title: Role of p38 in stress activation of Sp1.
abstract: Cell stressors such as physical forces can activate Sp1-dependent genes but the regulatory mechanisms are not defined. We determined if the stress-induced MAP kinase, p38, can phosphorylate Sp1 and thereby regulate the Sp1 target gene FLNA. We used Rat-2 cells and human gingival fibroblasts to examine stress-induced activation of an Sp1-dependent gene and SL2 cells, an Sp1-deficient model system, to facilitate interaction studies of transfected Sp1 with regulatory factors. Mechanical stress applied to Rat-2 cells increased promoter activity of the Sp1 target gene filamin A by >5-fold; activation was blocked by mutations to Sp1 binding sites in the filamin A promoter. Transfection experiments in SL2 cells with Sp1 expression vectors showed that when co-transfected with constitutively active p38, wild-type Sp1 but not an Sp1 binding mutant, increased promoter activity of the Sp1 target gene, filamin A, and enhanced binding of nuclear extracts to a filamin A promoter oligonucleotide. Filamin A promoter activity was blocked by dominant negative p38. Sp1 that was phosphorylated at Thr453 and Thr739 by constitutively active p38 bound to the filamin A promoter more effectively than un-phosphorylated Sp1. Recombinant active p38 phosphorylated wild-type Sp1 in vitro while the Sp1 Thr453Thr739 double mutant protein showed >3-fold reduction of phosphorylation. We conclude that stress activation of p38 phosphorylates Sp1 at specific threonine residues, modifications which in turn enhance the expression of Sp1-dependent genes.
authors.affiliation: CIHR Group in Matrix Dynamics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Mario
authors.initials: M
authors.lastname: D'Addario
authors.forename: Pamela D
authors.initials: PD
authors.lastname: Arora
authors.forename: C A
authors.initials: CA
authors.lastname: McCulloch
mesh_terms: D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D003285:Contractile Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D064448:Filamins; D005786:Gene Expression Regulation; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D051381:Rats; D015398:Signal Transduction; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D013314:Stress, Mechanical; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism / Q000502:physiology*
chemical_list: D003285:Contractile Proteins; D064448:Filamins; D008840:Microfilament Proteins; D016329:Sp1 Transcription Factor; D013912:Threonine; D048051:p38 Mitogen-Activated Protein Kinases