# Involvement of _E. coli_ 6S RNA in Oxidative Stress Response
## 3 Discussion
As alkyl hydroperoxide reductase AhpC, superoxide dismutase SodA and thiol peroxidase Tpx directly degrade H\({}_{2}\)O\({}_{2}\) and related ROS [42; 62; 63], their deficiency very likely contributes to the lethal phenotype of the \(\Delta\)_ssrS_ strain owing to insufficient or slow decomposition of the stress reagent. Finally, the H\({}_{2}\)O\({}_{2}\)-inducible global transcription regulator SoxS targets at least 20 genes [55], including _sodA_ and its own gene [63]. It is, therefore, likely that the attenuated H\({}_{2}\)O\({}_{2}\)-induced decrease in the level of _sodA_ mRNA (and potentially other mRNAs not yet identified) in the \(\Delta\)_ssrS_ strain has its origin in reduced expression of SoxS. Moreover, _alpC_ is under control of OxyR whose mRNA levels were not significantly affected by the lack of 6S RNA, in line with previous observations that _oxyR_ expression is not induced upon H\({}_{2}\)O\({}_{2}\) stress [35]. Thus, abolished H\({}_{2}\)O\({}_{2}\) induction of _alpC_ expression in the \(\Delta ssrS\) strain (Figure 6b) is expected to have other reasons than 6S RNA effects on \(oxyR\) expression. Finally, we performed a comparative proteomic analysis (based on 2D-PAGE) of the \(\Delta ssrS\) and parental _E. coli_ WT strain, which unveiled a number of dysregulated proteins, especially under oxidative stress conditions. Most interesting in the context of this study was, among other proteins, the identification of lower levels of proteins AhpC and Tpx in \(\Delta ssrS\) bacteria (Figure S16, Table S7). This correlates with the trend toward fewer amounts of their mRNAs in \(\Delta ssrS\) versus WT bacteria upon exposure to H\({}_{2}\)O\({}_{2}\) (Figure 6b), despite the fact that qRT-PCR and proteome analysis were performed in different conditions of cell growth for technical reasons (e.g., 20 versus 5 mM H\({}_{2}\)O\({}_{2}\), respectively). Interestingly, the dysregulated proteins identified so far in \(\Delta ssrS\) bacteria included AphA (acid phosphatase) and DsbA (thiol:disulfide interchange protein) whose mRNA levels were also found to be altered in microarray analyses of \(\Delta ssrS\) bacteria in standard growth conditions. Levels of \(alpha\) mRNA were reported to be upregulated (~1.7-fold) and those for \(dsbA\) mRNA to be downregulated (~1.6-fold) in \(\Delta ssrS\) cells [17]. These data perfectly correlate with our findings that AphA protein levels are increased in \(\Delta ssrS\) cells, while DsbA levels decreased (Table S7). Despite the fact that DsbA is not a primary factor of the oxidative stress response, it is yet a key player in the correct folding of proteins within the perplasmic space of _E. coli_ to counteract protein misfolding and aggregation in the presence of elevated ROS levels. Moreover, DsbA is a prospective target for novel antimicrobial agents and antibiotics [64]. Notably, we previously observed that AhpC and Tpx were upregulated in the proteomes of _B. subtilis_ strains that lack either 65-1 or 65-2 RNA, while SodA was only upregulated in the 6S-2 knockout strain [19]. In the recent study investigating 6S RNA-deficient strains derived from an undomesticated _B. subtilis_ strain, \(\Delta\)6S-1&2 RNA double knockout cells also showed a prolonged lag phase under mild oxidative stress conditions [27]. These findings suggest the involvement of 6S RNAs in the regulation of oxidative stress responses in phylogenetically distant bacterial species.