Abstract
Heat shock factor 1, the critical molecular regulator of the stress response is conserved throughout eukaryotic organisms and activates the transcription of heat shock genes. We now show that heat shock factor 1 inhibits the expression of c-fos, an immediate early gene that controls responses to extracellular stimuli for growth and differentiation. Heat shock factor 1 inhibits the transcription of the c-fos gene and antagonizes the activating effects of the signal transducing protein Ras on the c-fos promoter and on the promoter of another Ras responsive gene uPA. This property was specific for heat shock factor 1; c-fos repression was not seen with the structurally related protein heat shock factor 2. Repression involved different molecular mechanisms compared with those involved in transcriptional activation by heat shock factor 1 and specifically did not require binding to the c-fos promoter. Thus, in addition to its known role as a transcriptional activator of the cellular heat shock response, heat shock factor 1 also antagonizes the expression of Fos, a key component of the ubiquitous AP-1 transcription factor complex and as such could influence multiple aspects of cell regulation.
Highlights
Heat shock factor 1, the critical molecular regulator of the stress response is conserved throughout eukaryotic organisms and activates the transcription of heat shock genes
C-fos repression did not require the binding of heat shock factor 1 (HSF1) to the c-fos promoter because a HSF1 construct deficient in heat shock elements (HSE) binding activity and transcriptional activating function still repressed the c-fos promoter and no evidence was obtained for HSF1 binding to the promoter (Fig. 2)
The mechanisms of c-fos repression by HSF1 contrast with those underlying transcriptional activation, which are dependent on HSF1 binding to HSE sequences within hsp promoters [1, 2]
Summary
Human HSP70B promoter construct p2500CAT was purchased from StressGen (Victoria, BC, Canada), and the human HSP27 promoter was a gift from Drs Lee Weber and Eileen Hickey (Department of Biology, University of Nevada). PGL.hsp was constructed by inserting the HSP27 gene promoter fragment [19] from BamHI site to HindIII site (730 base pairs) into pGL.Basic 3 at the BglII and HindIII sites. Pfos 2000 was constructed by inserting the human c-fos promoter from Ϫ2000 to ϩ42 relative to the transcription start site into pGL.Basic 2. The fragment from pstI to VspI sites of pSV--Galactosidase plasmid, containing SV40 early promoter and enhancer segments, lacZ coding region and SV40 poly(A) signals, was inserted into the pHSF plasmids at NsiI and SapI sites to replace the Neomycin resistance gene, and these constructs were termed p-B-HSF1 and p-B-HSF2, respectively. P-B-control plasmid was derived from p-B-HSF1 by deleting HSF1 at the XhoI and EcoRI sites. Double stranded DNA templates were prepared from minipreps as described [21]
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