Structure and expression of the yeast heat-shock gene HSP26.

A search was made for genes in Saccharomyces cerevisiae which are controlled by tightly regulated promoters that direct growth phase-specific gene expression during growth in batch culture. HSP26, a gene which encodes the major yeast small heat-shock protein, Hsp26, was found to be transcriptionally repressed during normal exponential growth (log-phase) and strongly induced during entry into stationary-phase on a glucose medium. The close parallel between the steady-state levels of HSP26 mRNA and the synthesis of the Hsp26 polypeptide, indicated that the synthesis of Hsp26 during the fermentative batch growth cycle is regulated primarily at the level of gene transcription. Activation of HSP26 during stationary-phase does not, however, result from the release of carbon catabolite repression due to glucose exhaustion, nor does it require the function of the WHI2 gene, involved in the normal coordination between cell proliferation and nutrient availability. The possible mechanism by which HSP2 6 is regulated during the yeast growth cycle, and the ’signal’ for stationary-phase induction, are discussed in relation to the heat-inducibility of this gene.

Determination of the HSP2 6 gene nucleotide sequence, identified a number of sequence motifs in the promoter region which are characteristic of both heat-shock genes, and efficiently expressed genes in yeast. Sequence analysis of the HSP26 protein coding region identified a single open reading frame potentially encoding a 214 amino acid polypeptide of predicted molecular weight 27,630. This correlated well with the size of Hsp26 as estimated by it’s migration on a 2D-SDS PAGE gel. The predicted amino acid sequence of Hsp26 revealed homologies to other eukaryotic small Hsps, and also to bovine cc-crystall in A, homology being particularly strong in a predicted hydrophobic region of the Hsp26 polypeptide. In addition, with the exception of the amino-terminal methionine (Met), Hsp26 contains no Met residues, consistent with a failure to label the protein efficiently, .in vivo, with [3 5 S ] — methionine and demonstrating that the amino-terminal Met is cleaved post-translationally by an aminopeptidase.

The biological function of Hsp26 remains unclear. Contrary to evidence that the small Hsps of a number of eukaryotic organisms appear to be responsible for a tolerance of cells to potentially lethal high temperatures, overexpression of the HSP26 gene on a high copy number yeast episomal plasmid, demonstrated that overexpression of the protein following a heat-shock contributes little to acquired thermotolerance. However, overexpression of Hsp26 in stationary-phase cells revealed the presence of 45-55nm diameter cytoplasmic particles as putative Hsp26 aggregates. That the hydrophobicity of Hsp26 results in it’s self aggregation, is presumably central to the function of this protein.