The
yeast transcriptional activator Gcn4p is required for high-level
expression of genes involved in the biosynthesis of amino acids and
purines. Gcn4p, an unstable protein of t 1/2 < 5 min, is
degraded via the ubiquitin pathway. Gcn4 degradation is regulated: under
conditions of starvation for amino-acids, the half-life of the protein
is greatly extended. Here, we show that this extended half-life is
correlated with a reduction in Gcn4p ubiquitination. We further show
that low levels of cycloheximide also lead to Gcn4p stabilization,
suggesting that inhibition of protein synthesis, rather than reduced
intracellular amino acid concentration, constitutes the proximal signal
for Gcn4 stabilization. Gcn4p degradation requires the ubiquitin-conjugating enzyme Cdc34p (Ubc3p). Here, we show that mutants of
CDC4 and CDC53 , as well as certain alleles of SKP1 ,
are also defective in Gcn4 degradation. The effect of these mutations on
Gcn4p parallel their effect on the CDK inhibitor Sic1p. Thus, Sic1p and
Gcn4p may be ubiquitinated by the same complex. However, degradation of
Sic1p and of another SCF CDC4 substrate, Cdc6p, are unaffected
by amino-acid starvation. We discuss possible mechanisms for the
regulation of Gcn4 degradation.
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