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

Abstract

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.

From past to future: suppressor mutations in yeast genes encoding translation termination factors

“…However, since one of our viable sup45-n mutants (sup45-13) contained an allele identical to sup45-18 (Stansfield et al, 1996), the inviability of the sup45-0 mutants in the absence of SUQ5 cannot be explained by the influence of the context. A mathematical model predicting the Sup45 translational readthrough negative feedback loop control was created; this model anticipates that manifestation of sup45-n mutations will depend on nucleotide context, as well as on the level of tRNA suppression and amount of SUP45 mRNA (Betney et al, 2012).…”

Expression of the translation termination factor eRF1 is autoregulated by translational readthrough and 3'UTR intron‐mediated NMD in Neurospora crassa

“…Autoregulation, which significantly stabilizes gene expression [46], would be an efficient ‘solution’ to stabilize the expression of translation termination factors. However, these factors are required for the expression of all protein coding genes [47], and thus, for efficient autoregulation, they have to contain specific sensitizing elements that make them especially sensitive to the concentration of termination factor. In prokaryotes, the RF2 translation termination factor is autoregulated as the RF2 mRNA contains a sensitizing element, an early stop codon in a frameshift context.…”

“…In these cases, the functional protein is generated by stop codon recoding (readthrough and/or frameshift) events, whose frequencies depend on the concentration of the encoded translation termination factor. eRF1 autoregulated yeast mutants were also isolated, in which the eRF1 harbored an early stop codon in readthrough facilitating contexts [23,47,50]. However, these yeast mutants growth slowly.…”

Nonsense Mutations in the Yeast SUP35 Gene Affect the [PSI+] Prion Propagation

“…However, the situation may be more complicated. Previously in the laboratory of Ian Stansfield it was shown that transcriptional up-regulation of the nonsense allele sup45-18 leads to increased expression of full-length Sup45, even in the context of the negative feedback loop [31]. Authors could not prove this hypothesis by Western blotting because their antibodies did not recognize the N-terminal part of Sup45, but their data indicate that the presence of the sup45-n allele can affect the amount of full-length Sup45.…”