Chlamydial CT441 Is a PDZ Domain-Containing Tail-Specific Protease That Interferes with the NF-κB Pathway of Immune Response

Abstract

Chlamydia species are bacterial pathogens that affect over 140 million individuals worldwide. Ocular infection by Chlamydia trachomatis is the leading cause of preventable blindness, and urogenital tract infection by Chlamydia causes sexually transmitted disease. As obligate intracellular organisms, Chlamydia species have evolved mechanisms to evade the host immune system, including the degradation of the transcription factors regulatory factor X5 and upstream stimulation factor 1, which are required for the expression of major histocompatibility complex molecules I and II by CPAF and cleavage of p65 of the NF-B pathway by the encoded CT441 protein. Here, we report the characterization of CT441 as a tail-specific protease. CT441 contains a PDZ domain of protein-protein interactions and a Ser/Lys dyad catalytic unit. Mutation at either Ser455 or Lys481 in the active site ablated CT441 activity of p65 cleavage. In addition, we found that the production of CT441 Tsp, which was detected at the middle and late stages of an infection, correlated with p65 cleavage activity. In addition to high homology, human and mouse p65 proteins also contain an identical C-terminal tail of 22 amino acid (aa) residues. However, only human p65 was susceptible to cleavage. Using molecular biology approaches, we mapped the p65 cleavage site(s) to a region that differs from that of mouse p65 by 6 aa residues. Additionally, the substitution of T352 with a proline inhibited p65 cleavage. Together, the study demonstrates that CT441 is a tail-specific protease that is capable of interfering with the NF-B pathway of host antimicrobial and inflammatory responses.The carboxyl-terminal processing proteases (Ctp), including the bacterial tail-specific protease (Tsp), are a group of endoproteases of posttranslational protein modification, maturation, and disassembly or degradation. The Ctp proteases have been found in Archaea, plant chloroplasts, bacteria (reviewed in reference 20), and viruses (5). A well-characterized Ctp is the P1D protease that contains a PDZ domain of proteinprotein interaction and a domain of the S41B family peptidase (15,20). P1D catalyzes the C-terminal processing of the D1 protein of photosystem II, an essential event for the consequent water oxidation and the generation of oxygen molecules in oxygenic photosynthetic organisms. Tail-specific proteases have been identified from bacterial pathogens of medical importance, including Borrelia,

The lone S41 family C-terminal processing protease in Staphylococcus aureus is localized to the cell wall and contributes to virulence

“…either in Gram-negative bacteria (where many have been shown to affect virulence) or in non-pathogenic Grampositive species (where one example is known to affect sporulation) (Bandara et al, 2008;Lad et al, 2007;Marasco et al, 1996;Ostberg et al, 2004;Pan et al, 2003). In this work we have, for the first time, to our knowledge, identified and characterized a C-terminal protease from a Grampositive, pathogenic bacterial species.…”

“…Recent work by ourselves and others has shown that amino terminal peptidases and CTPs can play an important role in bacterial virulence (Bandara et al, 2008;Carroll et al, 2012Carroll et al, , 2013Lad et al, 2007;Wang et al, 2012). These findings, along with the observation that C-terminal processing peptidases are poorly understood in Gram-positive pathogens, led us to investigate the role of C-terminal processing peptidases in S. aureus.…”

“…Several of these studies have shown that mutation of ctpA leads to impaired interactions with components of the host immune system (Bandara et al, 2005(Bandara et al, , 2008Lad et al, 2007;Wang et al, 2012). To investigate if mutation of ctpA affects the interaction of S. aureus with components of the human immune system, we examined the survival of the ctpA 2 mutant in the presence of a variety of immune components.…”

“…Altered cell morphology, differences in osmotic/thermal stress resistance and altered susceptibility to antibiotics are common phenotypes of mutants from different species (and may result from alterations in the cell-wall integrity) (Hara et al, 1991;Kumru et al, 2011;Seoane et al, 1992). In addition, a number of Gram-negative bacteria, including E. coli, Brucella suis, Chlamydia trachomatis, Salmonella typhimurium, Pseudomonas aeruginosa and Burkholderia mallei, demonstrate reduced virulence upon inactivation of CTP (Bandara et al, 2005(Bandara et al, , 2008Bäumler et al, 1994;Lad et al, 2007;Seo & Darwin, 2013;Wang et al, 2012). An E. coli prc mutant demonstrates decreased resistance to complement-mediated killing, while in Brucella suis, Burkholderia mallei and S. typhimurium, CtpA has been implicated in the ability of bacteria to survive in the intracellular environment.…”

Production, crystallization and X-ray diffraction analysis of the protease CT441 fromChlamydia trachomatis

“…CT441 is a chlamydial tail-specific protease (Tsp) with a catalytic serine and a PDZ domain. It has been reported to interfere with the pro-inflammatory NF-B signalling pathway of the host cell, modulating antimicrobial and inflammatory responses Lad, Yang et al, 2007). However, in subsequent reports, the role of CT441 in the cleavage of NF-B has been questioned (Chen et al, 2012;Christian et al, 2010).…”