Project description:To examine whether human HSF1, chicken HSF1 and chicken HSF3 regulates the expression of different genes, we performed DNA microarray analysis.
Project description:To analyze target genes of human heat shock transcription factor 1 (HSF1), we first generated two independent HeLa clones (RDT1 and RDT2) expressing an actively mutated hHSF1 (hHSF1ΔRDT), which lacks the regulatory domain that masks its activation domain and possesses a glutamic acid at amino acid 395 instead of a leucine in the suppression domain of the trimerization domain (Fujimoto et al., J. Biol. Chem. 280, 34908-34916, 2005). We also generated a HeLa clone expressing chicken HSF1 (HeLa/cHSF1) to compare its profile of gene expression with those of RDT1 and RDT2 cells (Nakai and Morimoto, Mol. Cell. Biol. 13, 1983-1997, 1993). We then carried out DNA microarray analysis using total RNA isolated from HeLa, HeLa/cHSF1, RDT1, and RDT2 cells grown under normal growth conditions.
Project description:Transcription factor access to regulatory elements is prevented by the nucleosome. Heat shock factor 1 (HSF1) is a winged helix transcription factor that plays roles in control and stressed conditions by gaining access to target elements, but mechanisms of HSF1 access have not been well known in mammalian cells. We show a physical interaction between the wing motif of human HSF1 and replication protein A (RPA), which is involved in DNA metabolism. Depletion of RPA1 abolishes HSF1 access to the promoter of HSP70 in unstressed conditions, and delays its rapid activation in response to heat shock. The HSF1-RPA complex leads preloading of RNA polymerase II and opens chromatin structure by recruiting a histone chaperone FACT. Furthermore, this interaction is required for melanoma cell proliferation. These results provide a mechanistic basis for constitutive HSF1 access to nucleosomal DNA, which is important for both basal and inducible gene expression. To examine whether the HSF1-RPA complex regulates the expression of genes, mRNA levels in MEFs with/without HSF1- or RPA1-shRNA were analyzed by DNA microarray analysis using GeneChip Mouse Gene 1.0 ST Arrays (Affymetrix).
Project description:To analyze target genes of human heat shock transcription factor 1 (HSF1), we first generated two independent HeLa clones (RDT1 and RDT2) expressing an actively mutated hHSF1 (hHSF1?RDT), which lacks the regulatory domain that masks its activation domain and possesses a glutamic acid at amino acid 395 instead of a leucine in the suppression domain of the trimerization domain (Fujimoto et al., J. Biol. Chem. 280, 34908-34916, 2005). We also generated a HeLa clone expressing chicken HSF1 (HeLa/cHSF1) to compare its profile of gene expression with those of RDT1 and RDT2 cells (Nakai and Morimoto, Mol. Cell. Biol. 13, 1983-1997, 1993). We then carried out DNA microarray analysis using total RNA isolated from HeLa, HeLa/cHSF1, RDT1, and RDT2 cells grown under normal growth conditions. mRNA levels in human HeLa, RDT1, RDT2, and HeLa/cHSF1 were analyzed by DNA microarray analysis using GeneChip Human Genome U133 Plus 2.0 Array (Affymetrix).
Project description:Transcription factor access to regulatory elements is prevented by the nucleosome. Heat shock factor 1 (HSF1) is a winged helix transcription factor that plays roles in control and stressed conditions by gaining access to target elements, but mechanisms of HSF1 access have not been well known in mammalian cells. We show a physical interaction between the wing motif of human HSF1 and replication protein A (RPA), which is involved in DNA metabolism. Depletion of RPA1 abolishes HSF1 access to the promoter of HSP70 in unstressed conditions, and delays its rapid activation in response to heat shock. The HSF1-RPA complex leads preloading of RNA polymerase II and opens chromatin structure by recruiting a histone chaperone FACT. Furthermore, this interaction is required for melanoma cell proliferation. These results provide a mechanistic basis for constitutive HSF1 access to nucleosomal DNA, which is important for both basal and inducible gene expression.
Project description:To examine whether HSF1-PARP1-PARP13 complex regulates the expression of genes, we performed DNA microarray analysis. We also performed DNA microarray analysis to identify doxorubicin-induced genes,.