HCMV lytic replication is dependent on the regulatory interplay between viral and cellular proteins. In particular, the activities of protein kinases, such as cyclin-dependent kinases (CDKs) and the viral CDK ortholog (vCDK/pUL97), play an important role in both viral reproduction and virus-host interaction. Very recently, we reported on the complexes formed between vCDK/pUL97, human cyclin H, and CDK7. Major hallmarks of this interplay are the interaction between cyclin H and vCDK/pUL97, which is consistently detectable across various conditions and host cell types of infection, the decrease or increase in pUL97 kinase activity resulting from cyclin H knock-down or elevated levels, respectively, and significant trans-stimulation of human CDK7 activity by pUL97 in vitro. Due to the fact that even a ternary complex of vCDK/pUL97-cyclin H-CDK7 can be detected by coimmunoprecipitation and visualized by bioinformatic structural modeling, we postulated a putative impact of the respective kinase activities on the patterns of transcription in HCMV-infected cells. Here, we undertook a first vCDK/pUL97-specific transcriptomic analysis, which combined conditions of fully lytic HCMV replication with those under specific vCDK/pUL97 or CDK7 drug-mediated inhibition or transient cyclin H knockout. The novel results were further strengthened using bioinformatic modeling of the involved multi-protein complexes. Our data underline the importance of these kinase activities for the C-terminal domain (CTD) phosphorylation-driven activation of host RNA polymerase in HCMV-infected cells. The impact of the individual experimental conditions on differentially expressed gene profiles is described in detail and discussed.

KEGG tools for classification and analysis of viral proteins.

The KEGG database and analysis tools (https://www.kegg.jp) have been developed mostly for understanding genes and genomes of cellular organisms. The KO (KEGG Orthology) dataset, which is a collection of functional orthologs, plays the role of linking genes in the genome to pathways and other molecular networks, enabling KEGG mapping to uncover hidden features in the genome. Although viruses were part of KEGG for some time, they were not fully integrated in the KEGG analysis tools, because the KO assignment rate is very low for virus genes. To supplement KOs a new dataset named virus ortholog clusters (VOCs) is computationally generated, covering 90% of viral proteins in KEGG. VOCs can be used, in place of KOs, for taxonomy mapping to uncover relationships of sequence similarity groups and taxonomic groups and for identifying conserved gene orders in virus genomes. Furthermore, selected VOCs are used to define tentative KOs for characterizing protein functions. Here an overview of KEGG tools is presented focusing on these extensions for viral protein analysis.

Aon: a service to augment Alliance Genome Resource data with additional species.

OBJECTIVE: Cross-species comparative genomics requires access to accurate homology data across the entire range of annotated genes. The Alliance of Genome Resources (AGR) provides an open-source and comprehensive database of homology data calculated using a wide array of algorithms at differing stringencies to elucidate orthologous relationships. However, the current AGR application program interface (API) is limited to five homology endpoints for nine species. While AGR provides a robust resource for several canonical species, its utility can be greatly enhanced by increased filtering and data processing options and incorporating additional species. RESULTS: Here, we describe a novel API tool, AON, that expands access to the AGR orthology resource by creating a data structure that supports 50 additional endpoints. More importantly, it provides users with a framework for adding bespoke endpoints, custom species, and additional orthology data. We demonstrate AON's functionality by incorporating the service into the GeneWeaver ecosystem for supporting cross-species data analysis.

Requirements for anti-aquaporin 5 autoantibody production in a mouse model.

Several oral bacteria, including Prevotella melaninogenica (Pm), have aquaporin (AQP) proteins homologous to human AQP5, a major water channel protein targeted in Sjogren's syndrome. This study aimed to understand the antigenic characteristics that induce autoantibodies against an AQP5 "E" epitope (AQP5E) in a mouse model using C57BL/6 mice. Immunization with a PmE-L peptide derived from Pm AQP, which contains amino acid mismatches both at the B- and T-cell epitopes, efficiently induced anti-AQP5E autoantibodies accompanied by increased germinal center (GC) B and follicular helper T cells in the draining lymph nodes. However, PmE, a peptide lacking a T-cell epitope, and AQP5E-L, an AQP5-derived self-peptide, hardly induced either anti-AQP5E autoantibodies or GC responses. Surprisingly, OTII-AQP5E, a peptide that replaced the self T-cell epitope of AQP5E-L with an ovalbumin-derived foreign T-cell epitope, was not any better than AQP5E-L in the induction of anti-AQP5E autoantibodies and GC response, despite the substantial expansion of CD4+ T cells and production of anti-OTII-AQP5E antibodies. The complex of biotinylated PmE-L peptide and highly immunogenic streptavidin (SA) induced a strong extrafollicular B-cell response skewed toward the expansion of SA-specific B cells. However, the expansion of AQP5E-specific GC B cells was limited, resulting in the inefficient induction of anti-AQP5E autoantibodies. Collectively, our results have demonstrated that anti-AQP5E autoantibody production is only allowed when foreign B- and T-cell epitopes drive a strong GC response of AQP5E-specific B cells for affinity maturation. This study helps explain why cross-reactive anti-AQP5 autoantibodies are not produced during the immune response to Pm in most healthy people.

Subfunctionalisation of paralogous genes and evolution of differential codon usage preferences: The showcase of polypyrimidine tract binding proteins.

Gene paralogs are copies of an ancestral gene that appear after gene or full genome duplication. When two sister gene copies are maintained in the genome, redundancy may release certain evolutionary pressures, allowing one of them to access novel functions. Here, we focused our study on gene paralogs on the evolutionary history of the three polypyrimidine tract binding protein genes (PTBP) and their concurrent evolution of differential codon usage preferences (CUPrefs) in vertebrate species. PTBP1-3 show high identity at the amino acid level (up to 80%) but display strongly different nucleotide composition, divergent CUPrefs and, in humans and in many other vertebrates, distinct tissue-specific expression levels. Our phylogenetic inference results show that the duplication events leading to the three extant PTBP1-3 lineages predate the basal diversification within vertebrates, and genomic context analysis illustrates that local synteny has been well preserved over time for the three paralogs. We identify a distinct evolutionary pattern towards GC3-enriching substitutions in PTBP1, concurrent with enrichment in frequently used codons and with a tissue-wide expression. In contrast, PTBP2s are enriched in AT-ending, rare codons, and display tissue-restricted expression. As a result of this substitution trend, CUPrefs sharply differ between mammalian PTBP1s and the rest of PTBPs. Genomic context analysis suggests that GC3-rich nucleotide composition in PTBP1s is driven by local substitution processes, while the evidence in this direction is thinner for PTBP2-3. An actual lack of co-variation between the observed GC composition of PTBP2-3 and that of the surrounding non-coding genomic environment would raise an interrogation on the origin of CUPrefs, warranting further research on a putative tissue-specific translational selection. Finally, we communicate an intriguing trend for the use of the UUG-Leu codon, which matches the trends of AT-ending codons. Our results are compatible with a scenario in which a combination of directional mutation-selection processes would have differentially shaped CUPrefs of PTBPs in vertebrates: the observed GC-enrichment of PTBP1 in placental mammals may be linked to genomic location and to the strong and broad tissue-expression, while AT-enrichment of PTBP2 and PTBP3 would be associated with rare CUPrefs and thus, possibly to specialized spatio-temporal expression. Our interpretation is coherent with a gene subfunctionalisation process by differential expression regulation associated with the evolution of specific CUPrefs.

Apoptotic Factors Are Evolutionarily Conserved Since Mitochondrial Domestication.

The mechanisms initiating apoptotic programmed cell death in diverse eukaryotes are very similar. Basically, the mitochondrial permeability transition activates apoptotic proteases, DNases, and flavoproteins such as apoptosis-inducing factors (AIFs). According to the hypothesis of the endosymbiotic origin of apoptosis, these mechanisms evolved during mitochondrial domestication. Various phylogenetic analyses, including ours, have suggested that apoptotic factors were eubacterial protomitochondrial toxins used for killing protoeukaryotic hosts. Here, we tested whether the function of yeast Saccharomyces cerevisiae apoptotic proteases (metacaspases Mca1 and Nma111), DNase Nuc1, and flavoprotein Ndi1 can be substituted with orthologs from remotely related eukaryotes such as plants, protists, and eubacteria. We found that orthologs of remotely related eukaryotic and even eubacterial proteins can initiate apoptosis in yeast when triggered by chemical stresses. This observation suggests that apoptotic mechanisms have been maintained since mitochondrial domestication, which occurred approximately 1,800 Mya. Additionally, it supports the hypothesis that some of these apoptotic factors could be modified eubacterial toxins.

A Chicken Tapasin ortholog can chaperone empty HLA-B∗37:01 molecules independent of other peptide-loading components.

Human Tapasin (hTapasin) is the main chaperone of MHC-I molecules, enabling peptide loading and antigen repertoire optimization across HLA allotypes. However, it is restricted to the endoplasmic reticulum (ER) lumen as part of the protein loading complex (PLC), and therefore is highly unstable when expressed in recombinant form. Additional stabilizing co-factors such as ERp57 are required to catalyze peptide exchange in vitro, limiting uses for the generation of pMHC-I molecules of desired antigen specificities. Here, we show that the chicken Tapasin (chTapasin) ortholog can be expressed recombinantly at high yields in a stable form, independent of co-chaperones. chTapasin can bind the human HLA-B∗37:01 with low micromolar-range affinity to form a stable tertiary complex. Biophysical characterization by methyl-based NMR methods reveals that chTapasin recognizes a conserved ß2m epitope on HLA-B∗37:01, consistent with previously solved X-ray structures of hTapasin. Finally, we provide evidence that the B∗37:01/chTapasin complex is peptide-receptive and can be dissociated upon binding of high-affinity peptides. Our results highlight the use of chTapasin as a stable scaffold for protein engineering applications aiming to expand the ligand exchange function on human MHC-I and MHC-like molecules.

Biochemical functions and structure of Caenorhabditis elegans ZK177.8 protein: Aicardi-Goutières syndrome SAMHD1 dNTPase ortholog.

Mutations in sterile alpha motif domain and histidine-aspartate domain-containing protein 1 (SAMHD1) are found in a neurodevelopmental disorder, Aicardi-Goutières syndrome, and cancers, and SAMHD1, which is a deoxynucleoside triphosphate (dNTP) triphosphorylase, was identified as a myeloid-specific HIV-1 restriction factor. Here, we characterized the enzymology and structure of an SAMHD1 ortholog of Caenorhabditis elegans, ZK177.8, which also reportedly induces developmental defects upon gene knockdown. We found ZK177.8 protein is a dNTPase allosterically regulated by dGTP. The active site of ZK177.8 recognizes both 2' OH and triphosphate moieties of dNTPs but not base moiety. The dGTP activator induces the formation of the enzymatically active ZK177.8 tetramers, and ZK177.8 protein lowers cellular dNTP levels in a human monocytic cell line. Finally, ZK177.8 tetramers display very similar X-ray crystal structure with human and mouse SAMHD1s except that its lack of the canonical sterile alpha motif domain. This striking conservation in structure, function, and allosteric regulatory mechanism for the hydrolysis of the DNA building blocks supports their host developmental roles.

Cytomegalovirus cyclin-dependent kinase ortholog vCDK/pUL97 undergoes regulatory interaction with human cyclin H and CDK7 to codetermine viral replication efficiency.

Human cytomegalovirus (HCMV) infection is shaped by a tightly regulated interplay between viral and cellular proteins. Distinct kinase activities, such as the viral cyclin-dependent kinase ortholog (vCDK) pUL97 and cellular CDK7 are both crucial for efficient viral replication. Previously, we reported that both kinases, vCDK/pUL97 and CDK7, interact with cyclin H, thereby achieving an enhanced level of kinase activity and overall functionality in viral replication. Here we provide a variety of novel results, as generated on a methodologically extended basis, and present a concept for the codetermination of viral replication efficiency through these kinase activities: (i) cyclin H expression, in various human cell types, is substantially upregulated by strains of HCMV including the clinically relevant HCMV Merlin; (ii) vCDK/pUL97 interacts with human cyclin H in both HCMV-infected and plasmid-transfected cell systems; (iii) a doxycycline-inducible shRNA-dependent knock-down (KD) of cyclin H significantly reduces pUL97 activity (qSox in vitro kinase assay); (iv) accordingly, pUL97 in vitro kinase activity is seen significantly increased upon addition of recombinant cyclin H; (v) as a point of specific importance, human CDK7 activity shows an increase by vCDK/pUL97-mediated trans-stimulation (whereas pUL97 is not stimulated by CDK7); (vi) phosphosite-specific antibodies indicate an upregulated CDK7 phosphorylation upon HCMV infection, as mediated through a pUL97-specific modulatory effect (i.e. shown by pUL97 inhibitor treatment or pUL97-deficient viral mutant); (vii) finally, an efficient KD of cyclin H in primary fibroblasts generally results in an impaired HCMV replication efficiency as measured on protein and genomic levels. These results show evidence for the codetermination of viral replication by vCDK/pUL97, cyclin H and CDK7, thus supporting the specific importance of cyclin H as a central regulatory factor, and suggesting novel targeting options for antiviral drugs.

Analysis of the Arabidopsis venosa4-0 mutant supports the role of VENOSA4 in dNTP metabolism.

Human Sterile alpha motif and histidine-aspartate domain containing protein 1 (SAMHD1) functions as a dNTPase to maintain dNTP pool balance. In eukaryotes, the limiting step in de novo dNTP biosynthesis is catalyzed by RIBONUCLEOTIDE REDUCTASE (RNR). In Arabidopsis, the RNR1 subunit of RNR is encoded by CRINKLED LEAVES 8 (CLS8), and RNR2 by three paralogous genes, including TSO MEANING 'UGLY' IN CHINESE 2 (TSO2). In plants, DIFFERENTIAL DEVELOPMENT OF VASCULAR ASSOCIATED CELLS 1 (DOV1) catalyzes the first step of the de novo biosynthesis of purines. Here, to explore the role of VENOSA4 (VEN4), the most likely Arabidopsis ortholog of human SAMHD1, we studied the ven4-0 point mutation, whose leaf phenotype was stronger than those of its insertional alleles. Structural predictions suggested that the E249L substitution in the mutated VEN4-0 protein rigidifies its 3D structure. The morphological phenotypes of the ven4, cls8, and dov1 single mutants were similar, and those of the ven4 tso2 and ven4 dov1 double mutants were synergistic. The ven4-0 mutant had reduced levels of four amino acids related to dNTP biosynthesis, including glutamine and glycine, which are precursors in the de novo purine biosynthesis. Our results reveal high functional conservation between VEN4 and SAMHD1 in dNTP metabolism.

Regulatory mechanism of trichothecene biosynthesis in Fusarium graminearum.

Among the genes involved in the biosynthesis of trichothecene (Tri genes), Tri6 and Tri10 encode a transcription factor with unique Cys2His2 zinc finger domains and a regulatory protein with no consensus DNA-binding sequences, respectively. Although various chemical factors, such as nitrogen nutrients, medium pH, and certain oligosaccharides, are known to influence trichothecene biosynthesis in Fusarium graminearum, the transcriptional regulatory mechanism of Tri6 and Tri10 genes is poorly understood. Particularly, culture medium pH is a major regulator in trichothecene biosynthesis in F. graminearum, but it is susceptible to metabolic changes posed by nutritional and genetic factors. Hence, appropriate precautions should be considered to minimize the indirect influence of pH on the secondary metabolism while studying the roles of nutritional and genetic factors on trichothecene biosynthesis regulation. Additionally, it is noteworthy that the structural changes of the trichothecene gene cluster core region exert considerable influence over the normal regulation of Tri gene expression. In this perspective paper, we consider a revision of our current understanding of the regulatory mechanism of trichothecene biosynthesis in F. graminearum and share our idea toward establishing a regulatory model of Tri6 and Tri10 transcription.

The Whole Genome of Colletes collaris (Hymenoptera: Colletidae): An Important Step in Comparative Genomics of Cellophane Bees.

Gaining knowledge on bees is of the utmost importance due to the paramount role that they play in angiosperm pollination. Herein, we provide the first genome assembly of Colletes collaris, a pan-Eurasian cellophane bee. We sequenced 50.53 Gbp of long-read data plus 57.36 Gbp of short-read data in Oxford Nanopore Technologies and Illumina platforms, respectively. The genome assembly consisted of 374.75 Mbp distributed across 374 contigs, with L50 and N50 of 9 and 8.96 Mbp, respectively. We predicted the genome to comprise 20,399 protein-coding genes, 467,947 repeats, and 4,315 non-coding RNA genes. The transcriptome and mitochondrial genome of the species were also assembled. Gene family analysis with 15 insect species identified 14,417 families, 9,517 of them found in C. collaris. A dated phylogenomic analysis revealed high numbers of orthogroups experiencing rapid evolution within Colletes.

Large-scale analysis of putative Euphorbiaceae R2R3-MYB transcription factors identifies a MYB involved in seed oil biosynthesis.

BACKGROUND: MYB transcription factors are widely distributed in the plant kingdom and play key roles in regulatory networks governing plant metabolism and biochemical and physiological processes. RESULTS: Here, we first determined the R2R3-MYB genes in five Euphorbiaceae genomes. The three Trp (W) residues from the first MYB domain (R2) were absolutely conserved, whereas the first W residue from the second MYB domain (R3) was preferentially mutated. The R2R3-MYBs were clustered into 48 functional subfamilies, of which 34 had both R2R3-MYBs of Euphorbiaceae species and AtMYBs, and four contained only Euphorbiaceae R2R3-MYBs. The whole-genome duplication (WGD) and/or segmental duplication (SD) played key roles in the expansion of the R2R3-MYB family. Unlike paralogous R2R3-MYB family members, orthologous R2R3-MYB members contained a higher selective pressure and were subject to a constrained evolutionary rate. VfMYB36 was specifically expressed in fruit, and its trend was consistent with the change in oil content, indicating that it might be involved in oil biosynthesis. Overexpression experiments showed that VfMYB36 could significantly provide linolenic acid (C18:3) content, which eventually led to a significant increase in oil content. CONCLUSION: Our study first provides insight into understanding the evolution and expression of R2R3-MYBs in Euphorbiaceae species, and also provides a target for the production of biomass diesel and a convenient way for breeding germplasm resources with high linolenic acid content in the future.

InParanoiDB 9: Ortholog Groups for Protein Domains and Full-Length Proteins.

Prediction of orthologs is an important bioinformatics pursuit that is frequently used for inferring protein function and evolutionary analyses. The InParanoid database is a well known resource of ortholog predictions between a wide variety of organisms. Although orthologs have historically been inferred at the level of full-length protein sequences, many proteins consist of several independent protein domains that may be orthologous to domains in other proteins in a way that differs from the full-length protein case. To be able to capture all types of orthologous relations, conventional full-length protein orthologs can be complemented with orthologs inferred at the domain level. We here present InParanoiDB 9, covering 640 species and providing orthologs for both protein domains and full-length proteins. InParanoiDB 9 was built using the faster InParanoid-DIAMOND algorithm for orthology analysis, as well as Domainoid and Pfam to infer orthologous domains. InParanoiDB 9 is based on proteomes from 447 eukaryotes, 158 bacteria and 35 archaea, and includes over one billion predicted ortholog groups. A new website has been built for the database, providing multiple search options as well as visualization of groups of orthologs and orthologous domains. This release constitutes a major upgrade of the InParanoid database in terms of the number of species as well as the new capability to operate on the domain level. InParanoiDB 9 is available at https://inparanoidb.sbc.su.se/.

OrthoPhy: A Program to Construct Ortholog Data Sets Using Taxonomic Information.

Species phylogenetic trees represent the evolutionary processes of organisms, and they are fundamental in evolutionary research. Therefore, new methods have been developed to obtain more reliable species phylogenetic trees. A highly reliable method is the construction of an ortholog data set based on sequence information of genes, which is then used to infer the species phylogenetic tree. However, although methods for constructing an ortholog data set for species phylogenetic analysis have been developed, they cannot remove some paralogs, which is necessary for reliable species phylogenetic inference. To address the limitations of current methods, we developed OrthoPhy, a program that excludes paralogs and constructs highly accurate ortholog data sets using taxonomic information dividing analyzed species into monophyletic groups. OrthoPhy can remove paralogs, detecting inconsistencies between taxonomic information and phylogenetic trees of candidate ortholog groups clustered by sequence similarity. Performance tests using evolutionary simulated sequences and real sequences of 40 bacteria revealed that the precision of ortholog inference by OrthoPhy is higher than that of existing programs. Additionally, the phylogenetic analysis of species was more accurate when performed using ortholog data sets constructed by OrthoPhy than that performed using data sets constructed by existing programs. Furthermore, we performed a benchmark test of the Quest for Orthologs using real sequence data and found that the concordance rate between the phylogenetic trees of orthologs inferred by OrthoPhy and those of species was higher than the rates obtained by other ortholog inference programs. Therefore, ortholog data sets constructed using OrthoPhy enabled a more accurate phylogenetic analysis of species than those constructed using the existing programs, and OrthoPhy can be used for the phylogenetic analysis of species even for distantly related species that have experienced many evolutionary events.