Analysis of the Escherichia coli RNA degradosome composition by a proteomic approach

Regonesi, Maria Elena; Favero, Marta Del; Basilico, Fabrizio; Briani, Federica; Benazzi, Louise; Tortora, Paolo; Mauri, Pierluigi; Dehò, Gianni

doi:10.1016/j.biochi.2005.07.012

Cited by 71 publications

(48 citation statements)

References 42 publications

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“…Ribosomal protein S1 (GI 1,787,140, calculated MW of 61 kDa; apparent MW in SDS-PAGE of z68 kDa; Kimura et al 1982) turned out to be the most represented protein in the band, with 19 different peptides identified by amino acid sequencing and a SEQUEST score, a parameter correlated with protein abundance (Regonesi et al 2006), of 290. Two additional proteins, FtsH and DnaK, were also detected with lower score values (40 and 20, respectively).…”

Section: Identification Of a Protein Binding The Pnp Mrna Leader Regionmentioning

confidence: 99%

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Polynucleotide phosphorylase hinders mRNA degradation upon ribosomal protein S1 overexpression in Escherichia coli

Briani¹,

Curti²,

Rossi³

et al. 2008

RNA

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The exoribonuclease polynucleotide phosphorylase (PNPase, encoded by pnp) is a major player in bacterial RNA decay. In Escherichia coli, PNPase expression is post-transcriptionally regulated at the level of mRNA stability. The primary transcript is very efficiently processed by the endonuclease RNase III at a specific site and the processed pnp mRNA is rapidly degraded in a PNPase-dependent manner. While investigating the PNPase autoregulation mechanism we found, by UV-cross-linking experiments, that the ribosomal protein S1 in crude extracts binds to the pnp-mRNA leader region. We assayed the potential role of S1 protein in pnp gene regulation by modulating S1 expression from depletion to overexpression. We found that S1 depletion led to a sharp decrease of the amount of pnp and other tested mRNAs, as detected by Northern blotting, whereas S1 overexpression caused a strong stabilization of pnp and the other transcripts. Surprisingly, mRNA stabilization depended on PNPase, as it was not observed in a pnp deletion strain. PNPase-dependent stabilization, however, was not detected by chemical decay assay of bulk mRNA. Overall, our data suggest that PNPase exonucleolytic activity may be modulated by the translation potential of the target mRNAs and that, upon ribosomal protein S1 overexpression, PNPase protects from degradation a set of full-length mRNAs. It thus appears that a single mRNA species may be differentially targeted to either decay or PNPase-dependent stabilization, thus preventing its depletion in conditions of fast turnover.

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Section: Identification Of a Protein Binding The Pnp Mrna Leader Regionmentioning

confidence: 99%

“…As a loading control, the gels have been hybridized also with a 5S rRNA-specific radiolabeled oligonucleotide. and Bertani 1965; Piazza et al 1996), and C-5691 (C-1a Dpnp-751) (Regonesi et al 2006) were described previously. Dpnp-751 is an in frame deletion encompassing z95% of pnp coding sequence.…”

Section: Bacterial Strains and Plasmidsmentioning

confidence: 99%

Polynucleotide phosphorylase hinders mRNA degradation upon ribosomal protein S1 overexpression in Escherichia coli

Briani¹,

Curti²,

Rossi³

et al. 2008

RNA

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“…RNase E-Flag degradosomes were prepared as described previously (Regonesi et al 2006), with some modifications. E. coli.…”

Section: Purification Of Rnase E-flag Degradosomesmentioning

confidence: 99%

Small RNA-induced mRNA degradation achieved through both translation block and activated cleavage

Prévost¹,

Desnoyers²,

Jacques³

et al. 2011

Genes Dev.

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Small RNA (sRNA)-induced mRNA degradation occurs through binding of an sRNA to a target mRNA with the concomitant action of the RNA degradosome, which induces an endoribonuclease E (RNase E)-dependent cleavage and degradation of the targeted mRNA. Because many sRNAs bind at the ribosome-binding site (RBS), it is possible that the resulting translation block is sufficient to promote the rapid degradation of the targeted mRNA. Contrary to this mechanism, we report here that the pairing of the sRNA RyhB to the target mRNA sodB initiates mRNA degradation even in the absence of translation on the mRNA target. Remarkably, even though it pairs at the RBS, the sRNA RyhB induces mRNA cleavage in vivo at a distal site located >350 nucleotides (nt) downstream from the RBS, ruling out local cleavage near the pairing site. Both the RNA chaperone Hfq and the RNA degradosome are required for efficient cleavage at the distal site. Thus, beyond translation initiation block, sRNAinduced mRNA cleavage requires several unexpected steps, many of which are determined by structural features of the target mRNA.

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“…The formation of the complex with Hfq and SgrS requires the same region of RNase E that is necessary for the formation of the RNA degradosome, and evidence suggests that the degradosome is remodeled because of the new interaction. RraA and RraB, protein factors that downregulate RNase E activity, are also believed to remodel the RNA degradosome, and there is evidence that RNase E can form a "cold shock" RNA degradosome in which CsdA, another DEAD-box RNA helicase, is recruited to the complex (18,27,31,45,48). The noncatalytic region of RNase E can therefore be viewed as an interaction hub involved in a variety of proteinprotein interactions.…”

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confidence: 99%

Characterization of the RNA Degradosome ofPseudoalteromonas haloplanktis: Conservation of the RNase E-RhlB Interaction in the Gammaproteobacteria

Ait-Bara

Carpousis

2010

J Bacteriol

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The degradosome is a multienzyme complex involved in mRNA degradation in Escherichia coli. The essential endoribonuclease RNase E contains a large noncatalytic region necessary for protein-protein interactions with other components of the RNA degradosome. Interacting proteins include the DEAD-box RNA helicase RhlB, the glycolytic enzyme enolase, and the exoribonuclease PNPase. Pseudoalteromonas haloplanktis, a psychrotolerant gammaproteobacterium distantly related to E. coli, encodes homologs of each component of the RNA degradosome. In P. haloplanktis, RNase E associates with RhlB and PNPase but not enolase. Plasmids expressing P. haloplanktis RNase E (Ph-RNase E) can complement E. coli strains lacking E. coli RNase E (Ec-RNase E). Ph-RNase E, however, does not confer a growth advantage to E. coli at low temperature. Ph-RNase E has a heterologous protein-protein interaction with Ec-RhlB but not with Ec-enolase or EcPNPase. The Ph-RNase E binding sites for RhlB and PNPase were mapped by deletion analysis. The PNPase binding site is located at the C-terminal end of Ph-RNase E at the same position as that in Ec-RNase E, but the sequence of the site is not conserved. The sequence of the RhlB binding site in Ph-RNase E is related to the sequence in Ec-RNase E. Together with the heterologous interaction between Ph-RNase E and Ec-RhlB, our results suggest that the underlying structural motif for the RNase E-RhlB interaction is conserved. Since the activity of Ec-RhlB requires its physical interaction with Ec-RNase E, conservation of the underlying structural motif over a large evolutionary distance could be due to constraints involved in the control of RhlB activity.

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Analysis of the Escherichia coli RNA degradosome composition by a proteomic approach

Cited by 71 publications

References 42 publications

Polynucleotide phosphorylase hinders mRNA degradation upon ribosomal protein S1 overexpression in Escherichia coli

Polynucleotide phosphorylase hinders mRNA degradation upon ribosomal protein S1 overexpression in Escherichia coli

Small RNA-induced mRNA degradation achieved through both translation block and activated cleavage

Characterization of the RNA Degradosome ofPseudoalteromonas haloplanktis: Conservation of the RNase E-RhlB Interaction in the Gammaproteobacteria

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