Structure and Function of the Virulence-Associated High-Temperature Requirement A of <i>Mycobacterium tuberculosis</i>

Mohamedmohaideen, Nilofar N.; Palaninathan, S.K.; Morin, Paul; Williams, Brad J.; Braunstein, Miriam; Tichy, Shane E.; Locker, Joseph; Russell, David H.; Jacobs, William R.; Sacchettini, James C.

doi:10.1021/bi701929m

Cited by 58 publications

(88 citation statements)

References 59 publications

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“…⌬pepD::Hyg r mutants of M. tuberculosis Erdman are altered for virulence in mice. In particular, ⌬pepD::Hyg r mutant-infected animals exhibit increased time to death, and infection by ⌬pepD::Hyg r mutants induces less tissue pathology than in animals infected with wild-type or complemented mutant derivatives (33). Taken together, these results suggest that PepD is required for aspects of M. tuberculosis adaptation within the host.…”

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confidence: 79%

“…A subset of HtrA family members is also required for aspects of pathogenesis in both Gram-positive and Gramnegative organisms (14,16,45,49,57). M. tuberculosis possesses three HtrA-like proteases: Rv1223 (degP or htrA1), Rv0983 (pepD or htrA2), and Rv0215 (pepA or htrA3) (8,33), all of which share regions of sequence homology with one another. Initial studies of PepD and PepA in M. tuberculosis indicate that these proteins are secreted and are present within the culture filtrate (47).…”

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confidence: 99%

“…Initial studies of PepD and PepA in M. tuberculosis indicate that these proteins are secreted and are present within the culture filtrate (47). More recently, PepD has been shown to possess both protease and chaperone activities (33); however, the biological activities of DegP and PepA remain unclear. ⌬pepD::Hyg r mutants of M. tuberculosis Erdman are altered for virulence in mice.…”

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

White

Penoske

et al. 2010

J Bacteriol

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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...

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

White

Penoske

et al. 2010

J Bacteriol

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“…At the position corresponding to residue 207 in E. coli DegP, many orthologs have a wild-type proline (Mohamedmohaideen et al 2008;Cezairliyan and Sauer 2009). Based on our results, any potential deleterious effects of this ''activating'' proline are likely to be balanced by one or more ''inactivating'' residues elsewhere in the structure.…”

Section: A Critical Allosteric Balancementioning

confidence: 99%

Distinct regulatory mechanisms balance DegP proteolysis to maintain cellular fitness during heat stress

Kim

Sauer

2014

Genes Dev.

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Intracellular proteases combat proteotoxic stress by degrading damaged proteins, but their activity must be carefully controlled to maintain cellular fitness. The activity of Escherichia coli DegP, a highly conserved periplasmic protease, is regulated by substrate-dependent allosteric transformations between inactive and active trimer conformations and by the formation of polyhedral cages that confine the active sites within a proteolytic chamber. Here, we investigate how these distinct control mechanisms contribute to bacterial fitness under heat stress. We found that mutations that increase or decrease the equilibrium population of active DegP trimers reduce high-temperature fitness, that a mutation that blocks cage formation causes a mild fitness decrease, and that combining mutations that stabilize active DegP and block cage formation generates a lethal rogue protease. This lethality is suppressed by an extragenic mutation that prevents covalent attachment of an abundant outermembrane lipoprotein to peptidoglycan and makes this protein an inhibitor of the rogue protease. Lethality is also suppressed by intragenic mutations that stabilize inactive DegP trimers. In combination, our results suggest that allosteric control of active and inactive conformations is the primary mechanism that regulates DegP proteolysis and fitness, with cage formation providing an additional layer of cellular protection against excessive protease activity.

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“…The activation mechanism of DegS (another E. coli member of the HtrA family) is less defined, as DegS appears to also be activated by a DegP-like 'PDZ -L3' interaction [23], although alternate structural studies suggest that the activation occurs upon direct detection of the substrate C-terminus via a 'C-terminus -L3' interaction [24]. The Mycobacterium tuberculosis HtrA2 protein (MtHtrA2) has only been shown to form trimers in solution and remains in a constantly activated state with its PDZ1 domain appearing to bind autoproteolysis products [25]. This mechanistic diversity within the HtrA family is remarkable considering their structural homogeneity [21].…”

Section: Introductionmentioning

confidence: 99%

Proteolytic activation of Chlamydia trachomatis HTRA is mediated by PDZ1 domain interactions with protease domain loops L3 and LC and beta strand β5

Marsh

Lott

Tyndall

et al. 2013

Cellular and Molecular Biology Letters

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Abbreviations used: CD -circular dichroism; CtHtrA -Chlamydia trachomatis high temperature requirement A; MCA -7-methoxycoumarin-4-acetic acid; MOMP -major outer membrane protein; MRW -mean residue weight; OmpA -outer membrane protein A; PDZ -PSD-95/Dics-Large/ZO-1; PmpC -polymorphic membrane protein C; pNApara-nitroalinine; SDM -site-directed mutagenesis; SDS-PAGE -sodium dodecyl sulphate polyacrylamide gel electrophoresis Abstract: Chlamydia trachomatis is a bacterial pathogen responsible for one of the most prevalent sexually transmitted infections worldwide. Its unique development cycle has limited our understanding of its pathogenic mechanisms. However, CtHtrA has recently been identified as a potential C. trachomatis virulence factor. CtHtrA is a tightly regulated quality control protein with a monomeric structural unit comprised of a chymotrypsin-like protease domain and two PDZ domains. Activation of proteolytic activity relies on the C-terminus of the substrate allosterically binding to the PDZ1 domain, which triggers subsequent conformational change and oligomerization of the protein into 24-mers enabling proteolysis. This activation is mediated by a cascade of precise structural arrangements, but the specific CtHtrA residues and structural elements required to facilitate activation are unknown. Using in vitro analysis guided by homology modeling, we show that the mutation of residues Arg362 and Arg224, predicted to disrupt the interaction between the CtHtrA PDZ1 Unauthenticated Download Date | 5/13/18 1:07 AM CELLULAR & MOLECULAR BIOLOGY LETTERS 523 domain and loop L3, and between loop L3 and loop LD, respectively, are critical for the activation of proteolytic activity. We also demonstrate that mutation to residues Arg299 and Lys160, predicted to disrupt PDZ1 domain interactions with protease loop LC and strand β5, are also able to influence proteolysis, implying their involvement in the CtHtrA mechanism of activation. This is the first investigation of protease loop LC and strand β5 with respect to their potential interactions with the PDZ1 domain. Given their high level of conservation in bacterial HtrA, these structural elements may be equally significant in the activation mechanism of DegP and other HtrA family members.

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Structure and Function of the Virulence-Associated High-Temperature Requirement A of Mycobacterium tuberculosis

Cited by 58 publications

References 59 publications

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

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

Distinct regulatory mechanisms balance DegP proteolysis to maintain cellular fitness during heat stress

Proteolytic activation of Chlamydia trachomatis HTRA is mediated by PDZ1 domain interactions with protease domain loops L3 and LC and beta strand β5

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