Endoribonuclease-mediated control of hns mRNA stability constitutes a key regulatory pathway for Salmonella Typhimurium pathogenicity island 1 expression

Bacteria utilize endoribonuclease-mediated RNA processing and decay to rapidly adapt to environmental changes. Here, we report that the modulation of hns mRNA stability by the endoribonuclease RNase G plays a key role in Salmonella Typhimurium pathogenicity. We found that RNase G determines the half-life of hns mRNA by cleaving its 5′ untranslated region and that altering its cleavage sites by genome editing stabilizes hns mRNA, thus decreasing S. Typhimurium virulence in mice. Under anaerobic conditions, the FNR-mediated transcriptional repression of rnc encoding RNase III, which degrades rng mRNA, and simultaneous induction of rng transcription resulted in rapid hns mRNA degradation, leading to the derepression of genes involved in the Salmonella pathogenicity island 1 (SPI-1) type III secretion system (T3SS). Together, our findings show that RNase III and RNase G levels-mediated control of hns mRNA abundance acts as a regulatory pathway upstream of a complex feed-forward loop for SPI-1 expression.

RNase AM, a 5′ to 3′ exonuclease, matures the 5′ end of all three ribosomal RNAs in E. coli

Bacterial ribosomal RNAs (rRNAs) are transcribed as precursors and require processing by Ribonucleases (RNases) to generate mature and functional rRNAs. Although the initial steps of rRNA processing in Escherichia coli (E. coli) were described several decades ago, the enzymes responsible for the final steps of 5S and 23S rRNA 5′-end maturation have remained unknown. Here, I show that RNase AM, a recently identified 5′ to 3′ exonuclease, performs the last step of 5S rRNA 5′-end maturation. RNase AM was also found to generate the mature 5′ end of 23S rRNA, subsequent to a newly identified prior processing step. Additionally, RNase AM was found to mature the 5′ end of 16S rRNA, a reaction previously attributed to RNase G. These findings indicate a major role for RNase AM in cellular RNA metabolism and establish a biological role for the first 5′ to 3′ RNA exonuclease identified in E. coli.

The coordinated action of RNase III and RNase G controls enolase expression in response to oxygen availability in Escherichia coli

Rapid modulation of RNA function by endoribonucleases during physiological responses to environmental changes is known to be an effective bacterial biochemical adaptation. We report a molecular mechanism underlying the regulation of enolase (eno) expression by two endoribonucleases, RNase G and RNase III, the expression levels of which are modulated by oxygen availability in Escherichia coli. Analyses of transcriptional eno-cat fusion constructs strongly suggested the existence of cis-acting elements in the eno 5′ untranslated region that respond to RNase III and RNase G cellular concentrations. Primer extension and S1 nuclease mapping analyses of eno mRNA in vivo identified three eno mRNA transcripts that are generated in a manner dependent on RNase III expression, one of which was found to accumulate in rng-deleted cells. Moreover, our data suggested that RNase III-mediated cleavage of primary eno mRNA transcripts enhanced Eno protein production, a process that involved putative cis-antisense RNA. We found that decreased RNase G protein abundance coincided with enhanced RNase III expression in E. coli grown anaerobically, leading to enhanced eno expression. Thereby, this posttranscriptional up-regulation of eno expression helps E. coli cells adjust their physiological reactions to oxygen-deficient metabolic modes. Our results revealed a molecular network of coordinated endoribonuclease activity that post-transcriptionally modulates the expression of Eno, a key enzyme in glycolysis.