title: Taxol mediates serine phosphorylation of the 66-kDa Shc isoform.
abstract: In the human lung carcinoma cell line A549, Taxol (20 nM) causes a decreased electrophoretic mobility of the 66-kDa Shc isoform (p66shc), beginning 4 h after drug exposure, and reaching a maximum at 9-18 h. No shift was observed for the 52- and 46-kDa isoforms of Shc. The electrophoretic mobility shift of p66shc caused by Taxol is not the result of tyrosine phosphorylation, and there is no indication of a Shc/Grb2 complex in Taxol-treated A549 cells. This modification is blocked by the serine/threonine protein phosphatase 2A. In vivo 32P-labeling and subsequent phosphoamino acid analysis of p66shc indicated that both the original and the shifted p66shc were predominantly serine phosphorylated. Cyanogen bromide digestion of p66shc produced a phosphorylated fragment with an apparent molecular weight of approximately 7.9 kDa from the untreated cells and two phosphorylated fragments, of approximately 7.9 and approximately 9.6 kDa, from the Taxol-treated cells. The domain of Taxol-induced serine phosphorylation is thought to be in the cyanogen bromide fragment containing residues 2-65. The Taxol-induced electrophoretic mobility shift of p66shc was inhibited by the protein synthesis inhibitor, cycloheximide, but not by the mitogen-activated and extracellular signal-regulated protein kinase kinase (MEK) inhibitor, PD98059. This mobility shift did not occur in Taxol-resistant A549-T12 cells treated with 20 nM Taxol. In addition to Taxol, other microtubule-interacting drugs caused a decreased electrophoretic mobility of p66shc. This Taxol-mediated serine phosphorylation seen in p66shc may result from a MEK-independent signaling pathway that is activated in cells that have a prolonged or abnormal mitotic phase of the cell cycle and may play a role in signaling events that lead to cell death.
authors.affiliation: Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: C P
authors.initials: CP
authors.lastname: Yang
authors.forename: S B
authors.initials: SB
authors.lastname: Horwitz
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D002277:Carcinoma / Q000378:metabolism; D003488:Cyanogen Bromide / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D004347:Drug Interactions; D019008:Drug Resistance, Neoplasm; D015180:Electrophoresis, Gel, Two-Dimensional; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors; D008870:Microtubules / Q000187:drug effects; D017239:Paclitaxel / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D010446:Peptide Fragments / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000972:Antineoplastic Agents, Phytogenic; D004791:Enzyme Inhibitors; D005419:Flavonoids; D010446:Peptide Fragments; D020033:Protein Isoforms; D011500:Protein Synthesis Inhibitors; D011506:Proteins; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D012694:Serine; D003513:Cycloheximide; D019908:Proto-Oncogene Proteins c-raf; D020930:MAP Kinase Kinase Kinases; D003488:Cyanogen Bromide; D017239:Paclitaxel; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one