Phosphorylation of Fli1 at threonine 312 by protein kinase C delta promotes its interaction with p300/CREB-binding protein-associated factor and subsequent acetylation in response to transforming growth factor beta.
Previous studies have shown that transforming growth factor beta (TGF-beta)-induced collagen gene expression involves acetylation-dependent dissociation from the human alpha2(I) collagen (COL1A2) promoter of the transcriptional repressor Fli1. The goal of this study was to elucidate the regulatory steps preceding the acetylation of Fli1. We first showed that TGF-beta induces Fli1 phosphorylation on a threonine residue(s). The major phosphorylation site was localized to threonine 312 located in the DNA binding domain of Fli1. Using several independent approaches, we demonstrated that Fli1 is directly phosphorylated by protein kinase C delta (PKC delta). Additional experiments showed that in response to TGF-beta, PKC delta is recruited to the collagen promoter to phosphorylate Fli1 and that this step is a prerequisite for the subsequent interaction of Fli1 with p300/CREB-binding protein-associated factor (PCAF) and an acetylation event. The phosphorylation of endogenous Fli1 preceded its acetylation in response to TGF-beta stimulation, and the blockade of PKC delta abrogated both the phosphorylation and acetylation of Fli1 in dermal fibroblasts. Promoter studies showed that a phosphorylation-deficient mutant of Fli1 exhibited an increased inhibitory effect on the COL1A2 gene, which could not be reversed by the forced expression of PCAF or PKC delta. These data strongly suggest that the phosphorylation-acetylation cascade triggered by PKC delta represents the primary mechanism whereby TGF-beta regulates the transcriptional activity of Fli1 in the context of the collagen promoter.
Asano
Yoshihide
Y
Division of Rheumatology and Immunology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Trojanowska
Maria
M
D000107:Acetylation / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D024042:Collagen Type I; D020405:Dermis / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D007231:Infant, Newborn; D008297:Male; D010766:Phosphorylation / Q000187:drug effects; D010769:Phosphothreonine / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D051745:Protein Kinase C-delta / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics / Q000378:metabolism*; D051785:Smad Proteins / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D050880:p300-CBP Transcription Factors / Q000378:metabolism*; D000099224:p300-CBP-Associated Factor
D003094:Collagen; D024042:Collagen Type I; D010769:Phosphothreonine; D051745:Protein Kinase C-delta; D051789:Proto-Oncogene Protein c-fli-1; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C078433:FLI1 protein, human