B-Raf(V600E) signaling deregulates the mitotic spindle checkpoint through stabilizing Mps1 levels in melanoma cells

Oncogene. 2008 May 15;27(22):3122-33. doi: 10.1038/sj.onc.1210972. Epub 2007 Dec 10.

Abstract

The B-Raf(V600E) mutant, found in 65% of human melanomas, drives constitutive activation of the extracellular signal-regulated kinase (ERK) pathway and is implicated in tumorigenesis. Recently, we showed that B-Raf is important for spindle formation and the mitotic spindle checkpoint arrest. In this study, we demonstrate that B-Raf(V600E) signaling deregulates the spindle checkpoint as a consequence of stabilizing monopolar spindle 1 (Mps1) levels in human melanoma cells. Upon introducing the B-Raf(V600E) mutant into wild-type B-Raf melanoma cells, Mps1 protein and activity increased 3- and 10-fold, respectively. In addition, Mps1 became hyperphosphorylated, which correlated with stabilization of Mps1 protein levels. In contrast, reduction of B-Raf by RNAi or inactivation of ERK by the MEK inhibitor U0126 resulted in a precipitous decline in Mps1 levels. Together, these results suggest that B-Raf signaling through ERK regulates the stability of Mps1. Finally, B-Raf(V600E) expression induces a mitotic delay due to promoting robust activation of the mitotic spindle checkpoint. These effects were dependent on the induction of Mps1 levels by oncogenic B-Raf(V600E) as shown by depleting Mps1 with short interfering RNA. Collectively, our findings implicate a new mechanism through which B-Raf(V600E) exerts its oncogenic effects in melanoma.

Publication types

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

MeSH terms

  • Amino Acid Substitution / physiology
  • Cell Cycle Proteins / metabolism*
  • Cell Division / genetics
  • Cell Line, Tumor
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Genes, cdc
  • Glutamic Acid / genetics
  • Humans
  • Melanoma / genetics*
  • Melanoma / metabolism
  • Phosphorylation
  • Polymorphism, Single Nucleotide / physiology
  • Protein Binding
  • Protein Processing, Post-Translational / genetics
  • Protein Serine-Threonine Kinases / metabolism*
  • Protein-Tyrosine Kinases
  • Proto-Oncogene Proteins B-raf / genetics
  • Proto-Oncogene Proteins B-raf / metabolism
  • Proto-Oncogene Proteins B-raf / physiology*
  • Signal Transduction / genetics
  • Signal Transduction / physiology
  • Spindle Apparatus / genetics*
  • Spindle Apparatus / metabolism
  • Transfection
  • Valine / genetics

Substances

  • Cell Cycle Proteins
  • Glutamic Acid
  • Protein-Tyrosine Kinases
  • BRAF protein, human
  • Protein Serine-Threonine Kinases
  • Proto-Oncogene Proteins B-raf
  • Extracellular Signal-Regulated MAP Kinases
  • TTK protein, human
  • Valine