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Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β

Rong Guo1, Kotb Abdelmohsen, Patrice J Morin

  • 1Laboratory of Genetics, National Institute on Aging-Intramural Research Program, NIH, Baltimore, Maryland, United States of America.

Plos One
|July 11, 2013
PubMed

Insights

Glycogen synthase kinase 3 beta (GSK-3β) inhibition increases let-7 microRNA levels, a tumor suppressor, in ovarian cancer cells. This suggests GSK-3β is a potential therapeutic target for ovarian tumorigenesis.

Area of Science:

  • Molecular Biology
  • Oncology
  • Biochemistry

Background:

  • let-7 microRNAs function as tumor suppressors by downregulating oncogenic proteins.
  • Reduced let-7 levels are observed in various cancers, including ovarian cancer.
  • A sensitive assay is needed to systematically measure cellular let-7 levels.

Purpose of the Study:

  • To develop and validate a novel let-7 reporter assay system.
  • To identify pathways regulating let-7 levels in ovarian cancer cells.
  • To evaluate glycogen synthase kinase 3 beta (GSK-3β) as a therapeutic target.

Main Methods:

  • Developed a pmirGLO-let7 reporter construct expressing chimeric mRNA with luciferase and let-7 binding sites.
  • Transfected ovarian cancer cell lines (BG-1, UCI-101) with the reporter system.
  • Screened kinase inhibitors and utilized RNA interference to identify regulators of let-7 activity.

Main Results:

  • The pmirGLO-let7 assay showed inverse correlation between luciferase activity and let-7 levels.
  • Inhibition or silencing of GSK-3β significantly increased both mature and primary let-7 levels in ovarian cancer cells.
  • GSK-3β inhibition led to decreased cell survival and implicated p53 in the regulation of let-7 biosynthesis.

Conclusions:

  • GSK-3β plays a critical role in repressing let-7 biosynthesis in ovarian cancer.
  • Targeting GSK-3β represents a promising therapeutic strategy for ovarian tumorigenesis.
  • The pmirGLO-let7 assay is a valuable tool for studying let-7 regulation.

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