The Chlamydia trachomatis type III secretion chaperone Slc1 engages multiple early effectors, including TepP, a tyrosine-phosphorylated protein required for the recruitment of CrkI-II to nascent inclusions and innate immune signaling
| Autor: | Raphael H. Valdivia, Hector Alex Saka, Yi-Shan Chen, Victoria K. Carpenter, Gregory V. Plano, Kristian L. Richards, Robert J. Bastidas |
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| Rok vydání: | 2013 |
| Předmět: |
Scaffold protein
Fluorescent Antibody Technique Chlamydia trachomatis medicine.disease_cause purl.org/becyt/ford/1 [https] Adapter molecule crk Tandem Mass Spectrometry Molecular Cell Biology Tyrosine Kinase Signaling Cascade Gram Negative Phosphorylation lcsh:QH301-705.5 Host factor Oligonucleotide Array Sequence Analysis Effector Proto-Oncogene Proteins c-crk Signaling Cascades 3. Good health Cell biology Bacterial Pathogens Host-Pathogen Interaction CHLAMYDIA PROTEOMICS CIENCIAS NATURALES Y EXACTAS Signal Transduction Research Article lcsh:Immunologic diseases. Allergy Immunology Molecular Sequence Data Biology Real-Time Polymerase Chain Reaction Microbiology Ciencias Biológicas Biología Celular Microbiología Genetic Mutation Virology medicine Genetics Humans Immunoprecipitation Secretion Amino Acid Sequence purl.org/becyt/ford/1.6 [https] Molecular Biology Microbial Pathogens Innate immune system TYPE THREE SECRETION Molecular biology Immunity Innate lcsh:Biology (General) Mutagenesis VIRULENCE Parasitology lcsh:RC581-607 Chromatography Liquid HeLa Cells Molecular Chaperones |
| Zdroj: | PLoS Pathogens CONICET Digital (CONICET) Consejo Nacional de Investigaciones Científicas y Técnicas instacron:CONICET PLoS Pathogens, Vol 10, Iss 2, p e1003954 (2014) |
| ISSN: | 1553-7374 |
| Popis: | Chlamydia trachomatis, the causative agent of trachoma and sexually transmitted infections, employs a type III secretion (T3S) system to deliver effector proteins into host epithelial cells to establish a replicative vacuole. Aside from the phosphoprotein TARP, a Chlamydia effector that promotes actin re-arrangements, very few factors mediating bacterial entry and early inclusion establishment have been characterized. Like many T3S effectors, TARP requires a chaperone (Slc1) for efficient translocation into host cells. In this study, we defined proteins that associate with Slc1 in invasive C. trachomatis elementary bodies (EB) by immunoprecipitation coupled with mass spectrometry. We identified Ct875, a new Slc1 client protein and T3S effector, which we renamed TepP (Translocated early phosphoprotein). We provide evidence that T3S effectors form large molecular weight complexes with Scl1 in vitro and that Slc1 enhances their T3S-dependent secretion in a heterologous Yersinia T3S system. We demonstrate that TepP is translocated early during bacterial entry into epithelial cells and is phosphorylated at tyrosine residues by host kinases. However, TepP phosphorylation occurs later than TARP, which together with the finding that Slc1 preferentially engages TARP in EBs leads us to postulate that these effectors are translocated into the host cell at different stages during C. trachomatis invasion. TepP co-immunoprecipitated with the scaffolding proteins CrkI-II during infection and Crk was recruited to EBs at entry sites where it remained associated with nascent inclusions. Importantly, C. trachomatis mutants lacking TepP failed to recruit CrkI-II to inclusions, providing genetic confirmation of a direct role for this effector in the recruitment of a host factor. Finally, endocervical epithelial cells infected with a tepP mutant showed altered expression of a subset of genes associated with innate immune responses. We propose a model wherein TepP acts downstream of TARP to recruit scaffolding proteins at entry sites to initiate and amplify signaling cascades important for the regulation of innate immune responses to Chlamydia. Author Summary Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes a range of human diseases of significant public health importance. To create a suitable replicative niche within its host, Chlamydia delivers effector proteins across mammalian membranes via a syringe-like apparatus termed a Type III secretion (T3S) system. The lack of a robust system for the molecular genetic manipulation of these pathogens has hindered progress in identifying and characterizing T3S effectors. In this study, we took a mass spectrometry-based approach to identify Chlamydia effector proteins based on their interaction with Slc1, an abundant T3S chaperone. We identified a previously uncharacterized protein, Ct875/TepP, as a new T3S effector and determined that TepP is phosphorylated upon translocation into host cells, leading to the recruitment of the host scaffolding protein Crk and presumably manipulating Crk-dependent signaling functions. Finally, we provide genetic confirmation of the role of TepP in recruiting Crk and in modulating the expression of genes involved in innate immune responses to Chlamydia. This study is the first example of genetic validation of the function of a T3S effector in Chlamydia and a new example of a bacterial effector that directly co-opts the oncoprotein Crk to modulate host cell signaling events. |
| Databáze: | OpenAIRE |
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