σ^E‐dependent activation of RbpA controls transcription of the furA‐katG operon in response to oxidative stress in mycobacteria

Abstract

Mycobacterium tuberculosis adopts various strategies to cope with oxidative stress during infection. Transcriptional regulators, including σ factors, make important contributions to this stress response, but how these proteins cooperate with each other is largely unknown. In this study, the role of RbpA and its cooperation with σ factors in response to oxidative stress are investigated. Knock down expression of rbpA in Mycobacterium smegmatis attenuated bacterial survival in the presence of H2 O2 . Additionally, transcription of the rbpA gene was induced by H2 O2 in a σ(E) -dependent manner. After induction, RbpA interacts with the principal sigma factor, σ(A) , to control the transcription of furA-katG operon, which encodes an H2 O2 scavenging enzyme. Moreover, this regulation is responsible for the role of σ(E) in oxidative response because bacterial survival was attenuated and transcription of the furA-katG operon was down-regulated with H2 O2 treatment in sigE deletion mutant (ΔsigE), and over-expression of RbpA in ΔsigE strain restored all of these phenotypes. Taken together, our study first illustrated a mechanism for σ(E) in response to oxidative stress through regulation of rbpA transcription. This study was also the first to demonstrate that RbpA is required for the full response to oxidative stress by cooperating with the principal σ(A) .

RbpA and σB association regulates polyphosphate levels to modulate mycobacterial isoniazid‐tolerance

“…To investigate if RbpA was involved in susceptibility, we compared the survival of Msm strains with normal or repressed levels of RbpA in the presence of INH. Since rbpA was essential for mycobacterial growth (Bortoluzzi et al ., ), we constructed a rbpA knockdown strain named RbpAm‐p (Hu et al ., ), in which the rbpA native promoter was replaced by an anhydrotetracycline (ATc) depressed ptr promoter (Supporting Information Fig. S1A).…”

“…S1A). Addition of ATc to RbpAm‐p repressed the mRNA levels of RbpA and bacterial growth (Hu et al ., ), but did not prevent bacterial survival (Supporting Information Fig. S1B).…”

“…The expression of peroxidase catalase (KatG) has been shown to be important for tolerance to INH in mycobacterial cells (Wakamoto et al ., ). In our previous study, we showed that RbpA activates the expression of KatG under oxidative stress conditions (Hu et al ., ). Therefore, we examined if RbpA regulates the participation of KatG in INH‐tolerance by measuring the mRNA levels of katG in Msm WT‐p and RbpAm‐p under normal growth and PBST treated conditions.…”

“…Previous studies have shown that M. smegmatis ( Msm ) (Dey et al ., ) and Streptomyces coelicolor (Newell et al ., ) RbpA participate in rifampicin tolerance. Recently, we have shown that RbpA is also involved in tolerance to oxidative stress probably by its association with σ A in mycobacteria (Hu et al ., ). Nevertheless, the consequences of the association between RbpA and σ B in mycobacteria remain unknown.…”

Promoter mutagenesis for fine‐tuning expression of essential genes in Mycobacterium tuberculosis

“…Since then, the TetR/Pip‐OFF system has been successfully used by several groups to study essential genes and validate drug targets in vitro and in vivo in different mycobacterial species (Serafini et al ., 2009, 2013; Cortes et al ., 2011; Di Luca et al ., 2012; Mondino et al ., 2013; Ventura et al ., 2013; Ahmed et al ., 2014, 2016; Bazet Lyonnet et al ., 2014; Bhowmick et al ., 2014; Boldrin et al ., 2014; Kolly et al ., 2014a,b; Pandey and Rodriguez, 2014; Verma and Chatterji, 2014; Gola et al ., 2015; Gupta et al ., 2015; Mori et al ., 2015; Hu et al ., 2016; Degiacomi et al ., 2017). This broad experience taught us that the main drawback of this otherwise very succesful system was the strength of the P ptr promoter, which causes overexpression of the gene of interest, when this is physiologically expressed at low level, with resulting accumulation of its product.…”