Protein misfolding and temperature up-shift cause G(1) arrest via a commonmechanism dependent on heat shock factor in Saccharomyces cerevisiae

Accumulation of misfolded proteins in the cell at high temperature may caus e entry into a nonproliferating, heat-shocked state. The imino acid analog azetidine 2-carboxylic acid (AZC) is incorporated into cellular protein com petitively with proline and can misfold proteins into which it is incorpora ted. AZC addition to budding yeast cells at concentrations sufficient to in hibit proliferation selectively activates heat shock factor (HSF). We find that AZC treatment fails to cause accumulation of glycogen and trehalose (M sn2/4-dependent processes) or to induce thermotolerance (a protein kinase C -dependent process). However, AZC-arrested cells can accumulate glycogen an d trehalose and can acquire thermotolerance in response to a subsequent hea t shock. We find that AZC treatment arrests cells in a viable state and tha t this arrest is reversible. We find that cells at high temperature or cell s deficient in the ubiquitin-conjugating enzymes Ubc4 and Ubc5 are hypersen sitive to AZC-induced proliferation arrest. We find that AZC treatment mimi cs temperature up-shift in arresting cells in G1 and represses expression o f CLN1 and CLN2. Mutants with reduced G(1) cyclin-Cdc28 activity are hypers ensitive to AZC-induced proliferation arrest. Expression of the hyperstable Cln3-2 protein prevents G(1) arrest upon AZC treatment and temperature up- shift. Finally, we find that the EXA3-1 mutation, encoding a defective HSF, prevents efficient G(1) arrest in response to both temperature up-shift an d AZC treatment. We conclude that nontoxic levels of misfolded proteins (in duced by AZC treatment or by high temperature) selectively activate HSF. wh ich is required for subsequent G(1) arrest.