Expression and translocation of chlamydial protease during acute and persistent infection of the epithelial HEp-2 cells with Chlamydophila (Chlamydia) pneumoniae

Abstract

SummaryChlamydial protease-like activity factor (CPAF) is secreted to the cytoplasm of the infected cells where it proteolytically cleaves eukaryotic transcription factor RFX5. Here, we determined the localization pattern of CPAF during the course of an acute and persistent in vitro infection of the epithelial cell line HEp-2 with Chlamydophila pneumoniae strain VR1310. Using immunoblotting, confocal microscopy and electron microscopy, we found CPAF in the inclusion lumen or associated with bacteria during the first 48 h of an acute infection. Seventy-two hours and later, CPAF was present predominantly in the cytoplasm of the infected cells. Translocation of CPAF into cytoplasm correlated in time with degradation of the transcription factor RFX5, as confirmed by immunoblotting. Interestingly, during the persistent infection induced by either IFN-g g g g or iron limitation CPAF translocation to the cytoplasm was inhibited resulting in unaffected or only partially reduced levels of RFX5. Based on presented findings, we propose that CPAF translocation to the cytoplasm is separated from its production. The translocation mechanism appears to be fully active during an acute infection; however, it is fully or partially inhibited during persistent infection induced by IFN-g g g g or by iron limitation respectively. Consequently, this work demonstrates the importance of subcellular localization of CPAF for the characteristics of chlamydial acute and persistent infection in epithelial HEp-2 cells.

Tarp regulates earlyChlamydia-induced host cell survival through interactions with the human adaptor protein SHC1

“…Exact values are provided in Table S1. Although Chlamydia species continue to be refractory to genetic manipulation (Heuer et al, 2003), Tarp's function has been assessed using the heterologous type III secretion system of Yersinia pseudotuberculosis (Clifton et al, 2004). This study revealed a critical role of Tarp in the actin-driven uptake of bacteria by host epithelial cells.…”

Amphisomal Route of MHC Class I Cross-Presentation in Bacteria-Infected Dendritic Cells

“…To this end, CPAF is normally secreted from the chlamydial inclusion into the cytosol of the host cell (45), where it is essential to preserve the integrity of the inclusion (46). In view of our findings in Figs.…”

“…In this context, it was suggested that cytosolic CPAF is required for the chlamydial integral inclusion membrane protein turnover and/or processing of intermediate filaments at the inclusion (46). Heuer et al (45) found that, in situations of restricted chlamydial growth, cytosolic CPAF translocation and its virulence function are impaired. Restricted chlamydial growth in DCs seems to be controlled by TNF-a (58), which is released by Chlamydia-infected DCs and stimulates bactericidal activity by IDO (59).…”

“…9). Indeed, in infected epithelial cells, IFN-g restricts chlamydial growth (85), affects cytoplasmic CPAF translocation (45), rescues surface presentation of MHC I (86), and induces autophagic degradation of Chlamydia (62).…”

Localization of the Hypothetical Protein Cpn0797 in the Cytoplasm of Chlamydia pneumoniae -Infected Host Cells

“…It has been hypothesized that Chlamydia can actively manipulate host signal pathways (6,8,(22)(23)(24)25) via its secreted factors. Indeed, a protein encoded by the hypothetical open reading frame (ORF) Cpn1016 in the C. pneumoniae genome (designated chlamydial protease/proteasome-like activity factor in C. pneumoniae [CPAFcp]) has been shown to degrade host transcription factors required for major histocompatibility complex gene activation (5,13). More recently, it was reported that the N terminus, but not the C terminus, of another C. pneumoniae protein (designated Cpn0796n and Cpn0796c, respectively) encoded by the hypothetical ORF Cpn0796 was secreted into the host cell cytosol, although the biological significance of Cpn0796n secretion is still unclear (24).…”