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Polysome Fractionation and Analysis of Mammalian Translatomes on a Genome-wide Scale

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

Russell Betney¹, Eric de Silva, Christina Mertens

¹School of Medical Sciences, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, United Kingdom.

RNA (New York, N.Y.)

|October 30, 2012

Summary

Yeast release factor eRF1 (encoded by SUP45) has a feedback loop regulating stop codon readthrough. Mathematical modeling confirmed this loop dampens, but doesn't prevent, changes in eRF1 expression levels.

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Area of Science:

Molecular Biology
Genetics
Systems Biology

Background:

The essential eukaryote release factor eRF1, encoded by yeast SUP45, recognizes stop codons during translation.
SUP45 nonsense alleles are viable due to feedback-regulated readthrough of premature termination codons, where reduced eRF1 promotes readthrough, driving partial eRF1 production.

Purpose of the Study:

To develop and validate a deterministic mathematical model of the eRF1 feedback loop.
To analyze the control precision and damping mechanisms of this translational negative feedback loop.

Main Methods:

Development of a deterministic mathematical model of the eRF1 feedback loop with increasing complexity.
Experimental validation using yeast strains with SUP45 nonsense alleles and SUQ5 tRNA, including plasmid-borne SUQ5 and induced sup45 mRNA expression.

Main Results:

Model predictions accurately matched experimental observations regarding stop codon readthrough levels in different yeast strains.
The model predicted and experiments confirmed that eRF1 feedback control resists, but does not completely prevent, imposed changes in eRF1 expression.
The autogenous sup45 control mechanism acts as a damping mechanism, only partially resisting changes in release factor expression.

Conclusions:

The validated mathematical model is a valuable tool for analyzing translational negative feedback loops.
The degree of feedback damping is proportional to eRF1 affinity for premature stop codons.
The SUP45 feedback loop demonstrates a damping mechanism rather than precise control of eRF1 expression.