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Studying Proteolysis of Cyclin B at the Single Cell Level in Whole Cell Populations
10:54

Studying Proteolysis of Cyclin B at the Single Cell Level in Whole Cell Populations

Published on: September 17, 2012

Ubiquitin/proteasome-dependent degradation of D-type cyclins is linked to tumor necrosis factor-induced cell cycle

Xiaotang Hu1, Matthew Bryington, Ariana B Fisher

  • 1Interdisciplinary Oncology Program, Department of Internal Medicine, University of South Florida, and H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida 33612, USA. hu@moffitt.usf.edu

The Journal of Biological Chemistry
|February 28, 2002
PubMed

Insights

Tumor necrosis factor-alpha (TNF) induces G(1) cell cycle arrest and apoptosis in cancer cells. This process involves proteasome-dependent down-regulation of D-type cyclins, impacting cell cycle control.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Cancer Research

Background:

  • Tumor necrosis factor-alpha (TNF) is recognized for its cytotoxic effects on cancer cells.
  • The specific mechanisms by which TNF influences cell cycle regulation are less understood.

Purpose of the Study:

  • To investigate the role of TNF in cell cycle control.
  • To elucidate the molecular pathways involved in TNF-induced cell cycle arrest and apoptosis.

Main Methods:

  • TF-1 and MV4-11 cancer cell lines were treated with TNF.
  • Cell cycle progression was analyzed using flow cytometry.
  • Protein levels and kinase activities of cell cycle regulators (pRb, D-type cyclins, CDKs) were assessed.
  • Inhibitors of calpain (ALLN), apoptosis (zVAD-FMK), and proteasome (MG-132, lactacystin) were used to determine pathway involvement.
  • Proteasome activity was measured, and in vitro degradation assays were performed.

Main Results:

  • TNF induced G(1) cell cycle arrest and apoptosis in TF-1 and MV4-11 cells.
  • TNF treatment led to pRb dephosphorylation and reduced D-type cyclin expression, decreasing cyclin-dependent kinase activities.
  • Proteasome inhibitors (MG-132, lactacystin) blocked TNF-induced IkappaBalpha degradation and D-type cyclin down-regulation.
  • Calpain and apoptosis inhibitors did not prevent D-type cyclin down-regulation.
  • Proteasome activity increased in TNF-treated cells, and cyclin D3 was degraded by the 26 S proteasome in vitro.

Conclusions:

  • TNF-induced down-regulation of D-type cyclins is dependent on the proteasome pathway.
  • The proteasome plays a critical role in mediating TNF's effects on cell cycle regulation and apoptosis.

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