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Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin
07:56

Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin

Published on: January 17, 2012

A novel crosstalk mechanism between nuclear receptor-mediated and growth factor/Ras-mediated pathways through

Dujin Zhou1, Bin Chen, Jing-Jing Ye

  • 1Department of Surgical Research, Beckman Research Institute of the City of Hope, Duarte, CA 91010, USA.

Oncogene
|May 4, 2004
PubMed

Insights

Proline-rich nuclear receptor coregulatory protein (PNRC) interacts with Grb2, suppressing nuclear receptor and growth factor pathways. PNRC overexpression slowed cell growth, and its expression was lower in breast cancer tissues.

Area of Science:

  • Molecular Biology
  • Cellular Biology
  • Biochemistry

Background:

  • Proline-rich nuclear receptor coregulatory protein (PNRC) functions as a nuclear receptor coactivator.
  • PNRC interacts with nuclear receptors via an SH3-binding motif in its C-terminus.

Purpose of the Study:

  • To investigate the interaction between PNRC and Grb2.
  • To elucidate the functional consequences of this interaction on nuclear receptor and growth factor signaling pathways.
  • To assess the role of PNRC in cell growth and its expression in breast cancer.

Main Methods:

  • GST pull-down assay
  • Yeast two-hybrid assay
  • Coimmunoprecipitation
  • Cotransfection and fluorescence imaging
  • Transient transfection experiments
  • RT-PCR analysis of mRNA

Main Results:

  • PNRC physically interacts with Grb2, confirmed by multiple assays.
  • PNRC and Grb2 colocalize in mammalian cells.
  • Grb2 reduces PNRC's coactivator activity; PNRC suppresses Grb2-mediated Ras/MAP-kinase activation.
  • Overexpression of PNRC inhibits HeLa cell growth.
  • PNRC expression is significantly lower in breast cancer tissues compared to noncancerous tissues.

Conclusions:

  • PNRC and Grb2 interact to suppress both nuclear receptor-mediated and growth factor-mediated regulation in human breast tissue.
  • This interaction represents a novel crosstalk mechanism between these critical regulatory pathways.
  • PNRC may function as a tumor suppressor in breast cancer.

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