Domain Architecture Research Articles

Structural and functional insights from the sequences and complex domain architecture of adhesin-like proteins from Methanobrevibacter smithii and Methanosphaera stadtmanae

Methanogenic archaea, or methanogens, are crucial in guts and rumens, consuming hydrogen, carbon dioxide, and other fermentation products. While their molecular interactions with other microorganisms are not fully understood, genomic sequences provide information. The first genome sequences of human gut methanogens, Methanosphaera stadtmanae and Methanobrevibacter smithii, revealed genes encoding adhesin-like proteins (ALPs). These proteins were also found in other gut and rumen methanogens, but their characteristics and functions remain largely unknown. This study analyzes the ALP repertoire of M. stadtmanae and M. smithii using AI-guided protein structure predictions of unique ALP domains. Both genomes encode more than 40 ALPs each, comprising over 10% of their genomes. ALPs contain repetitive sequences, many of which are unmatched in protein domain databases. We present unique sequence signatures of conserved ABD repeats in ALPs and propose a classification based on domain architecture. Our study offers insights into ALP features and how methanogens may interact with other microorganisms.

Tuning ferroelectric domains and polarization properties of flexible BiFeO3 thin films by phase transition engineering

In the present investigation, thin films of flexible (Bi1-xSmx)(Fe0.95Mn0.05)O3 (denoted as BSFM0, BSFM4, BSFM8, and BSFM12 for x=0 %, 4 %, 8 %, and 12 %, respectively) were synthesized on metallic Ni-Cr substrates via the sol-gel technique. A comprehensive examination was conducted to elucidate the influence of varying concentrations of Sm3+ substitution on the phase constitution, oxygen vacancy concentration, and leakage behavior of the (Bi1-xSmx)(Fe0.95Mn0.05)O3 films. Utilizing X-ray diffraction (XRD) and Raman spectrometry, it was ascertained that all films exhibited a composite structure comprising orthorhombic (Pnma) and rhombohedral (R3c) phases. An augmentation in Sm3+ doping led to a non-monotonic variation in the R3c phase content, initially increasing and subsequently decreasing, while the nonpolar Pnma phase demonstrated a progressive enhancement. The distorted R3c phase was found to promote the flipping of the domain architecture, thereby augmenting the polarization attributes. Notably, at a Sm doping level of 4 %, the maximum polarization (Pmax) attained was 95 μC/cm2, with a remnant polarization (Pr) of 78 μC/cm2. The incorporation of a judicious amount of Sm was observed to curtail film leakage and decelerate the aging phenomenon. The compositional stability of this doping level was evaluated across a frequency spectrum of 0.1–12 kHz and a temperature gradient of 25–200 °C, revealing no propensity for degradation under conditions of polarization, a holding duration of 104 s, or fatigue scenarios involving a curvature of 5 mm and 108 switching cycles subjected to 103 instances of bending. These findings provide innovative insights for the development of next-generation lead-free transducer apparatus and flexible information storage mediums.

Characterization and Transcriptomic Analysis of Sorghum EIN/EIL Family and Identification of Their Roles in Internode Maturation.

Ethylene-insensitive 3/Ethylene-insensitive3-like proteins (EIN3/EIL) represent a group of transcription factors critical for the ethylene signaling transduction that manipulate downstream ethylene-responsive genes, thereby regulating plant growth, development, and stress responses. However, the identification, evolution, and divergence of the EIL family remain to be studied in Sorghum bicolor. Here, we identified eight SbEILs, which were expanded due to whole-genome-duplication (WGD) events. Characterization of the protein sequences and expression atlas demonstrates that the WGD-duplicated SbEILs could become divergent due to the differential expression patterns, rather than domain and motif architectures. Comparative expression analysis was performed between the RNA-seq data sets of internodes from several sorghum cultivars to understand the potential roles of SbEIL members in internode elongation and maturation. Our results identified SbEIL3 and 7 (the latter as a homolog of OsEIL7/OsEIL1) to be the highly expressed SbEIL genes in sorghum internodes and revealed a potential functional link between SbEIL7 and internode maturation. The co-expression analysis and comparative expression analysis with ethylene-regulated gene sets found that SbEIL7 was co-regulated with a set of ubiquitin-related protein degradation genes, suggesting possible involvement of SbEIL7 in protein degradation and processing during the post-anthesis stages. Altogether, our findings lay a foundation for future functional studies of ethylene signaling-mediated gene regulation and improvement of sorghum internode development.

Thrombospondins: Conserved mediators and modulators of metazoan extracellular matrix.

This review provides a personal overview of significant scientific developments in the thrombospondin field during the course of my career. Thrombospondins are multidomain, multimeric, calcium-binding extracellular glycoproteins with context-specific roles in tissue organisation. They act at cell surfaces and within ECM to regulate cell phenotype and signalling, differentiation and assembly of collagenous ECM, along with tissue-specific roles in cartilage, angiogenesis and synaptic function. More recently, intracellular, homeostatic roles have also been identified. Resolution of structures for the major domains of mammalian thrombospondins has facilitated major advances in understanding thrombospondin biology from molecule to tissue; for example, in illuminating molecular consequences of disease-causing coding mutations in human pseudoachrondroplasia. Although principally studied in vertebrates, thrombospondins are amongst the most ancient of animal ECM proteins, with many invertebrates encoding a single thrombospondin and the thrombospondin gene family of vertebrates originating through gene duplications. Moreover, thrombospondins form one branch of a thrombospondin superfamily that debuted at the origin of metazoans. The super-family includes additional sub-groups, present only in invertebrates, that differ in N-terminal domain organisation, share the distinctive TSP C-terminal region domain architecture and, to the limited extent studied to date, apparently contribute to tissue development and organisation. Finally, major lines of translational research are discussed, related to fibrosis; TSP1, TSP2 and inhibition of angiogenesis; and the alleviation of chronic cartilage tissue pathologies in pseudoachrondroplasia.

Identification and functional analysis of C-type lectin from mosquito Aedes albopictus in response to dengue virus infection

BackgroundC-type lectins (CTLs) are a large family of proteins with sugar-binding activity. CTLs contain an evolutionarily conserved C-type lectin domain (CTLD) that binds microbial carbohydrates in a calcium-dependent manner, thereby playing a key role in both microbial pathogenesis and innate immune responses. Aedes albopictus is an important vector for transmitting dengue virus (DENV) worldwide. Currently, the molecular characteristics and functions of CTLs in Ae. albopictus are largely unknown.MethodsTranscripts encoding CTL proteins in the Ae. albopictus genome assembly were analyzed via sequence blast. Phylogenetic analysis and molecular characterization were performed to identify the functional domains of the CTLs. Quantitative analysis was performed to determine the gene expression features of CTLs during mosquito development and in different tissues of female adults after blood feeding. In addition, the functional role of CTLs in response to DENV infection was investigated in Ae. albopictus mosquito cells.ResultsWe identified 39 transcripts encoding CTL proteins in the Ae. albopictus transcriptome. Aedes albopictus CTLs are classified into three groups based on the number of CTLDs and the domain architecture. These included 29 CTL-Ss (single-CTLDs), 1 immulectins (dual-CTLD) and 9 CTL-Xs (CTLDs with other domains). Phylogenetic analysis and structural modeling indicated that CTLs in Ae. albopictus are highly conserved with the homologous CTLs in Aedes aegypti. The expression profile assay revealed differential expression patterns of CTLs in both developmental stages and in adult female tissues. Knockdown and overexpression of three CTLs (CTL-S12, S17 and S19) confirmed that they can promote dengue virus infection in Ae. albopictus cells.ConclusionsThe CTL genes in Ae. albopictus mosquito and other mosquito species are evolutionarily conserved and exhibit different developmental and tissue expression features. The functional assay indicated that three CTLs in Ae. albopictus mosquitoes are involved in promoting dengue virus infection. Our study revealed that CTLs play important roles in both the physiological processes and viral infection in mosquito vectors.Graphical

Structural and Phylogenetic In Silico Characterization of Vitis vinifera PRR Protein as Potential Target for Plasmopara viticola Infection.

Fungi infection, especially derived from Plasmopara viticola, causes severe grapevine economic losses worldwide. Despite the availability of chemical treatments, looking for eco-friendly ways to control Vitis vinifera infection is gaining much more attention. When a plant is infected, multiple disease-control molecular mechanisms are activated. PRRs (Pattern Recognition Receptors) and particularly RLKs (receptor-like kinases) take part in the first barrier of the immune system, and, as a consequence, the kinase signaling cascade is activated, resulting in an immune response. In this context, discovering new lectin-RLK (LecRLK) membrane-bounded proteins has emerged as a promising strategy. The genome-wide localization of potential LecRLKs involved in disease defense was reported in two grapevine varieties of great economic impact: Chardonnay and Pinot Noir. A total of 23 potential amino acid sequences were identified, exhibiting high-sequence homology and evolution related to tandem events. Based on the domain architecture, a carbohydrate specificity ligand assay was conducted with docking, revealing two sequences as candidates for specific Vitis vinifera-Plasmopara viticola host-pathogen interaction. This study confers a starting point for designing new effective antifungal treatments directed at LecRLK targets in Vitis vinifera.

Genome-wide identification of sweet potato U-Box E3 ubiquitin ligases and roles of IbPUB52 in negative regulation of drought stress.

The plant U-box (PUB) proteins, a family of ubiquitin ligases (E3) enzymes, are pivotal in orchestrating many biological processes and facilitating plant responses to environmental stressors. Despite their critical roles, exploring the PUB gene family's characteristics and functional diversity in sweet potato (Ipomoea batatas (L.) Lam.) has been notably limited. There were 81 IbPUB genes identified within the sweet potato genome, and they were categorized into eight distinct groups based on domain architecture, revealing a non-uniform distribution across the 15 chromosomes of I. batatas. The investigation of cis-acting elements has shed light on the potential of PUBs to participate in a wide array of biological processes, particularly emphasizing their role in mediating responses to abiotic stresses. Transcriptome profiles revealed that IbPUB genes displayed a wide range of expression levels among different tissues and were regulated by salt or drought stress. IbPUB52 has emerged as a gene of significant interest due to its induction by salt and drought stresses. Localization studies have confirmed the presence of IbPUB52 in both the nucleus and the cytoplasm, and its ubiquitination activity has been validated through rigorous in vitro and in vivo assays. Intriguingly, the heterogeneous expression of IbPUB52 in Arabidopsis resulted in decreased drought tolerance. The virus-induced gene silencing (VIGS) of IbPUB52 in sweet potatoes led to enhanced resistance to drought. This evidence suggests that IbPUB52 negatively regulates the drought tolerance of plants. The findings of this study are instrumental in advancing our comprehension of the functional dynamics of PUB E3 ubiquitin ligases in sweet potatoes.

Role of protein domains in trafficking and localization of the voltage-gated sodium channel β2 subunit

The voltage-gated sodium (NaV) channel is critical for cardiomyocyte function since it is responsible for action potential initiation and its propagation throughout the cell. It consists of a protein complex made of a pore forming α subunit and associated β subunits, which regulate α subunit function and subcellular localization. We previously showed the implication of N-linked glycosylation and S-acylation of β2 in its polarized trafficking. Here, we present evidence of β2 dimerization. Moreover, we demonstrate the implication of the cytoplasmic tail, extracellular loop, and transmembrane domain on proper β2 folding and export to the cell surface of polarized Madin-Darby canine kidney cells. Substantial alteration, or lack of any of these domains, leads to accumulation of β2 in the endoplasmic reticulum, along with impaired complex N-glycosylation, which is needed for its efficient surface delivery. We also show that these alterations to β2 affected to certain extent NaV1.5 surface localization. Conversely, however, NaV1.5 had little or no influence on β2 trafficking, its localization to the surface, or homodimer formation. Altogether, our data link the architecture of the β2 domains to the establishment of its proper subcellular localization. These findings could provide valuable insights to gain a deeper comprehension of the elusive biology of β subunits in excitable cells, such as neurons and cardiomyocytes.

Genomic Underpinnings of Cytoplasmic Incompatibility: CIF Gene-Neighborhood Diversification Through Extensive Lateral Transfers and Recombination in Wolbachia.

Cytoplasmic incompatibility (CI), a non-Mendelian genetic phenomenon, involves the manipulation of host reproduction by Wolbachia, a maternally transmitted alphaproteobacterium. The underlying mechanism is centered around the CI Factor (CIF) system governed by two genes, cifA and cifB, where cifB induces embryonic lethality, and cifA counteracts it. Recent investigations have unveiled intriguing facets of this system, including diverse cifB variants, prophage association in specific strains, copy number variation, and rapid component divergence, hinting at a complex evolutionary history. We utilized comparative genomics to systematically classify CIF systems, analyze their locus structure and domain architectures, and reconstruct their diversification and evolutionary trajectories. Our new classification identifies ten distinct CIF types, featuring not just versions present in Wolbachia, but also other intracellular bacteria, and eukaryotic hosts. Significantly, our analysis of CIF loci reveals remarkable variability in gene composition and organization, encompassing an array of diverse endonucleases, variable toxin domains, deubiquitinating peptidases (DUBs), prophages, and transposons. We present compelling evidence that the components within the loci have been diversifying their sequences and domain architectures through extensive, independent lateral transfers and interlocus recombination involving gene conversion. The association with diverse transposons and prophages, coupled with selective pressures from host immunity, likely underpins the emergence of CIF loci as recombination hotspots. Our investigation also posits the origin of CifB-REase domains from mobile elements akin to CR (Crinkler-RHS-type) effectors and Tribolium Medea1 factor, which is linked to another non-Mendelian genetic phenomenon. This comprehensive genomic analysis offers novel insights into the molecular evolution and genomic foundations of Wolbachia-mediated host reproductive control.

Divergent Roles of the Auxin Response Factors in Lemongrass (Cymbopogon flexuosus (Nees ex Steud.) W. Watson) during Plant Growth.

Auxin Response Factors (ARFs) make up a plant-specific transcription factor family that mainly couples perception of the phytohormone, auxin, and gene expression programs and plays an important and multi-faceted role during plant growth and development. Lemongrass (Cymbopogon flexuosus) is a representative Cymbopogon species widely used in gardening, beverages, fragrances, traditional medicine, and heavy metal phytoremediation. Biomass yield is an important trait for several agro-economic purposes of lemongrass, such as landscaping, essential oil production, and phytoremediation. Therefore, we performed gene mining of CfARFs and identified 26 and 27 CfARF-encoding genes in each of the haplotype genomes of lemongrass, respectively. Phylogenetic and domain architecture analyses showed that CfARFs can be divided into four groups, among which groups 1, 2, and 3 correspond to activator, repressor, and ETTN-like ARFs, respectively. To identify the CfARFs that may play major roles during the growth of lemongrass plants, RNA-seq was performed on three tissues (leaf, stem, and root) and four developmental stages (3-leaf, 4-leaf, 5-leaf. and mature stages). The expression profiling of CfARFs identified several highly expressed activator and repressor CfARFs and three CfARFs (CfARF3, 18, and 35) with gradually increased levels during leaf growth. Haplotype-resolved transcriptome analysis revealed that biallelic expression dominance is frequent among CfARFs and contributes to their gene expression patterns. In addition, co-expression network analysis identified the modules enriched with CfARFs. By establishing orthologous relationships among CfARFs, sorghum ARFs, and maize ARFs, we showed that CfARFs were mainly expanded by whole-genome duplications, and that the duplicated CfARFs might have been divergent due to differential expression and variations in domains and motifs. Our work provides a detailed catalog of CfARFs in lemongrass, representing a first step toward characterizing CfARF functions, and may be useful in molecular breeding to enhance lemongrass plant growth.

SonicParanoid2: fast, accurate, and comprehensive orthology inference with machine learning and language models

Accurate inference of orthologous genes constitutes a prerequisite for comparative and evolutionary genomics. SonicParanoid is one of the fastest tools for orthology inference; however, its scalability and accuracy have been hampered by time-consuming all-versus-all alignments and the existence of proteins with complex domain architectures. Here, we present a substantial update of SonicParanoid, where a gradient boosting predictor halves the execution time and a language model doubles the recall. Application to empirical large-scale and standardized benchmark datasets shows that SonicParanoid2 is much faster than comparable methods and also the most accurate. SonicParanoid2 is available at https://gitlab.com/salvo981/sonicparanoid2 and https://zenodo.org/doi/10.5281/zenodo.11371108.

Evolutionary paths toward multi-level convergence of lactic acid bacteria in fructose-rich environments

Convergence provides clues to unveil the non-random nature of evolution. Intermediate paths toward convergence inform us of the stochasticity and the constraint of evolutionary processes. Although previous studies have suggested that substantial constraints exist in microevolutionary paths, it remains unclear whether macroevolutionary convergence follows stochastic or constrained paths. Here, we performed comparative genomics for hundreds of lactic acid bacteria (LAB) species, including clades showing a convergent gene repertoire and sharing fructose-rich habitats. By adopting phylogenetic comparative methods we showed that the genomic convergence of distinct fructophilic LAB (FLAB) lineages was caused by parallel losses of more than a hundred orthologs and the gene losses followed significantly similar orders. Our results further suggested that the loss of adhE, a key gene for phenotypic convergence to FLAB, follows a specific evolutionary path of domain architecture decay and amino acid substitutions in multiple LAB lineages sharing fructose-rich habitats. These findings unveiled the constrained evolutionary paths toward the convergence of free-living bacterial clades at the genomic and molecular levels.

Molecular insights into β-Galactosidase enzyme for its potential application in food industry

β-galactosidase generally catalyzes the hydrolysis of β-d-galactosides and α-L-arabinosides. β-galactosidases are well known for its wide range of applications in food industry. In the present study, β-galactosidase from many sources, including archaea, bacteria, fungi, plants, and animals, were analyzed using computational studies such as structural, functional analyses. Sixty sequences were selected for multiple sequence alignment, phylogenetic study, motif and Pfam recognition, identification of physiochemical characteristics. The sequences of β-galactosidase were grouped independently for phylogenetic analysis based on their source. Thermostable proteins were identified through physicochemical analysis. The selected thermostable proteins were then subjected to functional analysis, transmembrane region prediction, secondary structure and domain architecture analysis. The selected highly thermostable sequences were rich in hydrophobic amino acids. Most of the selected β-galactosidase sequences are composed of random coils. The 3D structure of high thermostable bacterial β-galactosidase from Cryobacteriummelibiosiphilum (RJT89331.1) was predicted and the validation results obtained prove that the built structure is of good quality. The modeled structure showed highest similarity with crystal structure of Bifidobacterium longum β-galactosidase. Finally, β-galactosidase from Cryobacterium melibiosiphilum (RJT89331.1) was subjected to docking studies using the CB-DOCK tool with lactose, galactose, galactan, and pectin, with and their binding affinities are −6.6, −5.4, −6.0, −6.4 kJ/mol, respectively. The molecular dynamics simulation was conducted for the docked complex that exhibited the lowest binding energy using NMA (normal mode analysis) to observe the stability and motion of the complex. This computational investigation suggests that the β-galactosidase protein could help in screening β-galactosidase from suitable source for food industry applications.

Amoebae as training grounds for microbial pathogens.

Grazing of amoebae on microorganisms represents one of the oldest predator-prey dynamic relationships in nature. It represents a genetic "melting pot" for an ancient and continuous multi-directional inter- and intra-kingdom horizontal gene transfer between amoebae and its preys, intracellular microbial residents, endosymbionts, and giant viruses, which has shaped the evolution, selection, and adaptation of microbes that evade degradation by predatory amoeba. Unicellular phagocytic amoebae are thought to be the ancient ancestors of macrophages with highly conserved eukaryotic processes. Selection and evolution of microbes within amoeba through their evolution to target highly conserved eukaryotic processes have facilitated the expansion of their host range to mammals, causing various infectious diseases. Legionella and environmental Chlamydia harbor an immense number of eukaryotic-like proteins that are involved in ubiquitin-related processes or are tandem repeats-containing proteins involved in protein-protein and protein-chromatin interactions. Some of these eukaryotic-like proteins exhibit novel domain architecture and novel enzymatic functions absent in mammalian cells, such as ubiquitin ligases, likely acquired from amoebae. Mammalian cells and amoebae may respond similarly to microbial factors that target highly conserved eukaryotic processes, but mammalian cells may undergo an accidental response to amoeba-adapted microbial factors. We discuss specific examples of microbes that have evolved to evade amoeba predation, including the bacterial pathogens- Legionella, Chlamydia, Coxiella, Rickettssia, Francisella, Mycobacteria, Salmonella, Bartonella, Rhodococcus, Pseudomonas, Vibrio, Helicobacter, Campylobacter, and Aliarcobacter. We also discuss the fungi Cryptococcus, and Asperigillus, as well as amoebae mimiviruses/giant viruses. We propose that amoeba-microbe interactions will continue to be a major "training ground" for the evolution, selection, adaptation, and emergence of microbial pathogens equipped with unique pathogenic tools to infect mammalian hosts. However, our progress will continue to be highly dependent on additional genomic, biochemical, and cellular data of unicellular eukaryotes.

Diversification of the Rho transcription termination factor in bacteria.

Correct termination of transcription is essential for gene expression. In bacteria, factor-dependent termination relies on the Rho factor, that classically has three conserved domains. Some bacteria also have a functional insertion region. However, the variation in Rho structure among bacteria has not been analyzed in detail. This study determines the distribution, sequence conservation, and predicted features of Rho factors with diverse domain architectures by analyzing 2730 bacterial genomes. About half (49.8%) of the species analyzed have the typical Escherichia coli like Rho while most of the other species (39.8%) have diverse, atypical forms of Rho. Besides conservation of the main domains, we describe a duplicated RNA-binding domain present in specific species and novel variations in the bicyclomycin binding pocket. The additional regions observed in Rho proteins exhibit remarkable diversity. Commonly, however, they have exceptional amino acid compositions and are predicted to be intrinsically disordered, to undergo phase separation, or have prion-like behavior. Phase separation has recently been shown to play roles in Rho function and bacterial fitness during harsh conditions in one species and this study suggests a more widespread role. In conclusion, diverse atypical Rho factors are broadly distributed among bacteria, suggesting additional cellular roles.

Bacterial protein acetylation: mechanisms, functions, and methods for study.

Lysine acetylation is an evolutionarily conserved protein modification that changes protein functions and plays an essential role in many cellular processes, such as central metabolism, transcriptional regulation, chemotaxis, and pathogen virulence. It can alter DNA binding, enzymatic activity, protein-protein interactions, protein stability, or protein localization. In prokaryotes, lysine acetylation occurs non-enzymatically and by the action of lysine acetyltransferases (KAT). In enzymatic acetylation, KAT transfers the acetyl group from acetyl-CoA (AcCoA) to the lysine side chain. In contrast, acetyl phosphate (AcP) is the acetyl donor of chemical acetylation. Regardless of the acetylation type, the removal of acetyl groups from acetyl lysines occurs only enzymatically by lysine deacetylases (KDAC). KATs are grouped into three main superfamilies based on their catalytic domain sequences and biochemical characteristics of catalysis. Specifically, members of the GNAT are found in eukaryotes and prokaryotes and have a core structural domain architecture. These enzymes can acetylate small molecules, metabolites, peptides, and proteins. This review presents current knowledge of acetylation mechanisms and functional implications in bacterial metabolism, pathogenicity, stress response, translation, and the emerging topic of protein acetylation in the gut microbiome. Additionally, the methods used to elucidate the biological significance of acetylation in bacteria, such as relative quantification and stoichiometry quantification, and the genetic code expansion tool (CGE), are reviewed.

Diversity, abundance, and domain architecture of plant NLR proteins in Fabaceae

The resistance (R) gene family in plants is a vital component of the plant defense system, enabling host resistance against pathogens through interactions with pathogen effector proteins. These R genes often encode nucleotide-binding (NB-ARC or N) and leucine-rich-repeat (LRR or L) domains, collectively forming the NLR protein family. The NLR proteins have been widely explored in crops from Poaceae and Brassicaceae, but limited studies are available for crops in other families, including Fabaceae. To address this gap, we conducted a comprehensive genome-wide analysis of putative NLR proteins in nine Fabaceae crops, including Glycine max, Lupinus angustifolius, Medicago truncatula, Pisum sativum, Phaseolus vulgaris, Trifolium pratense, Vigna angularis, Vigna radiata, and Vigna unguiculata. Our study revealed a substantial variation in the number of NLR proteins, independent of genome size. Notably, the NB-ARC domain exhibited a preferential co-occurrence with a specific LRR domain (IPR001611) in Fabaceae. Furthermore, through protein signature analysis, we identified both species-specific and shared domains across the nine crops. By classifying the identified proteins into seven distinct classes (N, L, CN, TN, NL, CNL, and TNL), we observed species-specific clustering within the CN, TN, and CNL classes, reflecting the diversification of species within Fabaceae. This genome-wide study enhances our understanding of the NLR protein repertoire and comprehensive protein signatures in nine Fabaceae species and provides valuable insights into plant defense mechanisms.

Nutrient Signaling-Dependent Quaternary Structure Remodeling Drives the Catalytic Activation of metazoan PASK.

The Per-Arnt-Sim (PAS) domains are characterized by diverse sequences and feature tandemly arranged PAS and PAS-associated C-terminal (PAC) motifs that fold seamlessly to generate the metabolite-sensing PAS domain. Here, using evolutionary scale sequence, domain mapping, and deep learning-based protein structure analysis, we deconstructed the sequence-structure relationship to unearth a novel example of signal-regulated assembly of PAS and PAC subdomains in metazoan PAS domain-regulated kinase (PASK). By comparing protein sequence, domain architecture, and computational protein models between fish, bird, and mammalian PASK orthologs, we propose the existence of previously unrecognized third PAS domain of PASK (PAS-C) formed through long-range intramolecular interactions between the N-terminal PAS fold and the C-terminal PAC fold. We experimentally validated this novel structural design using residue-level cross-linking assays and showed that the PAS-C domain assembly is nutrient-responsive. Furthermore, by combining structural phylogeny approaches with residue-level cross-linking, we revealed that the PAS-C domain assembly links nutrient sensing with quaternary structure reorganization in PASK, stabilizing the kinase catalytic core of PASK. Thus, PAS-C domain assembly likely integrates environmental signals, thereby relaying sensory information for catalytic control of the PASK kinase domain. In conclusion, we theorize that during their horizontal transfer from bacteria to multicellular organisms, PAS domains gained the capacity to integrate environmental signals through dynamic modulation of PAS and PAC motif interaction, adding a new regulatory layer suited for multicellular systems. We propose that metazoan PAS domains are likely to be more dynamic in integrating sensory information than previously considered, and their structural assembly could be targeted by regulatory signals and exploited to develop therapeutic strategies.

Plant Kinesin Repertoires Expand with New Domain Architecture and Contract with the Loss of Flagella.

Kinesins are eukaryotic microtubule motor proteins subdivided into conserved families with distinct functional roles. While many kinesin families are widespreadin eukaryotes, each organismal lineage maintains a unique kinesin repertoire composed of many families with distinct numbers of genes. Previous genomic surveys indicated that land plant kinesin repertoires differ markedly from other eukaryotes. To determine when repertoires diverged during plant evolution, we performed robust phylogenomic analyses of kinesins in 24 representative plants, two algae, two animals, and one yeast. These analyses show that kinesin repertoires expand and contract coincident with major shifts in the biology of algae and land plants. One kinesin family and five subfamilies, eachdefined by unique domain architectures, emerged in the green algae. Four of those kinesin groups expanded in ancestors of modern land plants, while six other kinesin groups were lost in the ancestors of pollen-bearing plants. Expansions of different kinesin families and subfamilies occurred in moss and angiosperm lineages. Other kinesin families remained stable and did not expand throughout plant evolution. Collectively these data support a radiation of kinesin domain architectures in algae followed by differential positive and negative selection on kinesins families and subfamilies in different lineages of land plants.

Evolutionary origins of archaeal and eukaryotic RNA-guided RNA modification in bacterial IS110 transposons.

Transposase genes are ubiquitous in all domains of life and provide a rich reservoir for the evolution of novel protein functions. Here we report deep evolutionary links between bacterial IS110 transposases, which catalyze RNA-guided DNA recombination using bridge RNAs, and archaeal/eukaryotic Nop5-family proteins, which promote RNA-guided RNA 2'-O-methylation using C/D-box snoRNAs. Based on conservation in the protein primary sequence, domain architecture, and three-dimensional structure, as well as common architectural features of the non-coding RNA components, we propose that programmable RNA modification emerged via exaptation of components derived from IS110-like transposons. Alongside recent studies highlighting the origins of CRISPR-Cas9 and Cas12 in IS605-family transposons, these findings underscore how recurrent domestication events of transposable elements gave rise to complex RNA-guided biological mechanisms.