Inactivation of NMD increases viability of sup45 nonsense mutants in Saccharomyces cerevisiae

Chabelskaya, Svetlana; Gryzina, Valentina; Moskalenko, Svetlana E.; Goff, Catherine Le; Zhouravleva, Galina A.

doi:10.1186/1471-2199-8-71

Cited by 16 publications

(23 citation statements)

References 46 publications

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“…The amount of eRF1 protein was taken from Moskalenko et al (2003) although lower (11% versus 29%) is associated with a higher quantity of protein (13% versus 8%) and relative stabilization of the mRNA (0.8-fold wild-type). For the sup45-105 mutant (8% readthrough), the accumulation of protein equivalent to that of the sup45-104 mutant can be explained by the stabilization of the SUP45 mRNA (1.7-fold wildtype) as a result of NMD machinery failure in sup45 mutants (Chabelskaya et al 2007). Taken together, these results show that PTC spread through the SUP45 sequence lead to a diVerential in mRNA stability.…”

Section: Mutant Strain Background Does Not Inxuence the Level Of Readmentioning

confidence: 92%

The paradox of viable sup45 STOP mutations: a necessary equilibrium between translational readthrough, activity and stability of the protein

et al. 2009

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The mechanisms leading to non-lethality of nonsense mutations in essential genes are poorly understood. Here, we focus on the factors influencing viability of yeast cells bearing premature termination codons (PTCs) in the essential gene SUP45 encoding translation termination factor eRF1. Using a dual reporter system we compared readthrough efficiency of the natural termination codon of SUP45 gene, spontaneous sup45-n (nonsense) mutations, nonsense mutations obtained by site-directed mutagenesis (76Q --> TAA, 242R --> TGA, 317L --> TAG). The nonsense mutations in SUP45 gene were shown to be situated in moderate contexts for readthrough efficiency. We showed that readthrough efficiency of some of the mutations present in the sup45 mutants is not correlated with full-length Sup45 protein amount. This resulted from modification of both sup45 mRNA stability which varies 3-fold among sup45-n mutants and degradation rate of mutant Sup45 proteins. Our results demonstrate that some substitutions in the place of PTCs decrease Sup45 stability. The viability of sup45 nonsense mutants is therefore supported by diverse mechanisms that control the final amount of functional Sup45 in cells.

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Section: Mutant Strain Background Does Not Inxuence the Level Of Readmentioning

confidence: 92%

The paradox of viable sup45 STOP mutations: a necessary equilibrium between translational readthrough, activity and stability of the protein

et al. 2009

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“…eRF1 forms a heterodimeric complex with another protein, eRF3, a class II GTPase release factor (Stansfield et al 1995;Zhouravleva et al 1995). Although eRF1 and eRF3 are both encoded by essential genes in yeast, a number of reports describe viable premature termination codon (PTC) mutants in both SUP45 (encoding eRF1) (Stansfield et al 1996;Moskalenko et al 2003) and SUP35 (eRF3-encoding) (Chabelskaya et al 2007). All such PTC release factor mutants exhibit increased frequencies of stop codon read-through and reduced levels of full-length release factor (Stansfield et al 1996;Chabelskaya et al 2007).…”

Section: Establishing Negative Feedback Control De Novo: Nonsense Allmentioning

confidence: 99%

“…Although eRF1 and eRF3 are both encoded by essential genes in yeast, a number of reports describe viable premature termination codon (PTC) mutants in both SUP45 (encoding eRF1) (Stansfield et al 1996;Moskalenko et al 2003) and SUP35 (eRF3-encoding) (Chabelskaya et al 2007). All such PTC release factor mutants exhibit increased frequencies of stop codon read-through and reduced levels of full-length release factor (Stansfield et al 1996;Chabelskaya et al 2007). Some of the SUP45 PTC mutants were identified by selecting for an allosuppressor phenotype, an enhancement of the suppressor activity of an ordinarily weak UAA suppressor tRNA SUQ5 oc (Stansfield et al 1996).…”

Section: Establishing Negative Feedback Control De Novo: Nonsense Allmentioning

confidence: 99%

Autoregulatory systems controlling translation factor expression: Thermostat-like control of translational accuracy

Betney

Silva²,

Krishnan³

et al. 2010

RNA

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In both prokaryotes and eukaryotes, the expression of a large number of genes is controlled by negative feedback, in some cases operating at the level of translation of the mRNA transcript. Of particular interest are those cases where the proteins concerned have cell-wide function in recognizing a particular codon or RNA sequence. Examples include the bacterial translation termination release factor RF2, initiation factor IF3, and eukaryote poly(A) binding protein. The regulatory loops that control their synthesis establish a negative feedback control mechanism based upon that protein's RNA sequence recognition function in translation (for example, stop codon recognition) without compromising the accurate recognition of that codon, or sequence during general, cell-wide translation. Here, the bacterial release factor RF2 and initiation factor IF3 negative feedback loops are reviewed and compared with similar negative feedback loops that regulate the levels of the eukaryote release factor, eRF1, established artificially by mutation. The control properties of such negative feedback loops are discussed as well as their evolution. The role of negative feedback to control translation factor expression is considered in the context of a growing body of evidence that both IF3 and RF2 can play a role in stimulating stalled ribosomes to abandon translation in response to amino acid starvation. Here, we make the case that negative feedback control serves primarily to limit the overexpression of these translation factors, preventing the loss of fitness resulting from an unregulated increase in the frequency of ribosome drop-off.

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“…The extent of this destabilization is dictated by the nonsense codon position, with more 59 positions being more destabilizing (Losson and Lacroute 1979). The NMD pathway is known to affect the behavior of some eRF1 premature stop codon mutants (Chabelskaya et al 2007), and it was therefore important to take this effect into account when developing a model to describe the behavior of these mutants. The position effect of premature nonsense codons has been experimentally determined with some accuracy (Cao and Parker 2001), and these data were used to define an NMD destabilization function (Materials and Methods, Eqs.…”

Section: Model Validationmentioning

confidence: 99%

“…eRF1 nonsense mutations of this type have been isolated in the presence of specific mutant suppressor tRNAs that help support readthrough of the premature stop codon (Stansfield et al 1996), but intriguingly, also in wild-type tRNA backgrounds (Moskalenko et al 2003). The level of readthrough of the premature eRF1 nonsense codons is regulated by a complex interaction between the suppressor tRNA efficiency (Stansfield et al 1996;de Silva et al 2010), the eRF1 protein stability and activity, and the modulatory effect of the nonsense-mediated mRNA decay system that acts to destabilize mRNAs carrying early stop codons (Chabelskaya et al 2007;Kiktev et al 2009). As in the case of the RF2 feedback loop, this system appears to be autoregulatory.…”

Section: Introductionmentioning

confidence: 99%

Regulation of release factor expression using a translational negative feedback loop: A systems analysis

Betney

Silva

Mertens

et al. 2012

RNA

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The essential eukaryote release factor eRF1, encoded by the yeast SUP45 gene, recognizes stop codons during ribosomal translation. SUP45 nonsense alleles are, however, viable due to the establishment of feedback-regulated readthrough of the premature termination codon; reductions in full-length eRF1 promote tRNA-mediated stop codon readthrough, which, in turn, drives partial production of full-length eRF1. A deterministic mathematical model of this eRF1 feedback loop was developed using a staged increase in model complexity. Model predictions matched the experimental observation that strains carrying the mutant SUQ5 tRNA (a weak UAA suppressor) in combination with any of the tested sup45 UAA nonsense alleles exhibit threefold more stop codon readthrough than that of an SUQ5 yeast strain. The model also successfully predicted that eRF1 feedback control in an SUQ5 sup45 UAA mutant would resist, but not completely prevent, imposed changes in eRF1 expression. In these experiments, the introduction of a plasmid-borne SUQ5 copy into a sup45 UAA SUQ5 mutant directed additional readthrough and full-length eRF1 expression, despite feedback. Secondly, induction of additional sup45 UAA mRNA expression in a sup45 UAA SUQ5 strain also directed increased full-length eRF1 expression. The autogenous sup45 control mechanism therefore acts not to precisely control eRF1 expression, but rather as a damping mechanism that only partially resists changes in release factor expression level. The validated model predicts that the degree of feedback damping (i.e., control precision) is proportional to eRF1 affinity for the premature stop codon. The validated model represents an important tool to analyze this and other translational negative feedback loops.

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Inactivation of NMD increases viability of sup45 nonsense mutants in Saccharomyces cerevisiae

Cited by 16 publications

References 46 publications

The paradox of viable sup45 STOP mutations: a necessary equilibrium between translational readthrough, activity and stability of the protein

The paradox of viable sup45 STOP mutations: a necessary equilibrium between translational readthrough, activity and stability of the protein

Autoregulatory systems controlling translation factor expression: Thermostat-like control of translational accuracy

Regulation of release factor expression using a translational negative feedback loop: A systems analysis

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