Regulation of HEF1 expression and phosphorylation by TGF-beta 1 and cell adhesion.
Transforming growth factor-beta1 (TGF-beta1) is a multipotential cytokine, which regulates remodeling of tissue extracellular matrix during early tumorigenesis and wound healing. Human enhancer of filamentation-1 (HEF1), a multifunctional docking protein, is involved in integrin-based signaling, which affects cell motility, growth, and apoptosis. Our studies reveal that TGF-beta1 is a potent inducer of HEF1 gene transcription in human dermal fibroblasts. TGF-beta1 promoted HEF1 expression in a dose-dependent manner and resulted in a 16-fold increase in HEF1 protein level. TGF-beta1 had no effect on the stability of either HEF1 protein or mRNA. The TGF-beta1-induced HEF1 expression was independent of cell adhesion and resistant to cytoskeleton disruption. TGF-beta1 increased levels of both p105 and p115 HEF1 in adherent fibroblasts. Digestion with specific phosphatases indicated that the p115HEF1 resulted from serine/threonine phosphorylation of p105HEF1. The appearance of the p115HEF1 as well as tyrosine phosphorylation of p105HEF1 required cell adhesion and/or an organized cytoskeleton. An in vitro kinase assay indicated that p105HEF1 was a substrate for Src. PP1, a specific Src kinase inhibitor, was able to block adhesion-dependent tyrosine phosphorylation of p105HEF1. These findings suggest that TGF-beta1 regulates HEF1 gene expression and that HEF1 phosphorylation is dependent on cell adhesion and Src kinase activity.
Center for Cell Biology & Cancer Research, Albany Medical College, Albany, New York 12208, USA. zhengm@mail.amc.edu
Mingzhe
M
Zheng
Paula J
PJ
McKeown-Longo
D048868:Adaptor Proteins, Signal Transducing; D017209:Apoptosis; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D002384:Catalysis; D002448:Cell Adhesion; D002455:Cell Division; D002460:Cell Line; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D015151:Immunoblotting; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D013379:Substrate Specificity; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013912:Threonine / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1; D014443:Tyrosine / Q000378:metabolism; D015854:Up-Regulation
D048868:Adaptor Proteins, Signal Transducing; D019086:Bridged Bicyclo Compounds, Heterocyclic; C099942:NEDD9 protein, human; D010750:Phosphoproteins; D011500:Protein Synthesis Inhibitors; C508960:TGFB1 protein, human; D013844:Thiazoles; D053778:Thiazolidines; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D003609:Dactinomycin; D015638:Cytochalasin D; D013912:Threonine; D014443:Tyrosine; D012694:Serine; C037067:latrunculin A