Consequences of RNase E scarcity in <i>Escherichia coli</i>

Jain, Chaitanya; Deana, Atilio; Belasco, Joel G.

doi:10.1046/j.1365-2958.2002.02808.x

Cited by 63 publications

(71 citation statements)

References 32 publications

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“…KSL2000 cells conditionally depleted for Rne by transferring bacteria to liquid media lacking arabinose underwent two to three cell divisions at a doubling rate similar to that observed for bacteria induced by 0.2% arabinose to express Rne at endogenous levels (data not shown). This result is consistent with the previous finding showing that E. coli cell division requires a cellular RNase E concentration at least 10-20% of normal ( Jain et al 2002).…”

Section: Resultssupporting

confidence: 94%

Identification of Amino Acid Residues in the Catalytic Domain of RNase E Essential for Survival of Escherichia coli: Functional Analysis of DNase I Subdomain

Shin

Yeom

et al. 2008

Genetics

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RNase E is an essential Escherichia coli endoribonuclease that plays a major role in the decay and processing of a large fraction of RNAs in the cell. To better understand the molecular mechanisms of RNase E action, we performed a genetic screen for amino acid substitutions in the catalytic domain of the protein (N-Rne) that knock down the ability of RNase E to support survival of E. coli. Comparative phylogenetic analysis of RNase E homologs shows that wild-type residues at these mutated positions are nearly invariably conserved. Cells conditionally expressing these N-Rne mutants in the absence of wild-type RNase E show a decrease in copy number of plasmids regulated by the RNase E substrate RNA I, and accumulation of 5S ribosomal RNA, M1 RNA, and tRNA Asn precursors, as has been found in Rne-depleted cells, suggesting that the inability of these mutants to support cellular growth results from loss of ribonucleolytic activity. Purified mutant proteins containing an amino acid substitution in the DNase I subdomain, which is spatially distant from the catalytic site posited from crystallographic studies, showed defective binding to an RNase E substrate, p23 RNA, but still retained RNA cleavage activity-implicating a previously unidentified structural motif in the DNase I subdomain in the binding of RNase E to targeted RNA molecules, demonstrating the role of the DNase I domain in RNase E activity.

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Section: Resultssupporting

confidence: 94%

Identification of Amino Acid Residues in the Catalytic Domain of RNase E Essential for Survival of Escherichia coli: Functional Analysis of DNase I Subdomain

Shin

Yeom

et al. 2008

Genetics

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“…[5][6][7][8][9][10][11]. The multicomponent complex consists of: the RNA endonuclease RNase E, whose activity is essential for Escherichia coli cell growth (12)(13)(14), RNA processing (15,16), and degradation (17,18); the 3Ј-5Ј exoribonuclease PNPase (19); RhlB RNA helicase (20); and enolase (21), an enzyme involved in the glycolytic pathway and other chaperonin proteins (3,22). Interestingly, in addition to mRNAs, highly structured, stable RNA fragments have also been found to be associated with RNA degradosome complexes (3,23), which implies quality control by the RNA degradosome for the biogenesis of stable RNAs.…”

mentioning

confidence: 99%

DEAD Box RhlB RNA Helicase Physically Associates with Exoribonuclease PNPase to Degrade Double-stranded RNA Independent of the Degradosome-assembling Region of RNase E

Liou

Chang

Lin

et al. 2002

Journal of Biological Chemistry

108

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The Escherichia coli RNA degradosome is a multicomponent ribonucleolytic complex consisting of three major proteins that assemble on a scaffold provided by the C-terminal region of the endonuclease, RNase E. Using an E. coli two-hybrid system, together with BIAcore apparatus, we investigated the ability of three proteins, polynucleotide phosphorylase (PNPase), RhlB RNA helicase, and enolase, a glycolytic protein, to interact physically and functionally independently of RNase E. Here we report that Rh1B can physically bind to PNPase, both in vitro and in vivo, and can also form homodimers with itself. However, binding of RhlB or PNPase to enolase was not detected under the same conditions. BIAcore analysis revealed real-time, direct binding for bimolecular interactions between Rh1B units and for the RhlB interaction with PNPase. Furthermore, in the absence of RNase E, purified RhlB can carry out ATP-dependent unwinding of double-stranded RNA and consequently modulate degradation of double-stranded RNA together with the exonuclease activity of PNPase. These results provide evidence for the first time that both functional and physical interactions of individual degradosome protein components can occur in the absence of RNase E and raise the prospect that the RNase E-independent complexes of RhlB RNA helicase and PNPase, detected in vivo, may constitute mini-machines that assist in the degradation of duplex RNA in structures physically distinct from multicomponent RNA degradosomes.RNA metabolism is a complex process affecting the control of gene expression. In bacteria, a multicomponent ribonucleolytic complex termed the RNA degradosome (1-4) has been identified as playing an important role in the control of mRNA degradation (for recent reviews, see Refs. 5-11). The multicomponent complex consists of: the RNA endonuclease RNase E, whose activity is essential for Escherichia coli cell growth (12-14), RNA processing (15, 16), and degradation (17, 18); the 3Ј-5Ј exoribonuclease PNPase (19); RhlB RNA helicase (20); and enolase (21), an enzyme involved in the glycolytic pathway and other chaperonin proteins (3,22). Interestingly, in addition to mRNAs, highly structured, stable RNA fragments have also been found to be associated with RNA degradosome complexes (3, 23), which implies quality control by the RNA degradosome for the biogenesis of stable RNAs. Degradosome complexity and its cooperation with individual protein components acting on degradation-targeted RNA, in vivo, remains to be discovered.Various approaches revealing protein-protein interactions in the degradosome indicate that the C-terminal region of RNase E serves as a scaffold that directly binds PNPase, RhlB RNA helicase, and enolase (24,25). No other interactions among these component proteins have been detected (25) or reported. Recently, a mini-degradosome complex (26) containing the scaffold region (without the N-terminal enzymatic region of RNase E), RhlB RNA helicase, and PNPase was reconstituted in vitro. These experiments revealed a functional inte...

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“…rRNAs in E. coli and many other proteobacteria (Ono and Kuwano 1979;Jain et al 2002;Carpousis 2007;Tsai et al 2012). Cleavage of mRNA by RNase E can occur via two pathways: 59-end monophosphate-dependent and 59-end independent (so-called "direct entry") (Anupama et al 2011;Bouvier and Carpousis 2011;Garrey and Mackie 2011).…”

Section: Discussionmentioning

confidence: 99%

Rapid Degradation of Host mRNAs by Stimulation of RNase E Activity by Srd of Bacteriophage T4

Alawneh

Yonesaki

et al. 2015

Genetics

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Escherichia coli messenger RNAs (mRNAs) are rapidly degraded immediately after bacteriophage T4 infection, and the host RNase E contributes to this process. Here, we found that a previously uncharacterized factor of T4 phage, Srd (Similarity with rpoD), was involved in T4-induced host mRNA degradation. The rapid decay of ompA and lpp mRNAs was partially alleviated and a decay intermediate of lpp mRNA rapidly accumulated in cells infected with T4 phage lacking srd. Exogenous expression of Srd in uninfected cells significantly accelerated the decay of these mRNAs. In addition, lpp(T) RNA, with a sequence identical to the decay intermediate of lpp mRNA and a triphosphate at 59-end, was also destabilized by Srd. The destabilization of these RNAs by Srd was not observed in RNase E-defective cells. The initial cleavage of a primary transcript by RNase E can be either direct or dependent on the 59-end of transcript. In the latter case, host RppH is required to convert the triphosphate at 59-end to a monophosphate. lpp(T) RNA, but not lpp and ompA mRNAs, required RppH for Srd-stimulated degradation, indicating that Srd stimulates both 59-end-dependent and -independent cleavage activities of RNase E. Furthermore, pull-down and immunoprecipitation analyses strongly suggested that Srd physically associates with the N-terminal half of RNase E containing the catalytic moiety and the membrane target sequence. Finally, the growth of T4 phage was significantly decreased by the disruption of srd. These results strongly suggest that the stimulation of RNase E activity by T4 Srd is required for efficient phage growth. KEYWORDS Srd; RNase E; Escherichia coli; bacteriophage T4; mRNA decay B ACTERIOPHAGE T4 shuts off gene expression of the host, Escherichia coli, immediately after infection and quickly starts to express its own genes (Kutter et al. 1994). Multiple mechanisms, such as modifications of apparatuses for transcription and translation, are involved in this shift of gene expression from E. coli to T4 (Carlson et al. 1994). Our previous work revealed that the representative stable E. coli messenger RNAs (mRNAs), lpp and ompA, were rapidly degraded after T4 infection (T4-induced host mRNA degra dation) and that RNase E, which is an essential endoribonuclease in E. coli (Marcaida et al. 2006), primarily functions in T4-induced host mRNA degradation (Ueno and Yonesaki 2004). This rapid degradation of host mRNAs may contribute to the quick shift from E. coli to T4 metabolism because it leads to immediate cessation of host gene expression and consequently generation of ribonucleotides and free ribosomes, each of which would stimulate transcription and translation of T4 genes. In fact, deficiency of RNase E resulted in a slow start of T4 gene transcription, reducing the level of transcription (Otsuka and Yonesaki 2005) and retarding the growth of T4 (Mudd et al. 1990a). In eukaryotic cells, the degradation of host mRNAs after viral infection, such as in alphaherpesvirus, gammaherpesvirus, or betacoronavirus, also contribu...

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Consequences of RNase E scarcity in Escherichia coli

Cited by 63 publications

References 32 publications

Identification of Amino Acid Residues in the Catalytic Domain of RNase E Essential for Survival of Escherichia coli: Functional Analysis of DNase I Subdomain

Identification of Amino Acid Residues in the Catalytic Domain of RNase E Essential for Survival of Escherichia coli: Functional Analysis of DNase I Subdomain

DEAD Box RhlB RNA Helicase Physically Associates with Exoribonuclease PNPase to Degrade Double-stranded RNA Independent of the Degradosome-assembling Region of RNase E

Rapid Degradation of Host mRNAs by Stimulation of RNase E Activity by Srd of Bacteriophage T4

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