title: Phosphatidylinositol 3-kinase is involved in alpha2(I) collagen gene expression in normal and scleroderma fibroblasts.
abstract: TGF-beta is implicated in the pathogenesis of fibrotic disorders. It has been shown that Smad3 promotes the human alpha2(I) collagen (COL1A2) gene expression by TGF-beta1 in human dermal fibroblasts. Here, we investigated the role of phosphatidylinositol 3-kinase (PI3K) in the COL1A2 gene expression in normal and scleroderma fibroblasts. In normal fibroblasts, the PI3K inhibitor, LY294002, significantly decreased the basal and the TGF-beta1-induced increased stability of COL1A2 mRNA. The TGF-beta1-induced COL1A2 promoter activity, but not the basal activity, was significantly attenuated by LY294002 or the dominant negative mutant of p85 subunit of PI3K, while the constitutive active mutant of p110 subunit of PI3K did not affect the basal or the TGF-beta1-induced COL1A2 promoter activity. LY294002 significantly decreased the phosphorylation of Smad3 induced by TGF-beta1. Furthermore, the transient overexpression of 2xFYVE, which induces the mislocalization of FYVE domain proteins, decreased the TGF-beta1-induced Smad3 phosphorylation to a similar extent to LY294002. In scleroderma fibroblasts, the blockade of PI3K significantly decreased the mRNA stability and the promoter activity of the COL1A2 gene. Furthermore, LY294002 and the transient overexpression of 2xFYVE completely diminished the constitutive phosphorylation of Smad3. These results indicate that 1) the basal activity of PI3K is necessary for the COL1A2 mRNA stabilization in normal and scleroderma fibroblasts, 2) there is an unidentified FYVE domain protein specifically interacting with Smad3, and 3) the basal activity of PI3K and the FYVE domain protein are indispensable for the efficient TGF-beta/Smad3 signaling in normal fibroblasts and for the establishment of the constitutive activation of TGF-beta/Smad3 signaling in scleroderma fibroblasts.
authors.affiliation: Department of Dermatology, Faculty of Medicine, University of Tokyo, Tokyo, Japan.
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Asano
authors.forename: Hironobu
authors.initials: H
authors.lastname: Ihn
authors.forename: Kenichi
authors.initials: K
authors.lastname: Yamane
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Jinnin
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Mimura
authors.forename: Kunihiko
authors.initials: K
authors.lastname: Tamaki
mesh_terms: D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D024042:Collagen Type I / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D045743:Scleroderma, Diffuse / Q000378:metabolism*; D051900:Smad3 Protein; D015534:Trans-Activators / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000494:pharmacology
chemical_list: D002867:Chromones; D024042:Collagen Type I; D004268:DNA-Binding Proteins; D009025:Morpholines; C497790:SMAD3 protein, human; D051900:Smad3 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D004247:DNA; D019869:Phosphatidylinositol 3-Kinases