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Supplementary site interactions are critical for the regulation of microsomal triglyceride transfer protein by

James Soh1, M Mahmood Hussain

  • 1Department of Cell Biology, SUNY Downstate Medical Center, Brooklyn, NY 11203, USA. mahmood.hussain@downstate.edu.

Nutrition & Metabolism
|September 7, 2013
PubMed

Insights

MicroRNA-30c specifically regulates Microsomal triglyceride transfer protein (MTTP) expression through supplementary binding sites, not just seed sequences. Enhancing base pairing, particularly at supplementary sites, boosts microRNA efficacy.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Microsomal triglyceride transfer protein (MTTP) is crucial for assembling apolipoprotein B-lipoproteins, essential for lipid transport.
  • MicroRNA (miR)-30c uniquely regulates MTTP expression, despite sharing a seed sequence with other miR-30 family members.
  • Previous studies identified the miR-30c seed sequence and MTTP mRNA 3'-untranslated region (UTR) interaction as critical for regulation.

Purpose of the Study:

  • To investigate the role of a supplementary binding site and intervening loop in miR-30c/MTTP mRNA interactions.
  • To elucidate the mechanism behind miR-30c's specific regulation of MTTP expression.
  • To evaluate the impact of varying base pairing at the supplementary site and loop on miR-30c efficacy.

Main Methods:

  • Cloning the MTTP 3'-UTR downstream of the luciferase gene.
  • Utilizing site-directed mutagenesis to alter the supplementary binding site and intervening loop.
  • Measuring luciferase activity to assess the functional impact of miR-30c interactions.

Main Results:

  • Reducing base pairs at the supplementary site abolished miR-30c's ability to decrease luciferase activity.
  • Increasing base pairs at the supplementary site, seed sequence, or intervening loop enhanced miR-30c's regulatory efficacy.
  • The supplementary site is critical for miR-30c binding to MTTP mRNA, while the intervening loop size is not.

Conclusions:

  • This study demonstrates that microRNAs may require both seed and supplementary interactions for target mRNA specificity.
  • The supplementary binding site is essential for miR-30c's specific regulation of MTTP mRNA.
  • Optimizing base pairing in the loop region could lead to the synthesis of more potent miR-30c mimics.

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