Activation of RegB endoribonuclease by S1 ribosomal protein requires an 11 nt conserved sequence

Durand, Sylvain; Richard, Graziella; Bisaglia, Marco; Laalami, Soumaya; Bontems, François; Uzan, Marc

doi:10.1093/nar/gkl911

Cited by 15 publications

(17 citation statements)

References 50 publications

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“…Finally, in the case of RegB activation, an 11-nucleotide sequence extending downstream the GGAG was found to be critical for S1 activity. A consensus sequence was defined (GGAGAAUAAAA), where the adenines in positions 6, 8, and 11 seem to be crucial (27).…”

Section: Discussionmentioning

confidence: 99%

“…We analyzed the interactions between the different domains in the F3-5 fragment by NMR and small angle x-ray scattering, showing that the domains D4 and D5 are in contact with each other, whereas domains D3 and D4 appear to be in equilibrium between a noninteracting and a weakly interacting state. Finally, we analyzed the interactions of the F3-5 fragment with three different biologically pertinent RNAs, F3-5 having a direct activity on two of them (26,27). Our results indicate that the three domains are involved in RNA binding, through systematic (common to the three RNAs) and specific (RNA-dependent) interactions.…”

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confidence: 91%

“…These were also selected as the best S1 ligands in a SELEX experiment (25). We observed that S1 promotes the cleavage by RegB of partially structured AG-rich RNAs (26) and of unstructured RNA characterized by an 11-nucleotide A-rich consensus sequence (27).…”

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confidence: 99%

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S1 Ribosomal Protein Functions in Translation Initiation and Ribonuclease RegB Activation Are Mediated by Similar RNA-Protein Interactions

Aliprandi

Sizun

Pérez

et al. 2008

Journal of Biological Chemistry

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The ribosomal protein S1, in Escherichia coli, is necessary for the recognition by the ribosome of the translation initiation codon of most messenger RNAs. It also participates in other functions. In particular, it stimulates the T4 endoribonuclease RegB, which inactivates some of the phage mRNAs, when their translation is no longer required, by cleaving them in the middle of their Shine-Dalgarno sequence. In each function, S1 seems to target very different RNAs, which led to the hypothesis that it possesses different RNA-binding sites. We previously demonstrated that the ability of S1 to activate RegB is carried by a fragment of the protein formed of three consecutive domains (domains D3, D4, and D5). The same fragment plays a central role in all other functions. We analyzed its structural organization and its interactions with three RNAs: two RegB substrates and a translation initiation region. We show that these three RNAs bind the same area of the protein through a set of systematic (common to the three RNAs) and specific (RNA-dependent) interactions. We also show that, in the absence of RNA, the D4 and D5 domains are associated, whereas the D3 and D4 domains are in equilibrium between open (noninteracting) and closed (weakly interacting) forms and that RNA binding induces a structural reorganization of the fragment. All of these results suggest that the ability of S1 to recognize different RNAs results from a high adaptability of both its structure and its binding surface.

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Section: Discussionmentioning

confidence: 99%

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confidence: 91%

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S1 Ribosomal Protein Functions in Translation Initiation and Ribonuclease RegB Activation Are Mediated by Similar RNA-Protein Interactions

Aliprandi

Sizun

Pérez

et al. 2008

Journal of Biological Chemistry

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“…Methods for RNA extractions, primer extension, and RNA sequencing are described in ref. [8][9][10]. Routinely, 700 μL infected cells were added to 77 μL SDS (10% wt/vol) and Na-EDTA (20 mM) and 500 μL water-saturated phenol maintained at 70°C.…”

Section: Methodsmentioning

confidence: 99%

“…It targets mostly but not uniquely Shine-Dalgarno sequences of early genes (9). The direct consequence of cleavages in Shine-Dalgarno sequences is the functional inactivation of mRNAs (10).…”

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Bacteriophage T4 polynucleotide kinase triggers degradation of mRNAs

Durand

Richard

Bontems

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The bacteriophage T4-encoded RegB endoribonuclease is produced during the early stage of phage development and targets mostly (but not exclusively) the Shine–Dalgarno sequences of early genes. In this work, we show that the degradation of RegB-cleaved mRNAs depends on a functional T4 polynucleotide kinase/phosphatase (PNK). The 5′-OH produced by RegB cleavage is phosphorylated by the kinase activity of PNK. This modification allows host RNases G and E, with activity that is strongly stimulated by 5′-monophosphate termini, to attack mRNAs from the 5′-end, causing their destabilization. The PNK-dependent pathway of degradation becomes effective 5 min postinfection, consistent with our finding that several minutes are required for PNK to accumulate after infection. Our work emphasizes the importance of the nature of the 5′ terminus for mRNA stability and depicts a pathway of mRNA degradation with 5′- to 3′-polarity in cells devoid of 5′–3′ exonucleases. It also ascribes a role for T4 PNK during normal phage development.

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Overview on the Expression of Toxic Gene Products in Escherichia coli

Saïda

2007

CP Protein Science

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Despite the development of various nonbacterial expression systems, Escherichia coli (E. coli) remains the host of choice for recombinant protein expression. Its culture is simple, fast, inexpensive, and highly efficient (tens of milligrams of pure proteins are typically obtained within 48 hours using as little as 1 liter of culture). Unfortunately, many toxic genes (from various organisms) severely interfere with the physiology of E. coli. As a result, expression yields are dramatically diminished, and sometimes abolished. In fact, some genes are so toxic that E. coli cannot maintain their expression vector during the growth phase (the phase during which recombinant gene expression is presumably repressed). Therefore, modified expression vectors, modified E. coli strains, and appropriate cultivation protocols are needed. This overview discusses several special strategies successfully used to express toxic genes in E. coli.

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Activation of RegB endoribonuclease by S1 ribosomal protein requires an 11 nt conserved sequence

Cited by 15 publications

References 50 publications

S1 Ribosomal Protein Functions in Translation Initiation and Ribonuclease RegB Activation Are Mediated by Similar RNA-Protein Interactions

S1 Ribosomal Protein Functions in Translation Initiation and Ribonuclease RegB Activation Are Mediated by Similar RNA-Protein Interactions

Bacteriophage T4 polynucleotide kinase triggers degradation of mRNAs

Overview on the Expression of Toxic Gene Products in Escherichia coli

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