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Assessment of Selective mRNA Translation in Mammalian Cells by Polysome Profiling
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Assessment of Selective mRNA Translation in Mammalian Cells by Polysome Profiling

Published on: October 28, 2014

Cytochrome P450 1B1 mRNA untranslated regions interact to inhibit protein translation

Andrea H Devlin1, Paul Thompson, Tracy Robson

  • 1Biomedical Sciences Research Institute, University of Ulster, Coleraine, Northern Ireland BT52 1SA, UK.

Molecular Carcinogenesis
|November 13, 2009
PubMed

Insights

Tumor cells show high levels of CYP1B1 protein, unlike normal tissues. This study reveals that translational control, specifically involving the 5' untranslated region (UTR) and microRNA-27b, regulates CYP1B1 protein expression.

Area of Science:

  • Molecular Biology
  • Cancer Biology
  • Biochemistry

Background:

  • CYP1B1 protein is typically undetectable in normal human tissues but highly expressed in tumors.
  • CYP1B1 mRNA is constitutively expressed across normal extrahepatic tissues.
  • Tumor-specific protein expression suggests posttranscriptional regulation of CYP1B1.

Purpose of the Study:

  • To investigate the regulatory mechanisms behind tumor-specific CYP1B1 protein expression.
  • To analyze the role of the 5' untranslated region (UTR) in CYP1B1 mRNA translational control.
  • To explore the interaction between CYP1B1 mRNA and microRNA-27b in regulating protein levels.

Main Methods:

  • Measurement of CYP1B1 mRNA and protein expression in cell lines.
  • Bioinformatic analysis using ONCOMINE to compare tumor and normal tissue expression.
  • In vitro translation assays using a luciferase reporter gene with the CYP1B1 5'UTR.
  • Site-directed mutagenesis of the upstream open-reading frame (uORF) start codon.
  • Analysis of microRNA-27b interaction with the CYP1B1 3'UTR.

Main Results:

  • CYP1B1 mRNA is not significantly overexpressed in tumors, indicating posttranscriptional regulation.
  • The CYP1B1 5'UTR, containing a uORF, significantly inhibits translation.
  • Mutation of the uORF start codon abrogates the inhibitory effect of the 5'UTR.
  • The CYP1B1 5'UTR interacts with microRNA-27b in the 3'UTR, leading to near-complete translational inhibition.
  • CYP1B1 is under stringent translational control, explaining its absence in normal cells.

Conclusions:

  • CYP1B1 expression is primarily regulated at the translational level.
  • The 5'UTR and its uORF, along with microRNA-27b interactions, are key regulators of CYP1B1 translation.
  • Altered translational control during malignant transformation likely contributes to high CYP1B1 protein levels in tumors.

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