title: Uncoupling Stress-Inducible Phosphorylation of Heat Shock Factor 1 from Its Activation.
abstract: In mammals the stress-inducible expression of genes encoding heat shock proteins is under the control of the heat shock transcription factor 1 (HSF1). Activation of HSF1 is a multistep process, involving trimerization, acquisition of DNA-binding and transcriptional activities, which coincide with several posttranslational modifications. Stress-inducible phosphorylation of HSF1, or hyperphosphorylation, which occurs mainly within the regulatory domain (RD), has been proposed as a requirement for HSF-driven transcription and is widely used for assessing HSF1 activation. Nonetheless, the contribution of hyperphosphorylation to the activity of HSF1 remains unknown. In this study, we generated a phosphorylation-deficient HSF1 mutant (HSF1Δ∼PRD), where the 15 known phosphorylation sites within the RD were disrupted. Our results show that the phosphorylation status of the RD does not affect the subcellular localization and DNA-binding activity of HSF1. Surprisingly, under stress conditions, HSF1Δ∼PRD is a potent transactivator of both endogenous targets and a reporter gene, and HSF1Δ∼PRD has a reduced activation threshold. Our results provide the first direct evidence for uncoupling stress-inducible phosphorylation of HSF1 from its activation, and we propose that the phosphorylation signature alone is not an appropriate marker for HSF1 activity.
authors.affiliation: Department of Biosciences, Åbo Akademi University, Turku, Finland Turku Center for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
authors.forename: Marek A
authors.initials: MA
authors.lastname: Budzyński
authors.affiliation: Department of Biosciences, Åbo Akademi University, Turku, Finland Turku Center for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
authors.forename: Mikael C
authors.initials: MC
authors.lastname: Puustinen
authors.affiliation: Department of Biosciences, Åbo Akademi University, Turku, Finland Turku Center for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
authors.forename: Jenny
authors.initials: J
authors.lastname: Joutsen
authors.affiliation: Department of Biosciences, Åbo Akademi University, Turku, Finland Turku Center for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland lea.sistonen@abo.fi.
authors.forename: Lea
authors.initials: L
authors.lastname: Sistonen
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002465:Cell Movement / Q000235:genetics; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D006367:HeLa Cells; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013312:Stress, Physiological; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D004268:DNA-Binding Proteins; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins; C490017:Hsf1 protein, mouse; D050505:Mutant Proteins; D014157:Transcription Factors; D004247:DNA