Characterization of RNA Helicase CshA and Its Role in Protecting mRNAs and Small RNAs of Staphylococcus aureus Strain Newman
The toxin MazFsainStaphylococcus aureusis a sequence-specific endoribonuclease that cleaves the majority of the mRNAsin vivobut spares many essential mRNAs (e.g.,secYmRNA) and, surprisingly, an mRNA encoding a regulatory protein (i.e.,sarAmRNA). We hypothesize that some mRNAs may be protected by RNA-binding protein(s) from degradation by MazFsa. Using heparin-Sepharose-enriched fractions that hybridized tosarAmRNA on Northwestern blots, we identified among multiple proteins the DEAD box RNA helicase CshA (NWMN_1985 or SA1885) by mass spectroscopy. Purified CshA exhibits typical RNA helicase activities, as exemplified by RNA-dependent ATPase activity and unwinding of the DNA-RNA duplex. A severe growth defect was observed in thecshAmutant compared with the parent when grown at 25°C but not at 37°C. Activation of MazFsain thecshAmutant resulted in lower CFU per milliliter accompanied by a precipitous drop in viability (∼40%) compared to those of the parent and complemented strains. NanoString analysis reveals diminished expression of a small number of mRNAs and 22 small RNAs (sRNAs) in thecshAmutant versus the parent upon MazFsainduction, thus implying protection of these RNAs by CshA. In the case of the sRNA teg049 within thesarAlocus, we showed that the protective effect was likely due to transcript stability as revealed by reduced half-life in thecshAmutant versus the parent. Accordingly, CshA likely stabilizes selective mRNAs and sRNAsin vivoand as a result enhancesS. aureussurvival upon MazFsainduction during stress.