Translational repression by a transcriptional elongation factor

Wilson, Helen Rose; Kameyama, Luis; Zhou, Jianguang; Guarneros, Gabriel; Court, Donald L.

doi:10.1101/gad.11.17.2204

Cited by 31 publications

(52 citation statements)

References 72 publications

(71 reference statements)

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“…Plasmid pSDF388 is a derivative of pERW12. SB1981 was crossed with each plasmid to select recombinants that had substituted the p L (N-lacZ ) fusion for the araC gene in the phage (Wilson et al, 1997). The phage was then used to make single-copy constructs of the fusions under control of the p L promoter and cI857 repressor (see Kameyama et al, 1991).…”

Section: Methodsmentioning

confidence: 99%

Genetic uncoupling of the dsRNA‐binding and RNA cleavage activities of the Escherichia coli endoribonuclease RNase III — the effect of dsRNA binding on gene expression

Dasgupta

Fernández

Kameyama

et al. 1998

Molecular Microbiology

108

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SummaryRNase III, a double-stranded RNA-specific endonuclease, is proposed to be one of Escherichia coli 's global regulators because of its ability to affect the expression of a large number of unrelated genes by influencing post-transcriptional control of mRNA stability or mRNA translational efficiency. Here, we describe the phenotypes of bacteria carrying point mutations in rnc, the gene encoding RNase III. The substrate recognition and RNA-processing properties of mutant proteins were analysed in vivo by measuring expression from known RNase III-modulated genes and in vitro from the proteins' binding and cleavage activities on known double-stranded RNA substrates. Our results show that although the point mutation rnc70 exhibited all the usual rnc null-like phenotypes, unlike other mutations, it was dominant over the wild-type allele. Multicopy expression of rnc70 could suppress a lethal phenotype of the wild-type rnc allele in a certain genetic background; it could also inhibit the RNase III-mediated activation of N gene translation by competing for the RNA-binding site of the wild-type endonuclease. The mutant protein failed to cleave the standard RNase III substrates in vitro but exhibited an affinity for double-stranded RNA when passed through poly(rI):poly(rC) columns. Filter binding and gel-shift assays with purified Rnc70 showed that the mutant protein binds to known RNase III mRNA substrates in a site-specific manner. In vitro processing reactions with purified enzyme and labelled RNA showed that the in vivo dominant effect of the mutant enzyme over the wild-type was not necessarily caused by formation of mixed dimers. Thus, the rnc70 mutation generates a mutant RNase III with impaired endonucleolytic activity but without blocking its ability to recognize and bind double-stranded RNA substrates.

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Section: Methodsmentioning

confidence: 99%

Genetic uncoupling of the dsRNA‐binding and RNA cleavage activities of the Escherichia coli endoribonuclease RNase III — the effect of dsRNA binding on gene expression

Dasgupta

Fernández

Kameyama

et al. 1998

Molecular Microbiology

108

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“…Among mutations that abrogate antitermination, the nutL mutations boxB44 and boxA5 block N-mediated translation repression, whereas nusA1, nusB5, nusE71, and boxA16 have only marginal effects on N translation (18). This finding raised the possibility that N binding to NUTL might suffice to block ribosome attachment.…”

mentioning

confidence: 65%

“…The pR promoter is activated by thermal denaturation of the cI857 repressor. Strains carrying a defective cI857 prophage in which the pL-nutL-N region is linked to lacZ by either an operon or a protein fusion are described (18), with the exception of the NUTL boxA69 mutant, which was constructed for this work by using the protocol described (18). All plasmids were constructed from pET-21d vector (Novagen).…”

Section: Methodsmentioning

confidence: 99%

“…When cleavage does not occur, either because of RNaseIII deficiency or deletion of the RNaseIII site (rIII⌬), N inhibits its own translation (17,18). It was suggested that the folding of pL RNA to form the rIII structure brings NUTL close to the beginning of N. If NUTL is bound to N and the antitermination complex, the ribosome-binding site of N is occluded.…”

Section: Inhibition Of N Translation By Nun K106͞107dmentioning

confidence: 99%

“…In addition, failure to cleave the RNaseIII site (rIII) induces a negative autoregulatory loop, in which N strongly represses its own translation (16)(17)(18). It is likely that the down-regulation of N is important for late stages in the lytic cycle as well as for entry into the lysogenic pathway (17,18).…”

mentioning

confidence: 99%

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Phage HK022 Nun protein represses translation of phage λ N (transcription termination/translation repression)

Kim¹,

Zhou²,

Wilson³

et al. 2003

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

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The N-terminal arginine-rich motif of phage HK022 Nun protein binds to NUT sequences in phage nascent transcripts and induces transcription termination. Interactions between the Nun C terminus and RNA polymerase as well as the DNA template are required for termination. We have isolated Nun C-terminal point and deletion mutants that are unable to block transcription. The mutants bind NUT RNA and inhibit translation of the N gene. Thus HK022 excludes both by terminating transcription on the phage chromosome and by preventing translation of the essential N gene. Like N autoregulation, translation repression by Nun requires host RNaseIII deficiency (rnc) or a mutation in the RNaseIII processing site (rIII) located between NUTL and the beginning of the N coding sequence. Our data support the idea that Nun bound at NUTL causes steric interference with ribosome attachment to the nearby N coding sequence. Two models, Nun acting alone or in complex with host proteins, are discussed.

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Bacteriophage λ N Protein Disorder‐Order Transitions upon Interactions with RNA or Proteins

Schweimer

Rösch

2011

Flexible Viruses

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Translational repression by a transcriptional elongation factor

Cited by 31 publications

References 72 publications

Genetic uncoupling of the dsRNA‐binding and RNA cleavage activities of the Escherichia coli endoribonuclease RNase III — the effect of dsRNA binding on gene expression

Genetic uncoupling of the dsRNA‐binding and RNA cleavage activities of the Escherichia coli endoribonuclease RNase III — the effect of dsRNA binding on gene expression

Phage HK022 Nun protein represses translation of phage λ N (transcription termination/translation repression)

Bacteriophage λ N Protein Disorder‐Order Transitions upon Interactions with RNA or Proteins

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