Processing and turn-over of small non-coding RNA OxyS in E. coli & post-transcriptional regulation of RpoS levels by small non-coding RNAs OxyS and DsrA and the Hfq protein in E. coli

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Basineni_Sobha_Rani_19.10.2010.pdf (2.05 MB)

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2010

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DOI:
http://dx.doi.org/10.22029/jlupub-10171

Abstract

rpoS mRNA is one of the first target RNA shown to be regulated by at least 3 small noncodingRNAs under various stress conditions. Two out of three sRNAs are involved in theactivation of rpoS mRNA translation. By contrast the third sRNA, OxyS has been implicatedin the repression of RpoS synthesis. OxyS RNA is strictly induced under oxidative stress andreduces the levels of the stationary phase sigma factor RpoS. In order to reveal a possibleimpact of processing on growth-dependent regulation, we analyzed the turn-over rates ofOxyS and rpoS mRNA in early exponential and in stationary growth phase in different E. colistrains. We were not able to assign a major role to any of the individual endo- orexoribonucleases (RNase E, RNase III, PNPase or RNase II) on OxyS turn-over inexponential growth phase. However, the simultaneous lack of RNase E, PNPase and RNase IIactivity resulted in some stabilization of OxyS in exponential growth phase, indicating thecombined action of both endo- and exoribonucleases on OxyS turn-over. A greater impact ofRNase E on OxyS turn-over was observed in stationary phase and this in turn was dependenton the presence of the RNA chaperone Hfq and of DsrA sRNAs (activator of rpoS mRNAtranslation). Our data also confirm a role of RNase III in rpoS turn-over, however, only inexponential growth phase. We conclude that OxyS and rpoS mRNA turn-over rates areinfluenced by different RNases and additional factors such as Hfq and DsrA and that theimpact of these factors is strongly dependent on growth phase.Furthermore, we analyzed the influence of OxyS inhibitor RpoS synthesis and Hfq andDsrA activators of RpoS synthesis, while all three regulators co-exist under oxidativestress. Our data reveal that growth phase has a strong effect on RpoS levels, while no majorinfluence of oxidative stress generated by H2O2 is observed. The lack of DsrA resulted in asmall decrease of RpoS levels, which is in agreement with the activating effect of this sRNAon RpoS translation. A lack of Hfq resulted in significantly reduced levels of RpoS, althoughthe turn-over of RpoS was reduced compared to the parental wild type. DsrA and OxySinfluenced RpoS turn-over in opposite way by yet unknown mechanisms. These findingsillustrate that main control of growth-phase dependent RpoS expression is at level of rpoStranscription and that growth phase has a stronger impact on RpoS levels than oxidativestress.

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