Processing and degradation of regulatory RNAs in Escherichia coli
Final Report Abstract
OxyS is together with RprA and DsrA one of at least three small non-coding RNAs, which affect rpoS expression. It is induced under oxidative stress and reduces the levels of the stationary phase sigma factor RpoS. We analyzed the turn-over of OxyS and rpoS mRNA in early exponential and in stationary growth phase in different E. coli strains to learn more about the mechanisms of processing and about a possible impact of processing on growthdependent regulation. We could not attribute a major role of RNase E, RNase III, PNPase or RNase II on OxyS turn-over in exponential growth phase. Only the simultaneous lack of RNase E, PNPase and RNase II activity resulted in some stabilization of OxyS in exponential growth phase, implying the action of multiple ribonucleases on OxyS turn-over. A major role of RNase E on OxyS stability was observed in stationary phase and was dependent on the presence of the RNA binding protein Hfq and of DsrA, one of the other small RNAs binding to rpoS mRNA. RNase III affects rpoS turn-over only in exponential growth phase and most likely indirectly. Our work revealed that OxyS and rpoS mRNA processing is influenced by different RNases and additional factors like Hfq and DsrA and that the impact of these factors is strongly dependent on growth phase. The sRNA RprA is known to activate rpoS translation in E. coli in an osmolarity-dependent manner. Our study revealed that osmolarity-dependent turn-over of RprA indeed contributes to its osmolarity-dependent abundance. RprA is stabilized by the RNA chaperone Hfq and in absence of Hfq its turn-over is no longer osmolarity-dependent. The stability of the RprA target mRNA rpoS shows a lower extent of osmolarity dependence, which differs from the profile observed for RprA. Thus the effect of sucrose is specific for individual RNAs. We can attribute a role of the endoribonuclease RNase E in turn-over of RprA and an indirect effect of the endoribonuclease III in vivo. In addition RprA is stabilized by the presence of rpoS suggesting that hybrid formation with its target may protect it against ribonucleases. In vitro RprA is cleaved by the RNase E containing degradosome and by RNase III and rpoS interferes with RNase III cleavage. This study demonstrated that environmental parameters can affect the stability of sRNAs and consequently their abundance.
Publications
- (2009) The influence of Hfq and ribonucleases on the stability of the small non-coding RNA OxyS and its target rpoS in E. coli is growth phase dependent. RNA Biol. 6: 584-594
Basineni, S. R., Madhugiri, R., Kolmsee, T., Hengge, R., Klug, G.
- (2010) Turn-over of the small non-coding RNA RprA in E. coli is influenced by osmolarity. Mol. Gen. Genet. 284:307-318
Madhugiri, R., Basineni, S. R., Klug, G.
