Transient Requirement of the PrrA-PrrB Two-Component System for Early Intracellular Multiplication of Mycobacterium tuberculosis

Abstract

Adaptive regulation of gene expression in response to environmental changes is a general property of bacterial pathogens. By screening an ordered transposon mutagenesis library of Mycobacterium tuberculosis, we have identified three mutants containing a transposon in the coding sequence or in the 5 regions of genes coding for two-component signal transduction systems (trcS, regX3, prrA). The intracellular multiplication capacity of the three mutants was investigated in mouse bone marrow-derived macrophages. Only the prrA mutant showed a defect in intracellular growth during the early phase of infection, and this defect was fully reverted when the mutant was complemented with prrA-prrB wild-type copies. The mutant phenotype was transient, as after 1 week this strain recovered full growth capacity to reach levels similar to that of the wild type at day 9. Moreover, a transient induction of prrA promoter activity was observed during the initial phase of macrophage infection, as shown by a prrA promoter-gfp fusion in M. bovis BCG infecting the mouse macrophages. The concordant transience of the prrA mutant phenotype and prrA promoter activity indicates that the PrrA-PrrB two-component system is involved in the environmental adaptation of M. tuberculosis, specifically in an early phase of the intracellular growth, and that, similar to other facultative intracellular parasites, M. tuberculosis can use genes temporarily required at different stages in the course of macrophage infection.

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

“…prrA-prrB is expressed by M. tuberculosis H37Rv during growth in human peripheral blood monocytederived macrophages but not during growth in standard laboratory medium, indicating that these genes may be specifically upregulated following infection (54). In agreement with these observations, infection of murine bone marrow-derived macrophages with an M. bovis BCG derivative containing an M. tuberculosis prrA (prrA M. tb )::gfp promoter fusion plasmid is transiently upregulated at 4 h postinfection (38). Consistent with these observations, while strain MT103 with a Tn9563 transposon insertion mutation of prrA exhibits no phenotype during the growth of M. tuberculosis in 7H9 or Sauton's medium, this mutant strain is attenuated during initial time points following infection of murine bone marrow-derived macrophages in vitro (Table 4) (38).…”

“…In agreement with these observations, infection of murine bone marrow-derived macrophages with an M. bovis BCG derivative containing an M. tuberculosis prrA (prrA M. tb )::gfp promoter fusion plasmid is transiently upregulated at 4 h postinfection (38). Consistent with these observations, while strain MT103 with a Tn9563 transposon insertion mutation of prrA exhibits no phenotype during the growth of M. tuberculosis in 7H9 or Sauton's medium, this mutant strain is attenuated during initial time points following infection of murine bone marrow-derived macrophages in vitro (Table 4) (38). Interestingly, growth of the mutant recovers to wild-type levels at later time points, suggesting that signaling through PrrA-PrrB may be important only for early stages of infection in this cell type.…”

High-Throughput Microplate Phosphorylation Assays Based on DevR-DevS/Rv2027c 2-Component Signal Transduction Pathway to Screen for Novel Antitubercular Compounds

“…Cell infection and animal models have shown that many proteins of this class, MprA, PhoP, DevR, KdpDE, TcrXY, TrcS, and SenX3-RegX3, play a role in the survival and multiplication of M. tuberculosis. 15,16,[36][37][38][39][40] Thus, multiple signaling systems could be inhibited by a single molecule targeting the active sites of sensor kinases or response regulators and interfere with pathways vital for M. tuberculosis survival within host tissues. Remove individual filters from the plate and count retained activity or add scintillation fluid to individual wells and count for activity retention in each well Inhibitor for step 3 ( Fig.…”

Modulation of Mycobacterium tuberculosis proliferation by MtrA, an essential two‐component response regulator

“…Of these, only the mtrA-mtrB system is essential (Via et al ., 1996;Zahrt and Deretic, 2000;Parish et al ., 2003a), while others are required under specific growth conditions, namely, mprA-B for maintenance of persistence (Zahrt and Deretic, 2001), devR-S for regulation of hypoxic response (Sherman et al ., 2001;Park et al ., 2003) and prrA-B for intramacrophage growth (Ewann et al ., 2002;. Deletion of regX-sensX , dev R, pho P-R and prrA-B and kdpD-kdp E affects growth and virulence in animal infection models (Perez et al ., 2001;Parish et al ., 2003a,b;Sassetti and Rubin, 2003;Zahrt et al ., 2003;Malhotra et al ., 2004;Roberts et al ., 2004;Tyagi and Sharma, 2004;Asensio et al ., 2006).…”