RNase Y in Bacillus subtilis: a Natively Disordered Protein That Is the Functional Equivalent of RNase E from Escherichia coli

Abstract

The control of mRNA stability is an important component of regulation in bacteria. Processing and degradation of mRNAs are initiated by an endonucleolytic attack, and the cleavage products are processively degraded by exoribonucleases. In many bacteria, these RNases, as well as RNA helicases and other proteins, are organized in a protein complex called the RNA degradosome. In Escherichia coli, the RNA degradosome is assembled around the essential endoribonuclease E. In Bacillus subtilis, the recently discovered essential endoribonuclease RNase Y is involved in the initiation of RNA degradation. Moreover, RNase Y interacts with other RNases, the RNA helicase CshA, and the glycolytic enzymes enolase and phosphofructokinase in a degradosome-like complex. In this work, we have studied the domain organization of RNase Y and the contribution of the domains to protein-protein interactions. We provide evidence for the physical interaction between RNase Y and the degradosome partners in vivo. We present experimental and bioinformatic data which indicate that the RNase Y contains significant regions of intrinsic disorder and discuss the possible functional implications of this finding. The localization of RNase Y in the membrane is essential both for the viability of B. subtilis and for all interactions that involve RNase Y. The results presented in this study provide novel evidence for the idea that RNase Y is the functional equivalent of RNase E, even though the two enzymes do not share any sequence similarity. mRNAs transmit the genetic information from DNA to proteins. In contrast to the high stability of tRNA and rRNA, bacterial mRNAs are readily degraded. This crucial feature ensures the ability of bacteria to respond quickly to changing environmental conditions by adding an extra layer of control, in addition to the regulation of transcription and enzymatic activities of proteins (1). Moreover, mRNA processing allows the adjustment of different expression levels for genes encoded in one operon, as observed for the ilv operon and the glycolytic gapA operon of the Gram-positive soil bacterium Bacillus subtilis (24,40,45).The enzymes that process and degrade RNAs are called RNases. In the two model organisms Escherichia coli and B. subtilis, a set of about 20 RNases has been described thus far, but only a small fraction of these are common to both bacteria (12). Some of these RNases (e.g., RNase M5 or RNase-Mini III in B. subtilis) are necessary for the maturation of one specific target, whereas others (e.g., RNase J1 or RNase Y in B. subtilis) are more promiscuous and have a more global impact on RNA metabolism (12, 43, 58). The individual RNases differ not only in their respective amino acid sequences and in their number of substrates but also in their enzymatic mode of action. Two different classes of RNases can be defined: exoribonucleases, which remove RNA nucleotides one at a time from either the 3Ј or the 5Ј end, and endoribonucleases, which cleave within the RNA molecule. For the degradation of mRNAs, the conc...

Cyanobacterial RNA Helicase CrhR Localizes to the Thylakoid Membrane Region and Cosediments with Degradosome and Polysome Complexes in Synechocystis sp. Strain PCC 6803

“…Thus, since CrhR is encoded by the only DEAD box RNA helicase in the Synechocystis genome, given the multitasking of other bacterial RNA helicases (18)(19)(20), CrhR may be performing divergent roles and thus may associate with a range of cellular complexes under different growth conditions. CrhR association with the translation and/or RNA degradation machinery is not unexpected, as RNA helicase association with the bacterial degradosome (4)(5)(6)(12)(13)(14) and with ribosome complexes (15)(16)(17) has been well documented. How CrhR is recruited to and interacts with the thylakoid membrane and translating polysome and/or degradosome components remains unknown.…”

“…Prokaryotes do not compartmentalize routine, everyday anabolic and catabolic processes such as transcription and translation or protein and RNA decay; however, there is a growing body of evidence indicating that prokaryotes spatially confine these cellular processes (3). A prime example is the localization of the RNA degradation complex, the degradosome, to the cytoplasmic membrane (CM) in Escherichia coli (4) and Bacillus subtilis (5).…”

The Blueprint of a Minimal Cell: MiniBacillus

“…Conflicting reports are available for RNases in B. subtilis. The endoribonuclease RNase Y and the 5=-to-3= exonuclease RNase J1 have long been considered to be essential, whereas a recent report suggests that these proteins are dispensable (34,(67)(68)(69). However, the strong expression of the corresponding genes suggests that they should be part of a minimal cell.…”

Current knowledge on regulatory RNAs and their machineries inStaphylococcus aureus

“…26,110,112 The two B. subtilis enzymes RNase J1/J2 are endowed with a dual activity of an endoribonuclease and a 5'-3' exoribonuclease 113 while RNase Y, is the functional equivalent of RNase E and cleaves mRNA at unpaired U/A rich sequences. 114,115 Hence, S. aureus and B. subtilis share the same components and three of them (enolase, PNPase, DEADbox RNA helicase) are also conserved in Gram-negative bacteria. More surprisingly, the protein subunit of RNase P, RnpA, is associated with CshA in S. aureus and this interaction takes place in B. subtilis too.…”

“…114 Interestingly enough, the localization of RNase Y at the membrane is essential in vivo indicating that subcellular localization is required for the turnover of a subset of mRNAs. 115 Although the target genes of RNase Y are not yet defined in S. aureus, deletion of cvfA represses the transcription of the agr operon resulting in the repression of exotoxin genes, and in the accumulation of protein A. 117 The third protein RpnA, which is the co-factor of RNase P, is an essential protein most likely due to its role in tRNA maturation.…”