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Serine phosphorylation of NPM-ALK, which is dependent on the auto-activation of the kinase activation loop,

Peng Wang1, Fang Wu, Jingdong Zhang

  • 1Department of Laboratory Medicine and Pathology, Cross Cancer Institute, University of Alberta, Edmonton, Alberta, Canada.

Carcinogenesis
|November 4, 2010
PubMed

Insights

This study identifies serine phosphorylation of nucleophosmin (NPM)-anaplastic lymphoma kinase (ALK) for the first time. This serine phosphorylation enhances the oncogenic protein

Area of Science:

  • Oncology
  • Molecular Biology
  • Biochemistry

Background:

  • Nucleophosmin (NPM)-anaplastic lymphoma kinase (ALK) is an oncogenic tyrosine kinase.
  • Its tumorigenic potential is linked to tyrosine phosphorylation.
  • The role of serine phosphorylation in NPM-ALK function was previously unexplored.

Purpose of the Study:

  • To investigate the role of serine phosphorylation in NPM-ALK.
  • To identify specific serine residues involved in NPM-ALK phosphorylation.
  • To determine the impact of serine phosphorylation on NPM-ALK tumorigenicity.

Main Methods:

  • Tandem affinity purification-mass spectrometry (TAP-MS) to identify phosphorylated serine residues.
  • Immunoprecipitation with anti-phosphoserine antibody to confirm serine phosphorylation.
  • Site-directed mutagenesis to assess the biological importance of serine residues.
  • Cell viability and clonogenic assays to evaluate tumorigenicity.
  • Pharmacological inhibition of serine kinases.

Main Results:

  • Three serine residues (Serine135, Serine164, Serine497) on NPM-ALK were identified as phosphorylated.
  • Serine phosphorylation was dependent on ALK activation loop status.
  • Mutating these serine residues significantly reduced NPM-ALK tumorigenicity.
  • Reduced tumorigenicity correlated with decreased phosphorylation of downstream signaling molecules (ERK1/2, JNK, STAT6).
  • Serine phosphorylation was regulated by multiple serine kinases, including GSK-3, CK1, and MAPKs.

Conclusions:

  • This study is the first to identify serine phosphorylation of NPM-ALK.
  • Serine phosphorylation of NPM-ALK enhances its tumorigenic potential.
  • NPM-ALK serine phosphorylation is regulated by various serine kinases and impacts downstream signaling pathways.

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