Invasive Acinetobacter baumannii ABC141 strain relies on the twin-arginine translocation (Tat) export system for adhesion to host cells
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PROVIDER: PRJNA1256846 | ENA |
REPOSITORIES: ENA
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Project description:Acinetobacter baumannii is associated with severe hospital-acquired multi-drug resistant infections worldwide, causing significant mortality and morbidity in intensive care patients or under prolonged hospitalization. Multiple studies have recently shown that a proportion of circulating clinical isolates establish a transient multiplication niche inside phagocytic and non-phagocytic eukaryotic cells. We have previously demonstrated that the A. baumannii ABC141 strain invades human endothelial and epithelial cells, where it efficiently multiplies without induction of cytotoxicity. Here, we show that ABC141 adhesion, invasion, and intracellular multiplication depend on the growth stage, being most efficient in the exponential growth phase. To define the gene expression signature most favorable to an intracellular lifestyle, a transcriptomic comparison was carried out between exponentially grown ABC141 and cultures at the stationary phase. Although most of the pathways identified reflected growth-related metabolic changes, we observed an upregulation of the twin-arginine translocation (Tat) export system. Analysis of a mutant strain lacking the tatABC operon revealed this secretion system is only required for adhesion to host cells but not invasion and intracellular multiplication. These data highlight a new role for the Tat export pathway in A. baumannii pathogenesis.
2025-08-12 | GSE295948 | GEO
Project description:Achromobacter xylosoxidans is an emerging pathogen, causing airway infections in patients with cystic fibrosis (CF). The knowledge about virulence factors and protein secretion systems in this bacterium is limited. The twin-arginine translocation (Tat) is a protein secretion system for the transportation of folded proteins across the inner cell membrane of Gram-negative bacteria. Here, we report a quantitative proteome analysis of fractionated Achromobacter xylosoxidans wt and Tat-mutant cells in iron-containing and iron-lacking conditions.
2023-06-15 | MSV000092175 | MassIVE
Project description:Brucella, a notorious intracellular pathogen, causes chronic infections in many mammals, including humans. The twin-arginine translocation (Tat) pathway transports folded proteins across the cytoplasmic membrane; protein substrates translocated by Brucella include ABC transporters, oxidoreductases, and cell envelope biosynthesis proteins. Previously, we showed that a Tat mutant of Brucella melitensis M28 exhibits reduced survival within murine macrophages. In this study, we compared the host responses elicited by wild-type M28 and its Tat-mutant strains ex vivo. We utilized label-free quantitative proteomics to assess proteomic changes in RAW264.7 macrophages after infection with M28 and its Tat mutants.
2021-07-05 | PXD024658 | Pride
Project description:Protein secretion is key for bacterial growth, survival and adaptation. In the Gram-positive bacterium Bacillus subtilis, the twin-arginine (Tat) pathway serves in the exclusive secretion of proteins with bound cofactors. The Bacillus Tat pathway stands out for its minimalist nature with two independently acting translocases being composed of dedicated TatA and TatC subunits only. Here we addressed the question whether one of these translocases, TatAyCy, recruits additional common cellular components for activity. To this end, TatAyCy was purified by affinity chromatography and gel filtration, and interacting co-purified proteins were identified by mass spectrometry. This revealed the cell envelope stress responsive LiaH protein as an appendix to the TatAyCy complex. Importantly, our functional studies show that Tat expression is tightly trailed by LiaH induction, and that LiaH itself determines the capacity and quality of Tat-dependent protein translocation. Altogether, our observations show that protein translocation by the minimal Tat translocase TatAyCy is tightly intertwined with an adequate bacterial response to cell envelope stress. This is consistent with a critical need to maintain cellular homeostasis, especially when the membrane is widely opened to permit passage of large fully-folded proteins via Tat.
2022-08-11 | PXD012842 | Pride
Project description:The twin arginine translocation (Tat) pathway transports fully-folded and assembled proteins in bacteria, archaea and plant thylakoids. The Tat pathway contributes to the virulence of numerous bacterial pathogens that cause disease in humans, cattle and poultry. Thus, the Tat pathway has the potential to be a novel therapeutic target. Deciphering the Tat protein transport mechanism has been challenging since the active translocon only assembles transiently in the presence of substrate and a proton motive force. To identify inhibitors of Tat transport that could be used as biochemical tools and possibly as drug development leads, we developed a high throughput screen (HTS) to assay the effects of compounds in chemical libraries against protein export by the Escherichia coli Tat pathway. The primary screen is a live cell assay based on a fluorescent Tat substrate that becomes degraded in the cytoplasm when Tat transport is inhibited. Consequently, low fluorescence in the presence of a putative Tat inhibitor was scored as a hit. Two diverse chemical libraries were screened, yielding average Z'-factors of 0.74 and 0.44, and hit rates of ~0.5% and 0.04%, respectively. Hits were evaluated by a series of secondary screens. Electric field gradient (Δψ) measurements were particularly important since the bacterial Tat transport requires a Δψ. Seven low IC50 hits were eliminated by Δψ assays, suggesting ionophore activity. As Δψ collapse is generally toxic to animal cells and efficient membrane permeability is generally favored during the selection of library compounds, these results suggest that secondary screening of hits against electrochemical effects should be done early during hit validation. Though none of the short-listed compounds inhibited Tat transport directly, the screening and follow-up assays developed provide a roadmap to pursue Tat transport inhibitors.
| S-EPMC4764201 | biostudies-literature
Project description:The Escherichia coli genome encodes at least 29 putative signal peptides containing a twin arginine motif characteristic of proteins exported via the twin arginine translocation (Tat) pathway. Fusions of the putative Tat signal peptides plus six to eight amino acids of the mature proteins to three reporter proteins (short-lived green fluorescent protein, maltose-binding protein (MBP), and alkaline phosphatase) and also data from the cell localization of epitope-tagged full-length proteins were employed to determine the ability of the 29 signal peptides to direct export through the Tat pathway, through the general secretory pathway (Sec), or through both. 27/29 putative signal peptides could export one or more reporter proteins through Tat. Of these, 11 signal peptides displayed Tat specificity in that they could not direct the export of Sec-only reporter proteins. The rest (16/27) were promiscuous and were capable of directing export of the appropriate reporter either via Tat (green fluorescent protein, MBP) or via Sec (PhoA, MBP). Mutations that conferred a >or=+1 charge to the N terminus of the mature protein abolished or drastically reduced routing through the Sec pathway without affecting the ability to export via the Tat pathway. These experiments demonstrate that the charge of the mature protein N terminus affects export promiscuity, independent of the effect of the folding state of the mature protein.
| S-EPMC2730154 | biostudies-literature
Project description:The twin-arginine translocation (Tat) system transports folded proteins across bacterial and plastid energy transducing membranes. Ion leaks are generally considered to be mitigated by the creation and destruction of the translocation conduit in a cargo-dependent manner, a mechanism that enables tight sealing around a wide range of cargo shapes and sizes. In contrast to the variable stoichiometry of the active translocon, the oligomerization state of the receptor complex is considered more consistently stable but has proved stubbornly difficult to establish. Here, using a single molecule photobleaching analysis of individual inverted membrane vesicles, we demonstrate that Tat receptor complexes are tetrameric in native membranes with respect to both TatB and TatC. This establishes a maximal diameter for a resting state closed pore. A large percentage of Tat-deficient vesicles explains the typically low transport efficiencies observed. This individual reaction chamber approach will facilitate examination of the effects of stochastically distributed molecules.
| S-EPMC9485244 | biostudies-literature
Project description:In Escherichia coli, the Twin-arginine (Tat) secretion system is one of the main routes of the protein export to the periplasm. The Tat pathway secretes a set of proteins with important physiological functions. In our study, we investigated the influence of the deactivation of the Tat pathway on the E. coli cells. We applied a comprehensive and comparative proteomic analysis of the E. coli wild type and tat mutant. This dataset provides mass spectrometry based details on the abundances of proteins in cytoplasmic, periplasmic and membrane fractions. We observed that a tat deletion increases abundances of proteins involved in protein folding, degradation, responses to heat, oxidation, osmolarity, and cold. Moreover, the impairment of E. coli outer membrane resulted in the activation of proteins responsible for cell wall biogenesis. The tat deletion negatively affects the synthesis of iron transporters and imbalances its homeostasis in the cell.
2019-02-18 | PXD008803 | Pride
Project description:The twin-arginine translocation (Tat) pathway is a protein transport system for the export of folded proteins. Substrate proteins are targeted to the Tat translocase by N-terminal signal peptides harboring a distinctive R-R-x-Phi-Phi "twin-arginine" amino acid motif. Using a combination of proteomic techniques, the protein contents from the cell wall of the model Gram-positive bacterium Streptomyces coelicolor were identified and compared with that of mutant strains defective in Tat transport. The proteomic experiments pointed to 43 potentially Tat-dependent extracellular proteins. Of these, 25 were verified as bearing bona fide Tat-targeting signal peptides after independent screening with a facile, rapid, and sensitive reporter assay. The identified Tat substrates, among others, include polymer-degrading enzymes, phosphatases, and binding proteins as well as enzymes involved in secondary metabolism. Moreover, in addition to predicted extracellular substrates, putative lipoproteins were shown to be Tat-dependent. This work provides strong experimental evidence that the Tat system is used as a major general export pathway in Streptomyces.
| S-EPMC1693849 | biostudies-literature
Project description:The surface structures of archaeal cells, many of which exist at high temperatures, high salinity, and non-physiological pH, are key factors for their adaptation to extreme living conditions. In the haloarchaeon Haloarcula hispanica, we have discovered a thin filamentous surface appendage called tat-fimbriae ("tafi"), which were identified to be composed of three protein subunits, TafA, TafC, and TafE, among which TafA is the major fimbrial subunit. Molecular genetic evidence demonstrates TafA was transported through the twin-arginine translocation pathway (Tat-pathway). Based on protein structure prediction (including AlphaFold 3), tafi exhibits a linear structure: TafC at the tip, TafE acting as an adapter, TafA forming the core filament, and they link the fourth subunit TafF, anchoring tafi to the cell wall. To our knowledge, this is the first case that the Tat-pathway has been linked to the secretion of protein subunits forming prokaryotic filamentous structures.
| S-EPMC11821415 | biostudies-literature