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miRNA length variation during macrophage stimulation confounds the interpretation of results: implications for miRNA

Katherine A Pillman1,2,3, Gregory J Goodall1,3,4, Cameron P Bracken1,3,4

  • 1Centre for Cancer Biology, SA Pathology and University of South Australia, Adelaide, South Australia 5000, Australia.

RNA (New York, N.Y.)
|November 30, 2018
PubMed

Insights

Most microRNAs (miRNAs) exist as length variants called isomiRs. Macrophage stimulation alters isomiR balance, potentially misinterpreting results from standard RT-qPCR methods.

Area of Science:

  • Molecular Biology
  • Immunology
  • Genomics

Background:

  • MicroRNAs (miRNAs) are crucial regulators of gene expression.
  • Most miRNAs are expressed as a mixture of length isoforms, known as isomiRs.
  • IsomiR stoichiometry can change upon cellular stimulation, impacting miRNA function.

Purpose of the Study:

  • To investigate changes in isomiR stoichiometry in macrophages stimulated with bacterial products.
  • To assess the impact of isomiR variation on miRNA quantification using RT-qPCR.
  • To emphasize the importance of isoform-level analysis in miRNA expression studies.

Main Methods:

  • Re-analysis of published RNA-sequencing datasets from human and mouse macrophages.
  • Analysis of miRNA expression at the isomiR level.
  • Comparison of isomiR profiles before and after stimulation with bacterial products.

Main Results:

  • Macrophage stimulation with bacterial products induced significant changes in isomiR stoichiometry for several miRNAs.
  • Standard RT-qPCR methods may misinterpret miRNA expression changes due to unequal targeting of isomiRs by primers.
  • Isoform-level analysis revealed complexities in miRNA regulation not apparent with bulk quantification.

Conclusions:

  • IsomiR stoichiometry is dynamically regulated in macrophages upon immune stimulation.
  • Accurate assessment of miRNA expression requires considering isomiR profiles.
  • Analysis of miRNA expression at the isoform level should be adopted as a standard practice in research.
Keywords:
LPSRT-qPCR miRNA assaysbacterial infectioninterferonisomiRmacrophagesmicroRNA isoform

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