Bryostatin 1 induces prolonged activation of extracellular regulated protein kinases in and apoptosis of LNCaP human prostate cancer cells overexpressing protein kinase calpha

Mol Pharmacol. 2000 Jun;57(6):1224-34.

Abstract

Previously, we reported that 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced apoptosis of LNCaP human prostate cancer cells was accompanied by prolonged translocation of protein kinase C (PKC)alpha to non-nuclear membranes and that TPA-resistant LNCaP cells had down-regulated PKCalpha. Here we show that 10 nM bryostatin 1 induced transient membrane translocation and down-regulation of PKCalpha, prolonged translocation of PKCdelta and epsilon to non-nuclear membranes, and did not induce cell death but blocked TPA-induced apoptosis. To test the hypothesis that inhibition of TPA-induced apoptosis by bryostatin 1 was due to down-regulation of PKCalpha, we inducibly overexpressed PKCalpha in LNCaP cells. Overexpression of PKCalpha alone did not induce apoptosis, even in clones that contained much more membrane-bound, active PKCalpha than was observed in TPA-treated untransfected LNCaP cells. However, the addition of 10 nM bryostatin 1 to PKCalpha-overexpressing LNCaP cells did not yield down-regulation of PKCalpha and induced extensive apoptosis. Immunoblot analysis revealed that TPA induced prolonged hyperphosphorylation of Raf-1 and activation of extracellular-regulated/mitogen-activated protein kinases 1 and 2 in untransfected LNCaP cells, as did bryostatin 1 in PKCalpha-overexpressing cells. On the other hand, bryostatin 1 induced only transient hyperphosphorylation of Raf-1 and activation of extracellular-regulated/mitogen-activated protein kinases 1 and 2 in untransfected LNCaP cells. These results confirm a role of prolonged membrane-associated PKCalpha in PKC activator-mediated LNCaP apoptosis and suggest involvement of the mitogen-activated protein kinase pathway.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antineoplastic Agents / pharmacology*
  • Apoptosis*
  • Biological Transport
  • Bryostatins
  • Carcinogens / pharmacology
  • Cell Division / drug effects
  • Down-Regulation
  • Enzyme Activation / drug effects
  • Humans
  • Isoenzymes / biosynthesis*
  • Lactones / pharmacology*
  • Macrolides
  • Male
  • Mitogen-Activated Protein Kinases / metabolism*
  • Phosphorylation
  • Prostatic Neoplasms / pathology
  • Protein Kinase C / biosynthesis*
  • Protein Kinase C-alpha
  • Proto-Oncogene Proteins c-raf / metabolism
  • Subcellular Fractions
  • Tetradecanoylphorbol Acetate / pharmacology
  • Tumor Cells, Cultured

Substances

  • Antineoplastic Agents
  • Bryostatins
  • Carcinogens
  • Isoenzymes
  • Lactones
  • Macrolides
  • bryostatin 1
  • Proto-Oncogene Proteins c-raf
  • PRKCA protein, human
  • Protein Kinase C
  • Protein Kinase C-alpha
  • Mitogen-Activated Protein Kinases
  • Tetradecanoylphorbol Acetate