Acetylation of Response Regulator Protein MtrA in M. tuberculosis Regulates Its Repressor Activity

Abstract

MtrA is an essential response regulator (RR) protein in M. tuberculosis, and its activity is modulated after phosphorylation from its sensor kinase MtrB. Interestingly, many regulatory effects of MtrA have been reported to be independent of its phosphorylation, thereby suggesting alternate mechanisms of regulation of the MtrAB two-component system in M. tuberculosis. Here, we show that RR MtrA undergoes non-enzymatic acetylation through acetyl phosphate, modulating its activities independent of its phosphorylation status. Acetylated MtrA shows increased phosphorylation and enhanced interaction with SK MtrB assessed by phosphotransfer assays and FRET analysis. We also observed that acetylated MtrA loses its DNA-binding ability on gene targets that are otherwise enhanced by phosphorylation. More interestingly, acetylation is the dominant post-translational modification, overriding the effect of phosphorylation. Evaluation of the impact of MtrA and its lysine mutant overexpression on the growth of H37Ra bacteria under different conditions along with the infection studies on alveolar epithelial cells further strengthens the importance of acetylated MtrA protein in regulating the growth of M. tuberculosis. Overall, we show that both acetylation and phosphorylation regulate the activities of RR MtrA on different target genomic regions. We propose here that, although phosphorylation-dependent binding of MtrA drives its repressor activity on oriC and rpf, acetylation of MtrA turns this off and facilitates division in mycobacteria. Our findings, thus, reveal a more complex regulatory role of RR proteins in which multiple post-translational modifications regulate the activities at the levels of interaction with SK and the target gene expression.

Exposure to nitric oxide drives transition to differential culturability inMycobacterium tuberculosis

“…Expression of Rpf coding genes is tightly controlled by multiple regulators [63], Fig. S7, including Lsr2 [64] and MtrA [65]. The precise mechanism underpinning the response to ND is unknown.…”

“…Fig. S7, including Lsr2 [64] and MtrA [65]. The precise mechanism underpinning the response to ND is unknown.…”

Effects of dual deletion ofglnRandmtrAon expression of nitrogen metabolism genes inStreptomyces venezuelae

“…The lower level of MtrA under nitrogen-limited and of GlnR under nutrient-rich conditions conditions could be caused at the transcriptional level and potentially at the post-transcriptional level. In addition to changes in the levels of these regulators, modification at the post-translational level has been reported for GlnR in Streptomyces and MtrA in Mycobacterium tuberculosis (Singh et al, 2020, Singhal et al, 2020, Amin et al, 2016. Acetylated and phosphorylated forms of GlnR have been identified (Amin et al, 2016); GlnR phosphorylation correlated with nitrogen-rich conditions, and phosphorylation inhibited the binding of GlnR to its target genes, whereas acetylation had only a minor influence on the binding of GlnR to its target genes (Amin et al, 2016).…”

“…Acetylation and methylation of MtrA influenced its repressor activity in M. tuberculosis (Singh et al, 2020, Singhal et al, 2020, and MtrA of Streptomyces may be similarly subject to post-translational modification, with different forms of MtrA having different binding affinities for target genes (Singh et al, 2020, Singhal et al, 2020. However, the role of any such modifications needs to be further investigated in Streptomyces.…”

Contribution of N ε -lysine Acetylation towards Regulation of Bacterial Pathogenesis

“…In M. tuberculosis H37Rv, MtrA of the MtrAB two-component system ( 49 , 50 ) enhances its interaction with its cognate sensor kinase MtrB and reduces DNA binding activity. The phosphorylated form of MtrA is a repressor that sequesters oriC to block cell division and blocks expression of resuscitation promoting factor (Rpf), which hydrolyzes peptidoglycan during cell division and is required for reactivation ( 51 – 53 ).…”