Taxol mediates serine phosphorylation of the 66-kDa Shc isoform

Huang Yang Chia-Ping Huang Yang, Susan Band Horwitz

Research output: Contribution to journalArticle

48 Citations (Scopus)

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. Tbe electropboretic 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 ~7.9 kDa from the untreated cells and two phosphorylated fragments, of ~7.9 and ~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.

Original languageEnglish (US)
Pages (from-to)5171-5178
Number of pages8
JournalCancer Research
Volume60
Issue number18
StatePublished - Sep 15 2000

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Paclitaxel
Serine
Protein Isoforms
Phosphorylation
Cyanogen Bromide
Phosphoamino Acids
MAP Kinase Kinase Kinases
Protein Phosphatase 2
Protein Synthesis Inhibitors
Mitogen-Activated Protein Kinase Kinases
Extracellular Signal-Regulated MAP Kinases
Threonine
Cycloheximide
Mitogens
Microtubules
Pharmaceutical Preparations
Protein Kinases
Tyrosine
Digestion
Cell Cycle

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Taxol mediates serine phosphorylation of the 66-kDa Shc isoform. / Chia-Ping Huang Yang, Huang Yang; Band Horwitz, Susan.

In: Cancer Research, Vol. 60, No. 18, 15.09.2000, p. 5171-5178.

Research output: Contribution to journalArticle

Chia-Ping Huang Yang, HY & Band Horwitz, S 2000, 'Taxol mediates serine phosphorylation of the 66-kDa Shc isoform', Cancer Research, vol. 60, no. 18, pp. 5171-5178.
Chia-Ping Huang Yang, Huang Yang ; Band Horwitz, Susan. / Taxol mediates serine phosphorylation of the 66-kDa Shc isoform. In: Cancer Research. 2000 ; Vol. 60, No. 18. pp. 5171-5178.
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N2 - 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. Tbe electropboretic 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 ~7.9 kDa from the untreated cells and two phosphorylated fragments, of ~7.9 and ~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.

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