Transcriptional regulation of a yeast HSP70 gene by heat shock factor and an upstream repression site-binding factor

Abstract

SSA1, one of the heat-inducible HSP70 genes in the yeast Saccharomyces cerevisiae, displays a significant basal level of expression under optimal growth conditions. Although multiple sites related to the heat shock element (HSE) consensus sequences are present in the SSA1 promoter region, one of these, HSE2, plays a key role in basal expression. An upstream repression site (URS) located adjacent to HSE2 causes repression of basal expression but has little effect on heat-inducible expression of SSA1. A series of DNase I footprinting assays suggests that heat shock factor (HSF) and a URS-binding factor (URSF) can bind simultaneously to the adjacent binding sites HSE2 and URS under optimal growth conditions. URSF in extracts from heat-shocked cells does not bind (or binds very poorly) to the URS adjacent to HSE2. However, URSF in these extracts is able to bind the URS if the URS is separated from HSE2 or if the HSE is mutated such that HSF binding is abolished. These in vitro experiments are consistent with in vivo results showing that the URS is able to repress transcription driven by HSE2 both before and after heat shock if it is separated from HSE2. Our results are consistent with a model of repression in which URSF and HSF bind simultaneously to the adjacent binding sites URS and HSE2 prior to heat shock. After heat shock, however, binding of the two proteins to the adjacent sites is exclusive, perhaps due to modification of HSF known to occur upon heat shock. Because HSF binding predominates, repression by URSF is relieved upon heat shock.