Peroxisome Proliferator-Activated Receptor- and Retinoic Acid X Receptor Represses the TGF 1 Gene via PTEN-Mediated p70 Ribosomal S6 Kinase-1 Inhibition: Role for Zf9 Dephosphorylation

Peroxisome proliferator-activated receptor (PPAR)and retinoic acid X receptor (RXR) heterodimer regulates cell growth and differentiation. Zinc finger transcription factor-9 (Zf9), whose phosphorylation promotes target genes, is a transcription factor essential for transactivation of the transforming growth factor (TGF)1 gene. This study investigated whether activation of PPAR -RXR heterodimer inhibits TGF 1 gene transcription and Zf9 phosphorylation and, if so, what signaling pathway regulates it. Either 15deoxy(12,14)-prostaglandin J2 (PGJ2) or 9-cis-retinoic acid (RA) treatment decreased the TGF 1 mRNA level in L929 fibroblasts. PGJ2 RA, compared with individual treatment alone, synergistically inhibited the TGF 1 gene expression, which was abrogated by PPAR antagonists. Likewise, PGJ2 RA decreased luciferase expression from the TGF 1 gene promoter. Promoter deletion analysis of the TGF 1 gene revealed that pGL3-323 making up to 323-base pair region, but lacking PPAR-responsive elements, responded to PGJ2 RA. PGJ2 RA treatment inhibited the activity of p70 ribosomal S6 kinase-1 (S6K1), abolishing Zf9 phosphorylation at serine as did rapamycin [a mammalian target of rapamycin (mTOR) inhibitor]. Zf9 dephosphorylation by PGJ2 RA was reversed by transfection of cells with the plasmid encoding constitutively active S6K1 (CA-S6K1). Transfection with dominant negative S6K1 inhibited the TGF 1 gene. TGF 1 gene repression by PGJ2 RA was consistently antagonized by CAS6K1. Ectopic expression of PPAR 1 and RXR repressed pGL3323 transactivation with S6K1 inhibition, which was abrogated by CA-S6K1 transfection. PGJ2 RA induced phosphatase and tensin homolog deleted on chromosome 10 (PTEN), whose overexpression repressed the TGF 1 gene through S6K1 inhibition, decreasing extracellular signal-regulated kinase 1/2–90-kDa ribosomal S6 kinase 1 and Akt-mTOR phosphorylations. Data indicate that activation of PPAR -RXR heterodimer represses the TGF 1 gene and induces Zf9 dephosphorylation via PTEN-mediated S6K1 inhibition, providing insight into pharmacological manipulation of the TGF 1 gene regulation. The human transforming growth factorisoforms constitute extracellular signaling molecules that have antiproliferative effects on mammalian cells, promoting the expression of cell adhesion molecules and extracellular matrix proteins. In particular, transforming growth factor (TGF)1 serves as a key fibrogenic mediator that can enhance extracellular matrix deposition and inhibit collagenase activity during fibrogenesis (Friedman, 1993). The regulation of TGF 1 expression is complex and occurs at multiple levels, orchestrated transcriptionally by the multiple transcription factors and post-translationally by maturation of the precursors bound with TGF 1 binding proteins (Kim et al., 1989a; Oklu