A specific endoribonuclease, RNase P, affects gene expression of polycistronic operon mRNAs

Abstract

The rnpA mutation, A49, in Escherichia coli reduces the level of RNase P at 43°C because of a temperature-sensitive mutation in C5 protein, the protein subunit of the enzyme. Microarray analysis reveals the expression of several noncoding intergenic regions that are increased at 43°C compared with 30°C. These regions are substrates for RNase P, and they are cleaved less efficiently than, for example, tRNA precursors. An analysis of the tna, secG, rbs, and his operons, all of which contain RNase P cleavage sites, indicates that RNase P affects gene expression for regions downstream of its cleavage sites.S everal years ago, it was reported that two of the enzymes in the lac operon were produced in different molar quantities, but no direct explanation was provided for this result (1). General speculative hypotheses for differing molar concentrations of proteins coded for in operons have also been suggested (2). Intergenic noncoding RNAs in operon mRNAs in Escherichia coli may provide endoribonuclease sites as one method of allowing production of operon mRNAs and proteins with differing molar concentrations. Products of endoribonuclease cleavage could be degraded rather quickly by an mRNA nuclease degradation mechanism involving RNase E. The small intergenic RNAs could be substrates for RNase P, and they would indicate that RNase P might process many more RNAs than was originally thought.RNase P itself is an essential enzyme responsible for the processing of the 5Ј ends of precursors to tRNA and other naturally occurring stable RNA molecules in E. coli (3). In Salmonella, there is one region in an operon mRNA that appears to be cleaved by RNase P (4). In this report, the noncoding intergenic regions and genes coding for proteins (ORFs) in several operons were cloned separately and examined as substrates for RNase P. Several of the intergenic transcripts were substrates for RNase P. These transcripts accumulate at the restrictive temperature in an E. coli mutant strain thermosensitive for RNase P function. ORFs in the relevant operons were not substrates. A mechanism of determining gene expression is described in which the action of a specific endoribonuclease controls regions downstream of its cleavage site in polycistronic operon mRNAs. Materials and MethodsStrains of E. coli. E. coli A49 and several derivatives of this strain are described in ref. 5 and references therein.Cloning and Nomenclature of Intergenic RNAs. Several intergenic regions, the expression of which was high at 43°C in A49 (5), were chosen for cloning. Clones were amplified by PCR starting generally with the first nucleotide after the stop codon of an upstream gene and the last nucleotide before the start codon of a downstream gene. Clones were numbered according to the length of the sequences cloned and w or c, depending on whether transcription was in the clockwise (w) or counterclockwise (c) direction on the chromosome. The start and stop sites for each fragment on the chromosome are listed below, with the numerology based on that of Blattner ...

Transcriptional linkage of Haloferax volcanii proteasomal genes with non-proteasomal gene neighbours including RNase P, MOSC domain and SAM-methyltransferase homologues

“…These include rnpA and rnpB, which encode the Rpp30 and Pop5 protein subunits of RNaseP, an enzyme responsible for processing the 59 ends of precursors to tRNA as well as cleaving other stable RNAs, intergenic transcripts (Li & Altman, 2003) and riboswitches (Altman et al, 2005). Although the RNA component of the archaeal RNase P alone is capable of catalysing 59-end tRNA maturation, substrate specificity (k cat /K m ) is dramatically enhanced in the presence of Pop5 and Rpp30 (Tsai et al, 2006).…”

Analysis of gene expression from theWolbachiagenome of a filarial nematode supports both metabolic and defensive roles within the symbiosis

“…The elevated expression of RNase P in the soma was also surprising, considering its canonical function in the catalysis of precursor tRNA maturation (Kazantsev and Pace 2006). However, RNase P is a remarkably pleiotropic ribozyme that can regulate the expression of the Sec pathway (Li and Altman 2003) and cleave transient secondary structures in riboswitches (Altman et al 2005). Thus, the loss of three transcriptional regulator proteins in wOo (Fig.…”

Novel function of C5 protein as a metabolic stabilizer of M1 RNA

“…In addition to assisting with tRNA processing, RNase P also contributes to the processing of other non-tRNA substrates (e.g. 4.5S RNA and tmRNA) and assists with decay of several mRNAs [2][3][4][5][6]. The RNase P holoenzyme is composed of two subunits, including a large RNA subunit (M1 RNA, 377 nucleotides) and a small basic protein (C5 protein, 119 amino acids).…”