In E. coli destabilization of RNase R by SmpB was shown to be dependent on previous acetylation of the enzyme. Acetylation only occurs during exponential growth and was proposed to release the C-terminal lysine-rich region of RNase R [29]. This

domain of RNase R is directly bound by SmpB in a tmRNA-dependent manner, and this interaction would ultimately target RNase R for proteolytic degradation [28, 29]. We have analysed the pneumococcal RNase R sequence and also identified a lysine-rich AZD9291 C-terminal domain, which could mediate an association between RNase R and SmpB. It seems reasonable to speculate that in S. pneumoniae, a similar interaction is taking place. Interestingly, the lysine-rich domain of RNase R is essential for the enzyme’s recruitment MLN2238 research buy to ribosomes that are stalled and for its activity on the degradation of defective transcripts [38]. A proper engagement of RNase R is dependent on both functional SmpB and tmRNA, and seems to be determinant for the enzyme’s role in trans-translation. All these observations point to an interaction between the pneumococcal RNase R and SmpB, which may destabilize the exoribonuclease. However, we believe that the strong increment of the rnr mRNA levels detected at 15°C may also account for the final expression levels of RNase R in the cell. A higher amount of mRNA may compensate the low translation levels under

cold-shock. One of the first indications for the involvement of E. coli RNase R in the quality control of proteins was its association with a ribonucleoprotein complex involved in ribosome rescue [39]. This exonuclease was subsequently

shown to be required for the maturation of E. coli tmRNA under cold-shock [12], and for its turnover in C. crescentus and P. syringae[23, 24]. Additional evidences included a direct role in the selective degradation of non-stop mRNAs [2, 27] and destabilization of RNase R by SmpB [28]. In this work we strengthen the functional relationship between RNase R and the trans-translation machinery by demonstrating that RNase R is also implicated in the modulation of SmpB levels. A marked accumulation of both smpB mRNA and SmpB protein was observed in a strain lacking RNase R. The increment in mRNA levels is particularly high at 15°C, the same condition where PLEK2 RNase R expression is higher. This fact suggests that the enzyme is implicated in the control of smpB mRNA levels. The higher smpB mRNA levels detected at 15°C could also suggest a temperature-dependent regulation of this message. However, the steady state levels of SmpB protein in the RNase R- strain were practically the same under cold-shock or at 37°C. Translational arrest caused by the temperature downshift may be responsible for the difference between the protein and RNA levels. Alternatively, we may speculate that the interaction between RNase R and SmpB could also mediate SmpB destabilization.