Regulation of human heat shock transcription factor I (hHSF1)

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)


Biochemistry and Molecular Biology

First Committee Member

Richard W. Voellmy, Committee Chair


Heat regulation of human heat shock genes is mediated by heat shock transcription factor I (hHSF1), that, like HSFs from other higher eukaryotes, contains three leucine zippers (LZ 1-3) or hydrophobic repeats of the 4-3 type. In the unstressed human cell (37$\sp\circ$C), hHSF1 appears to be in an inactive, monomeric form. Upon heat shock (40-42$\sp\circ$C), hHSF1 homotrimerizes and acquires HSE (heat shock element, present in all heat-inducible promoters) DNA-binding ability and transcriptional competence. When expressed in Xenopus oocytes, hHSF1 also appears to be in a monomeric, non-DNA-binding state and is converted to a trimeric, DNA-binding form upon heat shock (35-37$\sp\circ$C of this organism). In a systematic analysis of heat-regulated DNA-binding ability and trimerization of hHSF1, using a series of internal deletion mutants expressed in oocytes, three discrete regions containing LZ 1-3 were identified as being required for maintaining the factor as an inactive monomer at the non-heat shock temperature (20$\sp\circ$C). Disruption of LZ structures abolished heat regulation of hHSF1. Single amino acid substitutions of specific hydrophobic zipper residues in each LZ rendered the factor constitutively trimeric and DNA binding. That the same structural elements (except LZ1) maintain hHSF1 in an inactive monomeric state in the unstressed human cell was revealed by transient expression experiments. These results strongly suggest that the inactive hHSF1 monomer is maintained by hydrophobic interactions involving all three LZs which may form a triple-stranded coiled-coil. Model experiments with LexA-hHSF1 chimeras suggest that trimerization of hHSF1 serves to concentrate DNA-binding domains, facilitating their cooperative binding to target DNA.In transient expression experiments with human HeLa cells, overexpressed hHSF1 is predominantly in a trimeric form capable of DNA binding in the absence of stress. However, this overexpressed, trimeric hHSF1 is incapable of transactivating a heat shock gene. A second, apparently distinct conformational change, triggered by heat shock or mimicked by mutations in LZ2 or a specific region between LZs 2 and 3, is required to derepress the transcriptional ability of the factor. An extended region spanning the carboxy-terminal third of the hHSF1 sequence was identified as containing the transcription activation domain(s). A nuclear localization signal of hHSF3 was mapped to a cluster of basic residues preceding LZ1.


Biology, Molecular

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