Growth Phase-dependent Variation of RNase BN/Z Affects Small RNAs

Abstract

Edited by Charles SamuelRNase BN, the RNase Z family member in E. coli, can participate in the processing of tRNA precursors. However, this function only becomes apparent when other processing enzymes are absent, raising the question of its primary physiological role. Here, we show that RNase BN itself is subject to growth phasedependent regulation, because both rbn mRNA and RNase BN protein are at their highest levels in early exponential phase, but then decrease dramatically and are essentially absent in stationary phase. As a consequence of this variation, certain small RNAs, such as 6S RNA, remain low in exponential phase cells, and increase greatly in stationary phase. RNase BN affects 6S RNA abundance by decreasing its stability in exponential phase. RNase BN levels increase rapidly as cells exit stationary phase and are primarily responsible for the decrease in 6S RNA that accompanies this process. Purified RNase BN directly cleaves 6S RNA as shown by in vitro assays, and the 6S RNA:pRNA duplex is an even more favorable substrate of RNase BN. The exoribonuclease activity of RNase BN is unnecessary because all its action on 6S RNA is due to endonucleolytic cleavages. These data indicate that RNase BN plays an important role in determining levels of the global transcription regulator, 6S RNA, throughout the growth cycle.The RNase Z family of enzymes is widespread among organisms from prokaryotes to eukaryotes. These RNases generally participate in the 3Ј processing of tRNA precursors lacking an encoded CCA sequence, cleaving right after the discriminator nucleotide to generate a substrate for CCA addition by tRNA nucleotidyltransferase (1). In addition to tRNA precursors, some mRNAs have also been identified as substrates including some in rice (2) and Escherichia coli (3), raising the possibility of other putative substrates (4). This is of particular significance in E. coli, because the precursors of all 86 tRNAs already contain the CCA trinucleotide (5), obviating a need for the action of RNase Z.The RNase Z family member in E. coli is termed RNase BN and was originally identified as an enzyme required for maturation of those bacteriophage T4 tRNA precursors that lacked a CCA sequence (6, 7). In contrast to RNase Z in most organisms, RNase BN can act as an exoribonuclease or an endoribonuclease (8, 9). When acting on CCA-containing tRNA precursors, RNase BN cleaves after the CCA sequence using its endoribonuclease activity or trims up to the CCA sequence using its exoribonuclease activity, keeping the CCA sequence intact (10). Although both activities of RNase BN can function in vivo (9), a role for this enzyme in maturation of tRNA precursors only becomes evident when other processing ribonucleases are inactivated (11), suggesting that its primary function in wild type E. coli cells is still unknown.Ribonucleases play an important role in cellular RNA metabolism, and recent studies have revealed that these enzymes may act as important regulators of small RNAs (sRNAs) 2 by participating in their maturati...

Genes Contributing to Porphyromonas gingivalis Fitness in Abscess and Epithelial Cell Colonization Environments

“…RNase Z is widely distributed among bacteria (Condon and Putzer, 2002 ). In E. coli RNase Z plays a significant role in mRNA decay (Perwez and Kushner, 2006 ), and controls the levels of 6S RNA, a stable sRNA, and an important transcription regulator that acts by binding to the sigma 70-containing holoenzyme of RNA polymerase (Chen et al, 2016 ).…”

Characterization of 6S RNA in the Lyme disease spirochete

“…6S RNA is processed from a larger transcript in E. coli by mechanisms involving RNase BN or the endoribonucleases RNase G and RNase E (Chen, Dutta, & Deutscher, ; Kim & Lee, ) and further trimmed by exoribonucleases RNase T and RNase PH (Li, Pandit, & Deutscher, ). B. burgdorferi has a limited repertoire of ribonucleases, compared to E. coli and B. subtilis , and lacks genes encoding RNase G and RNase E; the only predicted endoribonuclease homologs are RNase III, RNase M5, RNase P, RNase Y, RNase Z, YbeY and RNase HII (Anacker, Drecktrah, LeCoultre, Lybecker, & Samuels, ; Archambault, Borchert, Bergeron, Snow, & Schlax, ; Fraser et al, ).…”

“…This is only the second identification of an sRNA-protein interaction in B. burgdorferi (Lybecker, Abel, Feig, & Samuels, 2010;Lybecker & Samuels, 2017). 6S RNA is processed from a larger transcript in E. coli by mechanisms involving RNase BN or the endoribonucleases RNase G and RNase E (Chen, Dutta, & Deutscher, 2016;Kim & Lee, 2004) and further trimmed by exoribonucleases RNase T and RNase PH (Li, Pandit, & Deutscher, 1998 (Anacker, Drecktrah, LeCoultre, Lybecker, & Samuels, 2018;Archambault, Borchert, Bergeron, Snow, & Schlax, 2013;Fraser et al, 1997). The size of Bb6S RNA was assayed by Northern blot in individual RNase mutant strains to investigate Bb6S RNA processing.…”

Porphyromonas gingivalis genes conferring fitness in a tobacco‐rich environment

“…PGN_0388 is an anti‐oxidant thiol peroxidase, thus the conditional essentiality of an oxidative stress gene is established. The PGN_0770 product is RNase Z, a 6S RNA‐/σ70‐related transcriptional regulator, at least in Escherichia coli (Chen, Dutta, & Deutscher, ). The gene targets of PGN_0770 in P. gingivalis , and their relevance to cigarette smoke, have yet to be ascertained.…”