TheSalmonella entericaSerovar Typhimurium-Encoded Type III Secretion Systems Can TranslocateChlamydia trachomatisProteins into the Cytosol of Host Cells

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Abstract
Chlamydia trachomatisis an obligate, intracellular pathogen that is a major cause of preventable blindness and infertility worldwide. Although the published genome sequence suggests thatC. trachomatisencodes a type III secretion system, the lack of genetic tools for studyingChlamydiahas hindered the examination of this potentially important class of virulence genes. We have developed a technique to identifyChlamydiaproteins that can be translocated into the host cell cytoplasm by a type III secretion system. We have selected severalChlamydiaproteins and tagged them with a multiple peptide motif element called F8M4. Epitopes contained in the F8M4 tag allow us to use tools corresponding to different arms of the adaptive immune system to detect the expression and translocation of these proteins bySalmonella entericaserovar Typhimurium. In particular, CD8+-T-cell reactivity can be used to detect the translocation of F8M4-tagged proteins into the cytoplasm of host cells. We have found that CD8+-T-cell activity assays are sensitive enough to detect translocation of even a small amount of F8M4-tagged protein. We have used CD8+-T-cell activity to show that CopN, aChlamydiaprotein previously shown to be translocated byYersiniatype III secretion, can be translocated by theSalmonellapathogenicity island 1 (SPI-1) type III secretion system. Additionally, we demonstrate that CopD and Pkn5, twoChlamydiaproteins hypothesized to be substrates of a type III secretion system, are translocated via the SPI-2 type III secretion system of serovar Typhimurium. The epitope tag system described here can be used more generally to examine the expression and subcellular compartmentalization of bacterial proteins deployed during the interaction of pathogens with mammalian cells.