Constitutive Synthesis Heat Shock Proteins and Altered Growth cerevisiae

A heat shock-resistant mutant of the budding yeast Saccharomyces cerevisiae was isolated at the mutation frequency of 10 -7 from a culture treated with ethyl methane sulfonate. Cells of the mutant are approximately 1,000-fold more resistant to lethal heat shock than those of the parental strain. Tetrad analysis indicates that phenotypes revealed by this mutant segregated together in the ratio 2+:2 from heterozygotes constructed with the wild-type strain of the opposite mating type, and are, therefore, attributed to a single nuclear mutation. The mutated gene in the mutant was herein designated hsrl (heat shock response). The hsrl allele is recessive to the HSR1 + allele of the wild-type strain. Exponentially growing cells of hsrl mutant were found to constitutively synthesize six proteins that are not synthesized or are synthesized at reduced rates in HSR1 + cells unless appropriately induced. These proteins include one hsp/G0-protein (hsp48A), one hsp (hsp48B), and two G0-proteins (p73, p56). Heterozygous diploid (hsrl/HSR1 +) cells do not synthesize the proteins constitutively induced in hsrl cells, which suggests that the product of the HSR1 gene might negatively regulate the synthesis of these proteins. The hsrl mutation also led to altered growth of the mutant cells. The mutation elongated the duration of G1 period in the cell cycle and affected both growth arrest by sulfur starvation and growth recovery from it. We discuss the problem of which protein(s) among those constitutively expressed in growing cells of the hsrl mutant is responsible for heat shock resistance and alterations in the growth control. Various organisms and cells are induced to synthesize a particular set of proteins, termed heat shock proteins (hsps), in response to an elevation in temperature (1, 19). These proteins are also induced as a result of cellular response to stress other than heat shock (1, 19). Evidence has been provided that the response protects cells from the stress (10, 11, 13, 14, 23), although the mechanism by which an accumulation of hsps within cells makes these cells resistant to further stress remains unclear. We have recently made a new finding that eucaryotic cells specifically synthesize hsps when they enter the resting state, Go (8). The induction of hsps in Go cells is distinct in two respects from that in heat-shocked cells. First the former is J Abbreviations used in this paper." hsps, heat shock proteins; 2DNEPHGE/SDS PAGE, two-dimensional nonequilibrium pH gradient electrophoresis/SDS PAGE. durable whereas the latter is transient. Second, Go cells synthesized mostly high molecular weight hsps and not low molecular weight hsps. These observations led us to hypothesize that high molecular weight hsps might function in the cellular transition from the proliferating state to Go and/or in the maintenance of the Go state. In the present study, we have intended to test this hypothesis by isolating heat shock-resistant mutants of the budding yeast, Saccharomyces cerevisiae, in which the hsp genes are expected to be constitutively expressed, and by examining the mutants for their properties with respect to growth control. The above finding (8) may indicate the alternative possibility that a particular class of hsps synthesized in Go cells do not function in the growth control whereas the synthesis of these hsps and that of functional proteins involved in the transition to Go are coordinatively regulated under the same mechanism. The isolation of hsp-constitutive mutants would THE JOURNAL OF CELL BIOLOGY • VOLUME 99 OCTOBER 1984 1441-1450 © The Rockefeller University Press • 0021-9525/84/10/1441/10 $1.0