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RNase Y is a key endoribonuclease that regulates global mRNA turnover and processing in Bacillus subtilis and likely many other bacteria. This enzyme is anchored to the cell membrane, creating a pseudo-compartmentalization that aligns with its role in initiating the decay of mRNAs primarily translated at the cell periphery. However, the reasons behind and the consequences of RNase Y's membrane attachment remain largely unknown. In our study, we examined a strain expressing wild-type levels of a cytoplasmic form of RNase Y from its chromosomal locus. This strain exhibits a slow-growth phenotype, similar to that of an RNase Y null mutant. Genome-wide data reveal a significant impact on the expression of hundreds of genes. While certain RNA substrates clearly depend on RNase Y's membrane attachment, others do not. We observed no correlation between mRNA stabilization in the mutant strains and the cellular location or function of the encoded proteins. Interestingly, the Y-complex, a specificity factor for RNase Y, also appears also recognize the cytoplasmic form of the enzyme, restoring wild-type levels of the corresponding transcripts. We propose that membrane attachment of RNase Y is crucial for its functional interaction with many coding and non-coding RNAs, limiting the cleavage of specific substrates, and potentially avoiding unfavorable competition with other ribonucleases like RNase J, which shares a similar evolutionarily conserved cleavage specificity. [ABSTRACT FROM AUTHOR]

The degradation of messenger RNA is of universal importance for controlling gene expression. It directly affects protein synthesis by modulating the amount of mRNA available for translation. Regulation of mRNA decay provides an efficient means to produce just the proteins needed and to rapidly alter patterns of protein synthesis. In bacteria, the half-lives of individual mRNAs can differ by as much as two orders of magnitude, ranging from seconds to an hour. Most of what we know today about the diverse mechanisms of mRNA decay and maturation in prokaryotes comes from studies of the two model organisms Escherichia coli and Bacillus subtilis. Their evolutionary distance provided a large picture of potential pathways and enzymes involved in mRNA turnover. Among them are three ribonucleases, two of which have been discovered only recently, which have a truly general role in the initiating events of mRNA degradation: RNase E, RNase J and RNase Y. Their enzymatic characteristics probably determine the strategies of mRNA metabolism in the organism in which they are present. These ribonucleases are coded, alone or in various combinations, in all prokaryotic genomes, thus reflecting how mRNA turnover has been adapted to different ecological niches throughout evolution.

The Bacillus subtilis valyl-transfer RNA synthetase (ValS) gene was analyzed and sequenced to characterize its transcription pattern and determine the importance of the specifier codon for induction of valine starvation. The expression of the putative Bacillus subtilis valS gene encoding ValS was autoregulated in a specifier-codon-dependent manner. The expression of the recombinant ValS overproducing plasmid into a strain carrying the wild-type valS-lacZ fusion also increased the expression of valS.

An analysis of in vivo and in vitro transcription of the 'house keeping' threonyl-tRNA synthetase gene (thrS) of Bacillus subtilis from a single promoter reveals that thrS is probably autoregulated at the transcriptional level. This is evident from the value of the repression of transcriptional and translational thrS-lacZ fusions by overexpression of thrS. Autogenous regulation does not function at the level of transcription initiation, but includes antitermination of the transcription process.

Abstract Background Formation of alternative structures in mRNA in response to external stimuli, either direct or mediated by proteins or other RNAs, is a major mechanism of regulation of gene expression in bacteria. This mechanism has been studied in detail using experimental and computational approaches in proteobacteria and Firmicutes, but not in other groups of bacteria. Results Comparative analysis of amino acid biosynthesis operons in Actinobacteria resulted in identification of conserved regions upstream of several operons. Classical attenuators were predicted upstream of trp operons in Corynebacterium spp. and Streptomyces spp., and trpS and leuS genes in some Streptomyces spp. Candidate leader peptides with terminators were observed upstream of ilvB genes in Corynebacterium spp., Mycobacterium spp. and Streptomyces spp. Candidate leader peptides without obvious terminators were found upstream of cys operons in Mycobacterium spp. and several other species. A conserved pseudoknot (named LEU element) was identified upstream of leuA operons in most Actinobacteria. Finally, T-boxes likely involved in the regulation of translation initiation were observed upstream of ileS genes from several Actinobacteria. Conclusion The metabolism of tryptophan, cysteine and leucine in Actinobacteria seems to be regulated on the RNA level. In some cases the mechanism is classical attenuation, but in many cases some components of attenuators are missing. The most interesting case seems to be the leuA operon preceded by the LEU element that may fold into a conserved pseudoknot or an alternative structure. A LEU element has been observed in a transposase gene from Bifidobacterium longum, but it is not conserved in genes encoding closely related transposases despite a very high level of protein similarity. One possibility is that the regulatory region of the leuA has been co-opted from some element involved in transposition. Analysis of phylogenetic patterns allowed for identification of ML1624 of M. leprae and its orthologs as the candidate regulatory proteins that may bind to the LEU element. T-boxes upstream of the ileS genes are unusual, as their regulatory mechanism seems to be inhibition of translation initiation via a hairpin sequestering the Shine-Dalgarno box.

Key message: Chlamydomonas RNase J is the first member of this enzyme family that has endo- but no intrinsic 5′ exoribonucleolytic activity. This questions its proposed role in chloroplast mRNA maturation.Abstract: RNA maturation and stability in the chloroplast are controlled by nuclear-encoded ribonucleases and RNA binding proteins. Notably, mRNA 5′ end maturation is thought to be achieved by the combined action of a 5′ exoribonuclease and specific pentatricopeptide repeat proteins (PPR) that block the progression of the nuclease. In Arabidopsis the 5′ exo- and endoribonuclease RNase J has been implicated in this process. Here, we verified the chloroplast localization of the orthologous Chlamydomonas (Cr) RNase J and studied its activity, both in vitro and in vivo in a heterologous B. subtilis system. Our data show that Cr RNase J has endo- but no significant intrinsic 5′ exonuclease activity that would be compatible with its proposed role in mRNA maturation. This is the first example of an RNase J ortholog that does not possess a 5′ exonuclease activity. A yeast two-hybrid screen revealed a number of potential interaction partners but three of the most promising candidates tested, failed to induce the latent exonuclease activity of Cr RNase J. We still favor the hypothesis that Cr RNase J plays an important role in RNA metabolism, but our findings suggest that it rather acts as an endoribonuclease in the chloroplast. [ABSTRACT FROM AUTHOR]