Escherichia coli RNase III (rnc) autoregulation occurs independently of rnc gene translation

Abstract

Control of mRNA stability is an established means of regulating gene expression. However, the detailed mechanisms by which such control is achieved are only now emerging. In particular, there remains a question about the involvement of translation. Escherichia coli ribonuclease III (RNase III) negatively autoregulates expression of its own gene (rnc) approximately 10-fold, by cleaving the untranslated leader and initiating approximately 10-fold more rapid decay of the rnc mRNA, after which RNase III plays no further role. Here, we define the mechanism of this control further. Mutations that increase rnc gene translation abolish autoregulation by increasing the stability of the RNase III-cleaved transcript RNA approximately 10-fold, with no effect on the uncleaved species. Mutations that decrease translation destabilize the rnc mRNA in the presence or absence of RNase III. In so doing, they reveal a pathway of rnc transcript decay distinct from the RNase III-dependent pathway. Stability of a 'mini-rnc' transcript containing the rnc leader and only the first two codons of the rnc gene is unaffected by decreased translation, presumably because sequences required for this pathway were removed. Importantly, this mini-rnc transcript is regulated normally by RNase III. Moreover, rnc transcripts synthesized in vitro do not decay in cell-free extracts lacking ribosomes, unless they are first cleaved by RNase III. These two results show that RNase III cleavage can initiate rnc transcript decay independently of rnc gene translation, unambiguously establishing that control of mRNA decay need not involve changes in translation. How rnc gene translation is optimized for efficient autoregulation will also be discussed.