Biology subjects

Andrianova, E. P.

Publications and source records attributed to Andrianova, E. P..

9 recordsLinked to original sources

bioRxiv2026

The last bacterial common ancestor encoded a complex flagellum

Bacterial flagella are rotary nanomachines that enable motility in diverse environments. Although more than 40 genes are required to assemble, operate, and regulate a functional flagellum in model organisms, only 24 flagellar genes have previously been inferred to be conserved across bacteria. This discrepancy raises a fundamental question: did the last bacterial common ancestor encode a simpler, partial flagellum that was elaborated later in a lineage-specific manner, or does the apparent absence of conserved components reflect limitations in detecting highly diverged homologs? Here we combine large-scale profile- and sequence-based searches across a comprehensive bacterial genome set with conserved sequence signatures, gene-tree clustering and flagellar gene-neighborhood evidence to reconstruct the ancestral complexity of bacterial flagellar systems. We identify 28 additional flagellar gene families whose distributions and evolutionary histories support an origin before major bacterial diversification, yielding a 52-gene ancestral flagellum. The ancestral flagellum included all proteins of the secretion/export apparatus, basal body, axial components, motor-force generators and regulatory checkpoints required to build and operate a functional, contemporary flagellum. These findings revise models of early bacterial evolution and overturn the notion that the ancestral flagellum was genetically minimal. Instead, they suggest that the last bacterial common ancestor possessed a highly complex flagellar system comprising more components than are typically found in extant bacteria, many of whose flagella appear to have been shaped by lineage-specific gene loss.

microbiology↗

bioRxiv2025

A cryptic START domain regulates deeply conserved transcription factors

Transcription factors (TFs) integrate a diverse array of inputs to achieve the exquisite control of gene expression necessary for life. In plants, this is exemplified by the deeply conserved CLASS III HOMEODOMAIN LEUCINE ZIPPER (HD-ZIPIII) family of TFs. HD-ZIPIII activity is controlled by inputs at transcriptional, post-transcriptional, and post-translational levels. As part of their multidomain architecture, HD-ZIPIII TFs contain a StAR-related lipid transfer (START) domain, a ubiquitously distributed evolutionary module that binds various types of lipophilic ligands. Here, we show that HD-ZIPIII and HD-ZIPIV proteins contain a cryptic, deeply conserved START domain which we term the disorder-containing START domain (dSTART). The dSTART domain is required for HD-ZIPIII developmental function, controlling their subcellular localization and DNA-binding properties. The dSTART domain also helps discriminate responsive from non-responsive binding sites across the HD-ZIPIII shared genetic network. Finally, we identify candidate ligands of the dSTART domain including several species of phosphatidylglycerol and phosphatidic acid. The identification and functional characterization of a cryptic START domain provides new mechanistic insights into a deeply conserved family of TFs with roles in nearly all aspects of plant development.

plant biology↗

bioRxiv2025

FliO is an evolutionarily conserved yet diversified core component of the bacterial flagellar type III secretion system

The bacterial flagellum is a complex nanomachine essential for motility, environmental sensing, and host colonization. While many of its core components have been well characterized, the relevance of proteins such as FliO, which are inconsistently annotated and poorly conserved at the sequence level, has remained ambiguous in their evolutionary and functional status. Here, we present a comprehensive phylogenomic and structural analysis of FliO across >30,000 representative genomes spanning >100 bacterial phyla. Additionally, during this analysis, we found that approximately 40% of bacterial genomes contain flagellar genes - significantly fewer than previously reported. Using a custom pipeline combining low-threshold HMM searches, operon context analysis, and structural information, we demonstrate that FliO is present in [~]95% of genomes encoding the core flagellar components FliP, FliQ, and FliR. This suggests that FliO is a nearly ubiquitous and ancestral core component of the flagellar type III secretion system (fT3SS). FliO exhibits considerable structural diversity, including lineage-specific acquisitions of LysM and AMIN domains. We identify FliO homologs not only in canonical flagellar systems but also in some virulence-associated T3SS and even some non-flagellated organisms, suggesting functional repurposing and highlighting its functional plasticity. Functional studies in Campylobacter jejuni reveal that FliO and its AMIN domain are critical for efficient bipolar flagellation, membrane stability of the export gate component FlhB, and colonization of the host. These findings establish FliO as a core, yet evolutionarily dynamic, component of flagella and provide new insights into the evolution and diversification of bacterial secretion systems.

microbiology↗

bioRxiv2024

FlhE functions as a chaperone to prevent formation of periplasmic flagella in Gram-negative bacteria

The bacterial flagellum is an organelle utilized by many Gram-negative bacteria to facilitate motility. The flagellum is composed of a several {micro}m long, extracellular filament that is connected to a cytoplasmic rotor-stator complex via a periplasmic rod. Composed of [~]20 structural proteins, ranging from a few subunits to several thousand building blocks, the flagellum is a paradigm of a complex macromolecular structure that utilizes a highly regulated assembly process. This process is governed by multiple checkpoints that ensure an ordered gene expression pattern coupled to the assembly of the various flagellar building blocks in order to produce a functional flagellum. Using epifluorescence, super-resolution STED and transmission electron microscopy, we discovered that in Salmonella, the absence of one periplasmic protein, FlhE, prevents proper flagellar morphogenesis and results in the formation of periplasmic flagella. The periplasmic flagella disrupt cell wall synthesis, leading to a loss of the standard cell morphology resulting in cell lysis. We propose a model where FlhE functions as a periplasmic chaperone to control assembly of the periplasmic rod to prevent formation of periplasmic flagella. Our results highlight that bacteria evolved sophisticated regulatory mechanisms to control proper flagellar assembly and minor deviations from this highly regulated process can cause dramatic physiological consequences.

microbiology↗

bioRxiv2023

Diverse domain architectures of CheA histidine kinase, a central component of bacterial and archaeal chemosensory systems

Chemosensory systems in bacteria and archaea are complex, multi-protein pathways that enable rapid cellular responses to environmental changes. The CheA histidine kinase is a central component of chemosensory systems. In contrast to other histidine kinases, it lacks a sensor (input) domain and utilizes dedicated chemoreceptors for sensing. CheA is a multi-domain protein; in model organisms as diverse as Escherichia coli and Bacillus subtilis, it contains five single-copy domains. Deviations from this canonical domain architecture have been reported, however, a broad genome-wide analysis of CheA diversity is lacking. Here, we present results of a genomic survey of CheA domain composition carried out using an unbiased set of thousands of CheA sequences from bacteria and archaea. We found that four out of five canonical CheA domains comprise a minimal functional unit (core domains), as they are present in all surveyed CheA homologs. The most common deviations from a classical five-domain CheA architecture are the lack of a P2/CheY-binding domain, which is missing from more than a half of CheA homologs and the acquisition of a response regulator receiver (CheY-like) domain, which is present in [~]35% of CheA homologs. We also document other deviations from classical CheA architecture, including bipartite CheA proteins, domain duplications and fusions, and reveal that phylogenetically defined CheA classes have pre-dominant domain architectures. This study lays a foundation for a better classification of CheA homologs and identifies targets for experimental investigations.

microbiology↗

bioRxiv2023

Evolutionary history of MEK1 illuminates the nature of cancer and RASopathy mutations

Mutations in signal transduction pathways lead to various diseases including cancers. MEK1 kinase, encoded by the human MAP2K1 gene, is one of the central components of the MAPK pathway and more than a hundred somatic mutations in MAP2K1 gene were identified in various tumors. Germline mutations deregulating MEK1 also lead to congenital abnormalities, such as the Cardiofaciocutaneous Syndrome and Arteriovenous Malformation. Evaluating variants associated with a disease is a challenge and computational genomic approaches aid in this process. Establishing evolutionary history of a gene improves computational prediction of disease-causing mutations; however, the evolutionary history of MEK1 is not well understood. Here, by revealing a precise evolutionary history of MEK1 we construct a well-defined dataset of MEK1 metazoan orthologs, which provides sufficient depth to distinguish between conserved and variable amino acid positions. We used this dataset to match known and predicted disease-causing and benign mutations to evolutionary changes observed in corresponding amino acid positions. We found that all known and the vast majority of suspected disease-causing mutations are evolutionarily intolerable. We selected several MEK1 mutations that cannot be unambiguously assessed by automated variant prediction tools, but that are confidently identified as evolutionary intolerant and thus "damaging" by our approach, for experimental validation in Drosophila. In all cases, evolutionary intolerant variants caused increased mortality and severe defects in fruit fly embryos confirming their damaging nature predicted by out computational strategy. We anticipate that our analysis will serve as a blueprint to help evaluate known and novel missense variants in MEK1 and that our approach will contribute to improving automated tools for disease-associated variant interpretation. Significance StatementHigh-throughput genome sequencing has significantly improved diagnosis, management, and treatment of genetic diseases and cancers. However, in addition to its indisputable utility, genome sequencing produces many variants that cannot be easily interpreted - so called variants of uncertain significance (VUS). Various automated bioinformatics tools can help predicting functional consequences of VUS, but their accuracy is relatively low. Here, by tracing precise evolutionary history of each amino acid position in MEK1 kinase, mutations in which cause neurodegenerative diseases and cancer in humans, we can establish whether VUS seen in humans are evolutionarily tolerant. Using published data and newly performed experiments in an animal model, we show that evolutionarily tolerable variants in MEK1 are benign, whereas intolerable substitutions are damaging. Our approach will help in diagnostics of MEK1-associated diseases, it is generalizable to many other disease-associated genes, and it can help improving automated predictors.

evolutionary biology↗

bioRxiv2022

TWO DISULFIDE-REDUCING PATHWAYS ARE REQUIRED FOR THE MATURATION OF PLASTID C-TYPE CYTOCHROMES IN CHLAMYDOMONAS REINHARDTII

AO_SCPLOWBSTRACTC_SCPLOWIn plastids, conversion of light energy into ATP relies on cytochrome f, a key electron carrier with a heme covalently attached to a CXXCH motif. Covalent heme attachment requires reduction of the disulfide bonded CXXCH motif by CCS5 and CCS4, a protein of unknown function. CCS5 receives electrons from the oxido-reductase CCDA at the thylakoid membrane. In Chlamydomonas reinhardtii, loss of CCS4 or CCS5 function yields a partial cytochrome f assembly defect. Here we report that the {Delta}ccs4ccs5 double mutant displays a synthetic photosynthetic defect due to a complete loss of holocytochrome f assembly, a phenotype that can be chemically corrected by reducing agents. In {Delta}ccs4, the CCDA protein accumulation is decreased, indicating that one function of CCS4 is to stabilize CCDA. Dominant suppressor mutations mapping to the CCS4 gene were identified in photosynthetic revertants of the {Delta}ccs4ccs5 mutants. The suppressor mutations correspond to changes in the stroma-facing domain of CCS4 and restore holocytochrome f assembly above the residual levels detected in {Delta}ccs5. Because disulfide reduction via CCS5 no longer takes place in {Delta}ccs5, we hypothesize the suppressor mutations enhance the supply of reducing power independently of CCS5, uncovering the participation of CCS4 in a distinct redox pathway. CCS4-like proteins occur in the green lineage and are related to mitochondrial COX16, a protein involved in a disulfide reducing pathway. We discuss the operation of two pathways controlling the redox status of the heme-binding cysteines of apocytochrome f and the possible function of CCS4 as a shared component between the two pathways. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=163 SRC="FIGDIR/small/512171v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@7f496forg.highwire.dtl.DTLVardef@ae5229org.highwire.dtl.DTLVardef@942b20org.highwire.dtl.DTLVardef@19b7576_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract.C_FLOATNO The {Delta}ccs4ccs5 mutant exhibits a photosynthetic growth defect due to a complete loss of cytochrome c assembly. Reduction of apocytochrome f in the thylakoid lumen requires the provision of reducing power through two different pathways, pathway 1 and 2. CCDA and CCS5, components of pathway 1, deliver electrons from stroma to apocytochrome f via thiol - disulfide exchange. CCS4 is involved in pathway 1 by stabilizing CCDA, but also functions through a CCS5 - independent pathway (pathway 2). In the absence of CCS5, gain - of - function mutations in the C terminus of CCS4 (indicated by a yellow star) enhance the delivery of reducing power either via CCDA or independently of CCDA to yet-to-be-discovered reductases C_FIG

genetics↗

bioRxiv2021

Amino acid sensor conserved from bacteria to humans

Amino acids are recognized as signals by various receptors in bacteria, archaea, and eukaryotes. However, no common mechanism for amino acid recognition is currently known. Here we show that a subclass of a ubiquitous extracellular domain dCache_1 contains a simple amino acid recognition motif, and it is found throughout the Tree of Life. In bacteria, this motif exclusively binds amino acids, including GABA, and it is present in all major receptor types. In humans, this motif is found in 2{delta} subunits of voltage-gated calcium channels that are implicated in neuropathic pain and neurodevelopmental disorders. Our findings suggest that GABA-derived drugs bind to the same motif in human 2{delta} subunits that binds natural GABA ligands in bacterial chemoreceptors.

evolutionary biology↗

bioRxiv2021

Phyletic distribution and diversification of the Phage Shock Protein stress response system in bacteria and archaea

The bacterial cell envelope is an essential structure that protects the cell from environmental threats, while simultaneously serving as communication interface and diffusion barrier. Therefore, maintaining cell envelope integrity is of vital importance for all microorganisms. Not surprisingly, evolution has shaped conserved protection networks that connect stress perception, transmembrane signal transduction and mediation of cellular responses upon cell envelope stress. The phage shock protein (PSP) stress response is one of such conserved protection networks. Most of the knowledge about the Psp response comes from studies in the Gram-negative model bacterium, Escherichia coli where the Psp system consists of several well-defined protein components. Homologous systems were identified in representatives of Proteobacteria, Actinobacteria, and Firmicutes; however, the Psp system distribution in the microbial world remains largely unknown. By carrying out a large-scale, unbiased comparative genomics analysis, we found components of the Psp system in many bacterial and archaeal phyla and demonstrated that the PSP system deviates dramatically from the proteobacterial prototype. Two of its core proteins, PspA and PspC, have been integrated in various (often phylum-specifically) conserved protein networks during evolution. Based on protein sequence and gene neighborhood analyses of pspA and pspC homologs, we built a natural classification system of PSP networks in bacteria and archaea. We performed a comprehensive in vivo protein interaction screen for the PSP network newly identified in the Gram-positive model organism Bacillus subtilis and found a strong interconnected PSP response system, illustrating the validity of our approach. Our study highlights the diversity of PSP organization and function across many bacterial and archaeal phyla and will serve as foundation for future studies of this envelope stress response beyond model organisms.

microbiology↗