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Chromosomal instability at a mutational hotspot in polyoma middle T-antigen affects its ability to activate the

P Rodriguez-Viciana1, C H Collins, M G Moule

  • 1UCSF Cancer Research Institute, 2340 Sutter Street, San Francisco, California 94115, USA.

Oncogene
|November 2, 2005
PubMed

Insights

Spontaneous polyoma virus middle T-antigen (PyMT) mutants were found to disrupt the ARF-p53 pathway, leading to untransformed cells. These PyMT mutations, located at a cytosine hotspot, alter protein structure and localization, affecting cell division.

Area of Science:

  • Virology
  • Molecular Biology
  • Cancer Research

Background:

  • Polyoma virus middle T-antigen (PyMT) is known to activate oncogenic pathways.
  • The ARF-p53 pathway plays a critical role in cell cycle regulation and tumor suppression.
  • Understanding PyMT's interaction with cellular pathways is crucial for cancer research.

Purpose of the Study:

  • To isolate and characterize spontaneous PyMT mutants.
  • To investigate the mechanism by which these mutants affect the ARF-p53 pathway and cell division.
  • To identify the specific mutational sites responsible for altered PyMT function.

Main Methods:

  • Isolation of spontaneous PyMT mutants in REF52 cells.
  • Analysis of cell morphology and ARF protein expression.
  • DNA sequencing to identify mutations in the PyMT gene, focusing on a cytosine-rich hotspot.
  • Assessment of PyMT protein localization and its impact on cell division.

Main Results:

  • Isolated PyMT mutants that fail to activate the ARF-p53 pathway, resulting in untransformed REF52 cells lacking ARF protein.
  • Identified a mutational hotspot (9 consecutive cytosines) in the PyMT sequence responsible for these mutations.
  • Frameshift mutations (deletion or insertion of cytosines) altered PyMT proteins, leading to loss of the C-terminal region and mislocalization.
  • Wild-type PyMT revertants were readily detected, originating from secondary mutations within the hotspot.

Conclusions:

  • PyMT mutations at the identified hotspot disrupt the ARF-p53 pathway by altering protein structure and localization.
  • These alterations lead to a loss of PyMT's transforming ability and cell cycle control.
  • The mutational hotspot provides a mechanism for generating functionally distinct PyMT variants and for reversion to wild-type activity.

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