The Response Regulator SprE (RssB) Modulates Polyadenylation and mRNA Stability in<i>Escherichia coli</i>

Carabetta, Valerie J.; Mohanty, Bijoy K.; Kushner, Sidney R.; Silhavy, Thomas J.

doi:10.1128/jb.00870-09

Cited by 19 publications

(24 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Thus, it has been proposed that RNA degradation may be a compartmentalized phenomenon. PAP I also localizes to the inner membrane during exponential growth (7,21). A common subcellular localization of both PAP I and the degradosome supports the idea that these two processes are coordinated.…”

supporting

confidence: 57%

“…Bacteria were grown at 37°C with aeration, and growth was monitored by measuring the optical density at 600 nm (OD 600 ). The ppcnB-GFP plasmid has been described previously (7). This plasmid expresses the PAP I-green fluorescent protein (GFP) fusion protein under the control of the native pcnB promoters.…”

Section: Methodsmentioning

confidence: 99%

“…During exponential growth, SprE plays a role in the normal maintenance of the poly(A) levels and, therefore, the stability of mRNAs. In exponential phase, PAP I is membrane localized; however, upon entry into stationary phase, PAP I is localized in a few punctate spots in the cytoplasm (7). Polyadenylation in stationary phase is poorly understood, but SprE appears to be required for the change in the intracellular localization of PAP I.…”

mentioning

confidence: 99%

See 2 more Smart Citations

The Response Regulator SprE (RssB) Is Required for Maintaining Poly(A) Polymerase I-Degradosome Association during Stationary Phase

2010

Self Cite

View full text Add to dashboard Cite

Poly(A) polymerase I (PAP I) is the enzyme responsible for the addition of poly(A) tails onto RNA molecules in Escherichia coli. Polyadenylation is believed to facilitate the destruction of such RNAs by the mRNA degradosome. Recently, it was discovered that the stationary-phase regulatory protein SprE (RssB) has a second function in the control of polyadenylation that is distinct from its known function in the regulated proteolysis of RpoS. In the work presented herein, we used a targeted proteomic approach to further investigate SprE's involvement in the polyadenylation pathway. Specifically, we used cryogenic cell lysis, immunopurifications on magnetic beads, and mass spectrometry to identify interacting partners of PAP I-green fluorescent protein. We provide the first in vivo evidence that PAP I interacts with the mRNA degradosome during both exponential and stationary phases and find that the degradosome can contain up to 10 different proteins under certain conditions. Moreover, we demonstrate that the majority of these PAP I interactions are formed via protein-protein interactions and that SprE plays an important role in the maintenance of the PAP I-degradosome association during stationary phase.Polyadenylation in Escherichia coli is important for proper RNA metabolism and, therefore, proper gene expression. Poly(A) polymerase I (PAP I) catalyzes the addition of poly(A) tails onto the 3Ј ends of RNA molecules, which ultimately facilitates their destruction (25,32,39). PAP I requires the RNA chaperone Hfq for efficient recognition of RNA substrates that contain 3Ј hairpin structures but not for unstructured RNA products (35). It was recently shown that ϳ90% of transcripts are polyadenylated in exponentially growing cells, demonstrating that polyadenylation is a frequent and fundamental process (33). A second enzyme involved in the polyadenylation of RNA species is polynucleotide phosphorylase (PNPase) (34). PNPase is a reversible enzyme, as it has a 3Ј 3 5Ј exoribonuclease activity and a 5Ј 3 3Ј templateindependent poly(A) polymerase activity (19,34,47

show abstract

supporting

confidence: 57%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

The Response Regulator SprE (RssB) Is Required for Maintaining Poly(A) Polymerase I-Degradosome Association during Stationary Phase

2010

Self Cite

View full text Add to dashboard Cite

show abstract

“…Accordingly, we conclude that phosphorylation of SprE is not integral to the carbon starvation signaling pathway that regulates RpoS. SprE was recently found to regulate mRNA stability (Carabetta et al 2009), and it is not known if phosphorylation plays a role in that activity.In response to other stresses, such as phosphate starvation, magnesium starvation, and DNA damage, a major mechanism for RpoS accumulation is antagonism of the adaptor SprE by the anti-adaptor proteins IraP, IraM, and IraD, respectively (Bougdour et al 2006(Bougdour et al , 2008Merrikh et al 2009). However, none of the Ira proteins have been associated with carbon starvation, and despite extensive screens, no other proteins have been identified in the carbon starvation signaling pathway.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

RpoS proteolysis is controlled directly by ATP levels in Escherichia coli

Peterson¹,

Levchenko²,

Rabinowitz³

et al. 2012

Genes Dev.

Self Cite

View full text Add to dashboard Cite

The master regulator of stationary phase in Escherichia coli, RpoS, responds to carbon availability through changes in stability, but the individual steps in the pathway are unknown. Here we systematically block key steps of glycolysis and the citric acid cycle and monitor the effect on RpoS degradation in vivo. Nutrient upshifts trigger RpoS degradation independently of protein synthesis by activating metabolic pathways that generate small energy molecules. Using metabolic mutants and inhibitors, we show that ATP, but not GTP or NADH, is necessary for RpoS degradation. In vitro reconstitution assays directly demonstrate that ClpXP fails to degrade RpoS, but not other proteins, at low ATP hydrolysis rates. These data suggest that cellular ATP levels directly control RpoS stability.Supplemental material is available for this article.Received November 17, 2011; revised version accepted February 9, 2012. RpoS, also known as s S or s 38 , is an alternate s factor that serves as the master regulator of stationary phase and the general stress response in many proteobacteria (Loewen and Hengge-Aronis 1994;Dong and Schellhorn 2009). In Escherichia coli, it has been reported to control transcription of up to 10% of the genome (Weber et al. 2005) and is necessary for survival in numerous environments (McCann et al. 1991). In healthy growing cells, RpoS is still produced but is immediately directed by the adaptor protein SprE (RssB) to the AAA + ATPase ClpXP protease for unfolding and degradation (Muffler et al. 1996;Pratt and Silhavy 1996;Baker and Sauer 2012). When carbon sources are depleted, RpoS proteolysis ceases, causing it to rapidly accumulate and compete with other s factors for binding to the core RNA polymerase. This s exchange in turn leads to changes in gene expression that protect against stresses and prepare the cell for long-term survival. Conversely, when carbon sources are replenished, RpoS is actively degraded, and RpoS-controlled genes are no longer expressed. We wish to understand the molecular signals that control RpoS stability.There are several plausible mechanisms of RpoS regulation. SprE is an adaptor specific for RpoS degradation and belongs to the response regulator family of twocomponent signaling proteins. It has a highly conserved N-terminal receiver domain with a conserved aspartic acid, D58, so it was possible that its activity was controlled by phosphorylation of this site. However, it is an orphan response regulator; there is no cognate sensor kinase. We and others have mutated the phosphorylation site of the chromosomal allele of sprE and found that RpoS proteolysis was still regulated upon carbon starvation in a SprE-dependent manner (Peterson et al. 2004;Zhou and Gottesman 2006). Accordingly, we conclude that phosphorylation of SprE is not integral to the carbon starvation signaling pathway that regulates RpoS. SprE was recently found to regulate mRNA stability (Carabetta et al. 2009), and it is not known if phosphorylation plays a role in that activity.In response to other stress...

show abstract

Bacterial/archaeal/organellar polyadenylation

Mohanty

Kushner

2010

WIREs RNA

Self Cite

View full text Add to dashboard Cite

Although the first poly(A) polymerase (PAP) was discovered in Escherichia coli in 1962, the study of polyadenylation in bacteria was largely ignored for the next 30 years. However, with the identification of the structural gene for E. coli PAP I in 1992, it became possible to analyze polyadenylation using both biochemical and genetic approaches. Subsequently, it has been shown that polyadenylation plays a multifunctional role in prokaryotic RNA metabolism. While the bulk of our current understanding of prokaryotic polyadenylation comes from studies on E. coli, recent experiments with Cyanobacteria, organelles and Archaea, although limited, have widened our view on the diversity, complexity, and universality of the polyadenylation process.For example, the identification of polynucleotide phosphorylase (PNPase), a reversible phosphorolytic enzyme that is highly conserved in bacteria, as an additional PAP in E. coli caught everyone by surprise. In fact, PNPase has now been shown to be the source of post-transcriptional RNA modifications in a wide range of cells of prokaryotic origin including those that lack a eubacterial PAP homologue. Accordingly, the past few years have witnessed increased interest in the mechanism and role of post-transcriptional modifications in all species of prokaryotic origin. However, the fact that many of the poly(A) tails are very short and unstable as well as the presence of polynucleotide tails has posed significant technical challenges to the scientific community trying to unravel the mystery of polyadenylation in prokaryotes. This review discusses the current state of knowledge regarding polyadenylation and its functions in bacteria, organelles and Archaea. Keywords poly(A) polymerase; polynucleotide phosphorylase; Hfq; RNA degradation Polyadenylation is a post-transcriptional event that involves the addition of untemplated adenosine residues to the 3' ends of RNA substrates. The first bacterial poly(A) polymerase (PAP) was identified almost 50 years ago in Escherichia coli [1,2]. A PAP was also identified in eukaryotic cells at about the same time [3,4]. However, polyadenylation in bacteria was virtually ignored for next 30 years, in part because eukaryotic poly(A) tails were relatively long, nearly uniform in length, and were found on almost all mRNAs. Furthermore, even though poly(A) tails were detected in E. coli and Bacillus subtilis [5][6][7][8][9], the overall low abundance of polyadenylated transcripts and the apparent lack of evidence for a physiological role led to the belief that polyadenylation was only important in higher organisms (See definition of polyadenylation in the glossary of Lewin, Genes I through Genes VIII).* To whom correspondence should be addressed. NIH Public Access Author ManuscriptWiley Interdiscip Rev RNA. Author manuscript; available in PMC 2011 September 27. Published in final edited form as:Wiley Interdiscip Rev RNA. 2010 September 27; 2(2): 256-276. doi:10.1002/wrna.51. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscri...

show abstract

The Response Regulator SprE (RssB) Modulates Polyadenylation and mRNA Stability inEscherichia coli

Cited by 19 publications

References 51 publications

The Response Regulator SprE (RssB) Is Required for Maintaining Poly(A) Polymerase I-Degradosome Association during Stationary Phase

The Response Regulator SprE (RssB) Is Required for Maintaining Poly(A) Polymerase I-Degradosome Association during Stationary Phase

RpoS proteolysis is controlled directly by ATP levels in Escherichia coli

Bacterial/archaeal/organellar polyadenylation

Contact Info

Product

Resources

About