MULTIPLE PROTEIN-DNA INTERACTIONS OVER THE YEAST HSC82 HEAT-SHOCK GENE PROMOTER

We have utilized DNase I and micrococcal nuclease (MNase) to map the c hromatin structure of the HSC82 heat shock gene of Saccharomyces cerev isiae, The gene is expressed at a high basal level which is enhanced 2 -3-fold by thermal stress, A single, heat-shock invariant DNase I hype rsensitive domain is found within the HSC82 chromosomal locus; it maps to the gene's 5' end and spans 250 bp of promoter sequence, DNase I g enomic footprinting reveals that within this hypersensitive region are four constitutive protein-DNA interactions, These map to the transcri ption initiation site, the TATA box, the promoter-distal heat shock el ement (HSE1) and a consensus GRF2 (REB1/Factor Y) sequence, However, t wo other potential regulatory sites, the promoter-proximal heat shock element (HSE0) and a consensus upstream repressor sequence (URS1), are not detectably occupied under either transcriptional state, In contra st to its sensitivity to DNase I, the nucleosome-free promoter region is relatively protected from MNase; the enzyme excises a stable nucleo protein fragment of similar to 210 bp, As detected by MNase, there are at least two sequence-positioned nucleosomes arrayed 5' of the promot er; regularly spaced nucleosomes exhibiting an average repeat length o f 160-170 bp span several kilobases of both upstream and downstream re gions, Similarly, the body of the gene, which exhibits heightened sens itivity to DNase I, displays a nucleosomal organization under both bas al and induced states, but these nucleosomes are not detectably positi oned with respect to the underlying DNA sequence and may be irregularl y spaced and/or structurally altered, We present a model of the chroma tin structure of HSC82 and compare it to one previously derived for th e closely related, but differentially regulated, HSP82 heat shock gene .