We have isolated a gene from the yeast Saccharomyces cerevisiae that encodes a 2.0-kilobase heat-inducible mRNA. This gene, which we have designated STII, for stress inducible, was also induced by the amino acid analog canavanine and showed a slight increase in expression as cells moved into stationary phase. The STII gene encodes a 66-kilodalton protein, as determined from the sequence of the longest open reading frame. The putative STII protein, as identified by two-dimensional gel electrophoresis, migrated in the region of 73 to 75 kilodaltons as a series of four isoforms with different isoelectric points. STII is not homologous to the other conserved HSP70 family members in yeasts, despite similarities in size and regulation. Cells carrying a disruption mutation of the STI) gene grew normally at 30°C but showed impaired growth at higher and lower temperatures. Overexpression of the STII gene resulted in substantial trans-activation of SSA4 promoterreporter gene fusions, indicating that STII may play a role in mediating the heat shock response of some HSP70 genes.Cells from early every organism thus far tested respond to an increase in temperature by inducing synthesis of a stereotypical set of proteins, the so-called heat shock proteins. Synthesis of these proteins is also induced by a number of other stimuli that stress the cell, indicating that the response is a general one to organismal (and cellular) trauma. The most abundant of the heat shock proteins found after temperature upshift is a 70-kilodalton (kDa) protein called hsp70. hsp70 is the most highly conserved of all heat shock proteins, showing between 60 and 78% identity among eucaryotes and 40 to 50% identity between Escherichia coli hsp70, the dnaK gene product, and the corresponding eucaryotic proteins (reviewed in reference 22).Among the best characterized of the HSP70 genes are those from the yeast Saccharomyces cerevisiae. Eight genes related to HSP70 have been isolated as recombinant DNA clones in our laboratory. These eight genes have been divided into four phenotypic subfamilies, SSA through SSD (SS denotes stress seventy related; A through D indicate the subfamilies). The expression patterns and functional interrelationships of members of the SSA subfamily (encompassing SSAJ through SSA4) have been the most thoroughly investigated. The SSAI gene is expressed at moderate levels at 23°C but is induced approximately 10-fold after a temperature shift to 37°C, whereas SSA2 is expressed at a high level at all temperatures. SSA3 and SSA4 are expressed at extremely low levels at 23°C, but after a temperature upshift the amounts of SSA3 and SSA4 mRNAs increase dramatically (9