Reassessing the role of the secreted protease CPAF in Chlamydia trachomatis infection through genetic approaches. Issue 3 (16th May 2014)

• Record Type: Journal Article
• Authors: Snavely, Emily A.; Kokes, Marcela; Dunn, Joe Dan; Saka, Hector A.; Nguyen, Bidong D.; Bastidas, Robert J.; McCafferty, Dewey G.; Valdivia, Raphael H.
• Is Part Of: Pathogens and disease. Volume 71:Issue 3(2014:Aug.)
• Page Start: 336
• Page End: 351
• Publication Date: 2014-05-16
• Subjects: Medical microbiology -- Periodicals; Pathogenic microorganisms -- Periodicals; Communicable diseases -- Microbiology -- Periodicals; Communicable diseases -- Pathogenesis -- Periodicals; Host-parasite relationships -- Periodicals; Systems biology -- Periodicals; 616.904105
• DOI: 10.1111/2049-632X.12179
• Languages: English
• ISSNs: 2049-632X

Abstract

<abstract abstract-type="main" id="fim12179-abs-0001"> <title>Abstract</title> <p>The secreted <italic>Chlamydia</italic> protease CPAF cleaves a defined set of mammalian and <italic>Chlamydia</italic> proteins <italic>in vitro</italic>. As a result, this protease has been proposed to modulate a range of bacterial and host cellular functions. However, it has recently come into question the extent to which many of its identified substrates constitute <italic>bona fide</italic> targets of proteolysis in infected host cell rather than artifacts of postlysis degradation. Here, we clarify the role played by CPAF in cellular models of infection by analyzing <italic>Chlamydia trachomatis</italic> mutants deficient for CPAF activity. Using reverse genetic approaches, we identified two <italic>C. trachomatis</italic> strains possessing nonsense, loss‐of‐function mutations in <italic>cpa</italic> (CT858) and a third strain containing a mutation in type II secretion (T2S) machinery that inhibited CPAF activity by blocking zymogen secretion and subsequent proteolytic maturation into the active hydrolase. HeLa cells infected with T2S<sup>−</sup> or CPAF<sup>−</sup><italic>C. trachomatis</italic> mutants lacked detectable <italic>in vitro</italic> CPAF proteolytic activity and were not defective for cellular traits that have been previously attributed to CPAF activity, including resistance to staurosporine‐induced apoptosis, Golgi fragmentation, altered NFκB‐dependent gene expression, and resistance to reinfection. However, CPAF‐deficient mutants did display impaired generation of infectious elementary bodies (EBs), indicating an important role for this protease in the full replicative potential of <italic>C. trachomatis</italic>. In addition, we provide compelling evidence in live cells that CPAF‐mediated protein processing of at least two host protein targets, vimentin filaments and the nuclear envelope protein lamin‐associated protein‐1 (LAP1), occurs rapidly after the loss of the inclusion membrane integrity, but before loss of plasma membrane permeability and cell lysis. CPAF‐dependent processing of host proteins correlates with a loss of inclusion membrane integrity, and so we propose that CPAF plays a role late in infection, possibly during the stages leading to the dismantling of the infected cell prior to the release of EBs during cell lysis.</p> </abstract>