Chromatin remodeling in the mammalian heat shock response.

摘要

Transcriptional activators regulate gene expression by a variety of mechanisms. They stimulate transcription through interactions with the basal transcription machinery, general transcription factors and chromatin remodeling factors. The cellular stress response is a highly regulated transcriptional response in which all of these interactions are called into play.; Upon exposure to a variety of stressors, the transactivator Heat Shock Factor One (HSF1) binds to the hsp70 promoter, induces release of a stable promoter-proximal paused RNA polymerase II complex, and stimulates production of full length hsp70 transcripts. These events are accompanied by transcription-independent chromatin remodeling. In vitro, mutations in acidic residues of the HSF1 activation domains impair reinitiation and targeting of the SWI/SNF chromatin remodeling complex while mutations in phenylalanine residues impair elongation on chromatin templates and abrogate SWI/SNF targeting.; The goal of the work described in this thesis is to elucidate the relationship between transcriptional elongation and chromatin remodeling during HSF1 function in vivo. To examine the phenotypes of the HSF1 mutations described above in a native chromatin context, immortalized fibroblasts from an HSF1 (−/−) mouse were used to make stable cell lines that express wild-type, acidic-mutant and phenylalanine-mutant human HSF1. In these cell lines, HSF1 is required for stress tolerance, hsp70 mRNA induction and heat-induced chromatin remodeling of the hsp70 gene. SWI/SNF is recruited to the hsp70 promoter by HSF1 following heat stress. Wild-type human HSF1 restores all of the functions of mouse HSF1. While cells expressing the acidic mutant survive heat stress as well as those expressing wild-type HSF1, they show reduced levels of hsp70 mRNA induction, heat shock-induced chromatin remodeling, and SWI/SNF recruitment. Phenylalanine-mutant hHSF1 does not confer stress tolerance, hsp70 induction, remodeling, or recruitment. However, this mutant does direct the production of high levels of abortive hsp70 transcripts following heat shock.; The ability to stimulate elongation on the hsp70 gene is critical for the heat shock response and transcriptional elongation by HSF1 is strongly correlated with its ability to direct chromatin remodeling by the SWI/SNF complex. This work supports a model of hsp70 activation in which HSF1-mediated recruitment of SWI/SNF and subsequent remodeling drive elongation through the gene.