Messenger RNA degradation represents an essential lever for the control of gene expression in bacteria, where mRNA half-lives can range from a few seconds to more than an hour. In Bacillus subtilis, this degradation is largely initiated by a major endoribonuclease, RNase Y, which cleaves its substrates at single-stranded sequences rich in adenine and uracil. This enzyme influences the overall stability of the transcriptome: the intracellular abundance of most transcripts is altered upon its depletion. Consequently, the amount of RNase Y present in the cell must itself be finely tuned. The present work establishes that the enzyme regulates its own synthesis by modulating the longevity of its messenger RNA, the rny transcript.
To dissect this mechanism, the team used translational rny-lacZ fusions of various lengths, combining β-galactosidase activity measurements with transcript quantification by Northern blot. This approach proved particularly informative: the fusion encoding the first 210 amino acids of RNase Y behaves very similarly to the wild-type rny transcript. The autoregulation appears extremely sensitive, since an overexpression of only 1.5-fold is sufficient to strongly reduce rny mRNA levels, whereas these same transcripts accumulate in a strain lacking the enzyme. The authors show that this regulation is direct and strictly requires endonucleolytic activity: a catalytic mutant, even when strongly overexpressed, does not decrease rny expression but instead exerts a dominant-negative effect.
Two regions of the transcript are targeted. On the one hand, cleavages within the first hundred nucleotides of the open reading frame immediately inactivate the mRNA for any new round of translation; a fragment containing only 39 nucleotides (13 codons) of rny sequence is already sufficient to render the transcript susceptible to the enzyme. On the other hand, cleavages in the 5' untranslated region, mainly within the first fifty nucleotides, create entry sites for the 5' exonuclease RNase J1. The progression of the latter is halted around position −15, presumably by initiating ribosomes bound to a particularly strong Shine-Dalgarno sequence. Transcript inactivation is thus coupled to translation efficiency, according to the ribosomal occupancy of the initiation site.
This model contrasts with that proposed for the autoregulation of RNase E in Escherichia coli, where a stem-loop structure in the 5' untranslated region acts without being cleaved. Through these combined mechanisms, RNase Y can initiate the degradation of its own mRNA when it is not engaged by other substrates, thereby avoiding its overproduction beyond the needs of RNA metabolism. The authors emphasize that the precise role of secondary structures at the translation initiation site, as well as the possible interference of other partner factors, remains to be elucidated.