Identification and characterization of chlamydia trachomatis-secreted proteins.

摘要

Chlamydia trachomatis are Gram-negative, obligate intracellular bacterial pathogens. Chronic inflammation from recurring C.trachomatis infection can lead to severe complications ranging from blindness to pelvic inflammatory disease and infertility. The ability of C.trachomatis to establish intracellular infection and to persist in the infected host significantly contributes to C.trachomatis pathogenicity. To establish and maintain a successful intracellular infection, the C.trachomatis organisms have evolved the ability to secrete proteins into host cells. The secreted proteins may modify host cellular processes and facilitate C.trachomatis invasion, intracellular survival/replication and transmission to new cells. Thus, identification and characterization of secreted proteins have been an area under intensive investigation.;This project was to identify novel Chlamydia trachomatis-secreted proteins, characterize their secretion and explore their biological functions. First, by using anti-chlamydial fusion protein antibodies to localize endogenous chlamydial proteins in chlamydia-infected cells, I identified two chlamydial hypothetical proteins (CT311 and CT795) in host cell cytosol. The cytosolic localization of these two proteins was confirmed independently. Second, the secretions of CT311 and CT795 were monitored over the infection time course and both were secreted at 24 hours post infection and in both cell lines and primary cells. A Sec-dependent secretion pathway was required for secretion of both CT311 and CT795. Third, the biological functions of CT311 and CT795 were explored. CT795 was found to bind host cell COPB and BAT3 in yeast two hybrid and pull down assays. However, the biological relevance of the binding remained unclear since no obvious co-localization of CT795 with COPB in chlamydia-infected Hela cells was detected and knocking down COPI complex subunit COPB or COPE failed to alter chlamydial growth or lipid trafficking to chlamydial inclusions. Interestingly, both CT311 and CT795 were also localized in host nuclei in chlamydial infected cells. The nuclear localization was especially obvious when CT311 and CT795 were overexpressed in mammalian cells. A bipartite nuclear localization signal was identified in CT311 although no clear signal sequence was found in CT795. Surprisingly, nuclear expression of either CT311 or CT795 failed to alter inflammatory signaling pathway or inflammatory cytokine gene expression in mammalian cells with or with chlamydial infection. Efforts are ongoing to further understand the biological relevance of CT311 and CT795 secretion into host cell cytosol and nuclei.