Protein kinase C-independent activation of protein kinase D is involved in BMP-2-induced activation of stress mitogen-activated protein kinases JNK and p38 and osteoblastic cell differentiation

J Biol Chem. 2004 Jan 2;279(1):259-64. doi: 10.1074/jbc.M308665200. Epub 2003 Oct 22.

Abstract

An important role for JNK* and p38 has recently been discovered in the differentiating effect of bone morphogenetic protein 2 (BMP-2) on osteoblastic cells. In this study, we investigated the molecular mechanism by which BMP-2 activates JNK and p38 in MC3T3-E1 osteoblastic cells. Activation of JNK and p38 induced by BMP-2 was blocked by the protein kinase C/protein kinase D (PKC/PKD) inhibitor Go6976 but not by the related compound, Go6983, a selective inhibitor of conventional PKCs. Associated with this inhibitory effect of Go6976, BMP-2 induced a selective and a dose-dependent Ser916 phosphorylation/activation of PKD, which was also blocked by Go6976. In contrast to the recently described PKC-dependent molecular mechanism involved in activation of PKD by G protein-coupled receptor agonists, BMP-2 did not induce a phosphorylation of PKD on Ser744/748. To further document an implication of PKD in activation of JNK and p38 induced by BMP-2, we constructed MC3T3-E1 cells stably expressing PKD antisense oligonucleotide (AS-PKD). In AS-PKD clones having low PKD levels, activation of JNK and p38 by BMP-2, but not of Smad1/5, was markedly impaired compared with empty vector transfected (V-PKD) cells. Analysis of osteoblastic cell differentiation in AS-PKD compared with V-PKD cells showed that mRNA and protein expressions of alkaline phosphatase and osteocalcin induced by BMP-2 were markedly reduced in AS-PKD. In conclusion, results presented in this study indicate that BMP-2 can induce activation of PKD in osteoblastic cells by a PKC-independent mechanism and that this kinase is involved in activation of JNK and p38 induced by BMP-2. Thus, this pathway, in addition to Smads, appears to be essential for the effect of BMP-2 on osteoblastic cell differentiation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3T3 Cells
  • Animals
  • Base Sequence
  • Bone Morphogenetic Protein 2
  • Bone Morphogenetic Proteins / pharmacology*
  • Carbazoles / pharmacology
  • Cell Differentiation
  • Enzyme Activation
  • Enzyme Inhibitors / pharmacology
  • Indoles / pharmacology
  • Kinetics
  • Mice
  • Mitogen-Activated Protein Kinases / metabolism*
  • Molecular Sequence Data
  • Osteoblasts / cytology
  • Osteoblasts / enzymology*
  • Protein Kinase C / antagonists & inhibitors
  • Protein Kinase C / genetics
  • Protein Kinase C / metabolism*
  • RNA, Messenger / genetics
  • Transforming Growth Factor alpha / pharmacology
  • Transforming Growth Factor beta*
  • p38 Mitogen-Activated Protein Kinases

Substances

  • Bmp2 protein, mouse
  • Bone Morphogenetic Protein 2
  • Bone Morphogenetic Proteins
  • Carbazoles
  • Enzyme Inhibitors
  • Indoles
  • RNA, Messenger
  • Transforming Growth Factor alpha
  • Transforming Growth Factor beta
  • Go 6976
  • protein kinase D
  • Protein Kinase C
  • Mitogen-Activated Protein Kinases
  • p38 Mitogen-Activated Protein Kinases