title: Cyclic AMP enhances Smad-mediated BMP signaling through PKA-CREB pathway.
abstract: We present experimental results indicating involvement of cyclic AMP (cAMP)-mediated signaling in bone morphogenetic protein (BMP)-induced osteoblastic gene expression at the transcriptional level by luciferase activity assay in C2C12 cells using the promoter sequence of the Id1 gene, an early-response gene to BMPs, which contains both a BMP-responsive element (BRE) and a cAMP-response element (CRE). In cells transfected with luciferase gene driven by wild-type Id1 promoter, treatment with BMP-4 increased luciferase expression, which was further enhanced by the addition of dibutyryl cAMP (dbcAMP). This dbcAMP-enhanced luciferase expression was significantly suppressed when the CRE site in the Id1 promoter was replaced by mutated CRE or endogenous CRE-binding protein (CREB) was knocked down by transfection of CREB RNAi. Pretreatment of cells with protein kinase A (PKA) inhibitor, H89, also dramatically reduced dbcAMP-enhanced luciferase expression. Immunoprecipitation assay showed phosphorylated-Smad1/5/8, phosphorylated-CREB, and CREB-binding protein (CBP) formed the transcriptional complex. These data indicate that cAMP-PKA/CREB/CRE signaling potentially enhances BMP-induced transcription through the BRE in the promoter of the BMP-responsive gene through a PKA-mediated pathway.
authors.affiliation: Department of Orthopaedic Surgery, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka, Japan. ohta@msic.med.osaka-cu.ac.jp
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Ohta
authors.forename: Keisuke
authors.initials: K
authors.lastname: Nakagawa
authors.forename: Yuuki
authors.initials: Y
authors.lastname: Imai
authors.forename: Takenobu
authors.initials: T
authors.lastname: Katagiri
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Koike
authors.forename: Kunio
authors.initials: K
authors.lastname: Takaoka