Characterization of constitutive HSF2 DNA-binding activity in mouse embryonal carcinoma cells.
Two distinct murine heat shock transcription factors, HSF1 and HSF2, have been identified. HSF1 mediates the transcriptional activation of heat shock genes in response to environmental stress, while the function of HSF2 is not understood. Both factors can bind to heat shock elements (HSEs) but are maintained in a non-DNA-binding state under normal growth conditions. Mouse embryonal carcinoma (EC) cells are the only mammalian cells known to exhibit HSE-binding activity, as determined by gel shift assays, even when maintained at normal physiological temperatures. We demonstrate here that the constitutive HSE-binding activity present in F9 and PCC4.aza.R1 EC cells, as well as a similar activity found to be present in mouse embryonic stem cells, is composed predominantly of HSF2. HSF2 in F9 EC cells is trimerized and is present at higher levels than in a variety of nonembryonal cell lines, suggesting a correlation of these properties with constitutive HSE-binding activity. Surprisingly, transcription run-on assays suggest that HSF2 in unstressed EC cells does not stimulate transcription of two putative target genes, hsp70 and hsp86. Genomic footprinting analysis indicates that HSF2 is not bound in vivo to the HSE of the hsp70 promoter in unstressed F9 EC cells, although HSF2 is present in the nucleus and the promoter is accessible to other transcription factors and to HSF1 following heat shock. Thus trimerization and nuclear localization of HSF2 do not appear to be sufficient for in vivo binding of HSF2 to the HSE of the hsp70 promoter in unstressed F9 EC cells.
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, Illinois 60208.
S P
SP
Murphy
J J
JJ
Gorzowski
K D
KD
Sarge
B
B
Phillips
D000818:Animals; D001483:Base Sequence; D018236:Carcinoma, Embryonal; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D005786:Gene Expression Regulation; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism*; D006358:Hot Temperature; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D017931:DNA Primers; D006360:Heat-Shock Proteins; D046911:Macromolecular Substances; D012333:RNA, Messenger; D014157:Transcription Factors; C070389:HSF2 protein, human