PepD Participates in the Mycobacterial Stress Response Mediated through MprAB and SigE

Abstract

Currently, one-third of the world's population is believed to be latently infected with Mycobacterium tuberculosis. The mechanisms by which M. tuberculosis establishes latent infection remain largely undefined. mprAB encodes a two-component signal transduction system required by M. tuberculosis for aspects of persistent infection. MprAB regulates a large and diverse group of genetic determinants in response to membrane stress, including the extracytoplasmic function (ECF) sigma factor sigE and the HtrA-like serine protease pepD. Recent studies have demonstrated that PepD functions as both a protease and chaperone in vitro. In addition, inactivation of pepD alters the virulence of M. tuberculosis in a mouse model system of infection. Here, we demonstrate that PepD plays an important role in the stress response network of Mycobacterium mediated through MprAB and SigE. In particular, we demonstrate that the protease activity of PepD requires the PDZ domain, in addition to the catalytic serine at position 317. pepD expression initiates from at least three promoters in M. tuberculosis, including one that is regulated by SigE and is located upstream of the mprA coding sequence. Deletion of pepD or mprAB in Mycobacterium smegmatis and M. tuberculosis alters the stress response phenotypes of these strains, including increasing sensitivity to SDS and cell wall antibiotics and upregulating the expression of stress-responsive determinants, including sigE. Taking these data together, we hypothesize that PepD utilizes its PDZ domain to recognize and process misfolded proteins at the cell membrane, leading to activation of the MprAB and SigE signaling pathways and subsequent establishment of a positive feedback loop that facilitates bacterial adaptation.Currently, one-third of the world's population is believed to be latently infected with Mycobacterium tuberculosis, the causative agent of tuberculosis. There are an estimated 9 million new cases of infection caused by this organism each year, accounting for more than 2 million deaths annually (3). This makes M. tuberculosis one of the leading causes of death due to a single infectious agent. Humans are the only known reservoir of M. tuberculosis, and as such, part of its success as a pathogen lies in its ability to maintain a significant persistent reservoir within the population. Latently infected individuals are asymptomatic and are harder to treat, thereby complicating measures to control the spread of disease (27). In addition, the emergence of multi-and extensively drug-resistant strains has further compromised established strategies for treatment. A better understanding of the mechanisms by which M. tuberculosis establishes persistent infection is urgently needed so that better therapeutics for the latently infected population can be developed.M. tuberculosis is a facultative intracellular pathogen that primarily infects alveolar macrophages. The organism survives within these cells by blocking key steps in the phagosome maturation process (38). Infection by M. tubercul...

Adaptation to Environmental Stimuli within the Host: Two-Component Signal Transduction Systems of Mycobacterium tuberculosis

“…Regulation of sigE by MprA seems especially critical, since MprA and SigE participate in a positive-feedback loop in response to cell wall/membrane-perturbing agents, such as SDS (63,101,113,173,174). Consistent with this observation, both mprA and sigE mutants exhibit increased sensitivity to SDS (101,173).…”

“…MprA positively regulates its own expression and that of downstream genes pepD and moaB2 by directly binding a 17-bp sequence containing two 6-bp direct-repeat motifs separated by 5 nucleotides (Table 3) (64,112,173). MprA also directly regulates downstream genes pepD and moaB2 by binding three tandemly positioned MprA recognition sequences located in the end of mprB and extending into the mprB-pepD intergenic region (64,173), and MprA directly regulates, apart from its own locus, the expression of other determinants, including those encoding the Acr2 alpha-crystallin-like protein (112), a predicted 18-kDa chaperone.…”

MprAB Regulates theespAOperon in Mycobacterium tuberculosis and Modulates ESX-1 Function and Host Cytokine Response

“…MprAB activates the response to envelope stresses through sigma factor SigE and the SigE regulon (22,31). Genes of the MprAB/SigE regulons are induced by agents or environments that impair the integrity of the cell envelope (22,31,35,(44)(45)(46), and sigE mutants are more susceptible to envelope stress, including thioridazine (42,44,45). Using real-time PCR and promoter fusion constructs, we determined that espA expression was increased in our mprAB deletion mutant Rv-D981.…”

“…MprAB activates the response to envelope stress induced by surfactants (22,31). The response regulator MprA binds a repeated hexamer motif (MprA box) found primarily in the promoters of genes associated with envelope stress (22,(31)(32)(33)(34)(35). The majority of these promoters are activated by MprAB, but MprAB can also repress expression under some conditions (34).…”

Mycobacterium tuberculosis Transcription Machinery: Ready To Respond to Host Attacks

“…• The MprAB TCS regulates expression of a subset of genes in the DosR regulon, the stress-responsive chaperone pepD, and the espA operon, which encodes ESX-1 substrates (115)(116)(117)(118). The MprAB TCS also activates expression of sigB and sigE in response to envelope stress and indirectly regulates the stringent response mediator M. tuberculosis rel gene (rel Mtb ) through E activity (119,120).…”