MicroRNAs and their isomiRs function cooperatively to target common biological pathways

Abstract

BackgroundVariants of microRNAs (miRNAs), called isomiRs, are commonly reported in deep-sequencing studies; however, the functional significance of these variants remains controversial. Observational studies show that isomiR patterns are non-random, hinting that these molecules could be regulated and therefore functional, although no conclusive biological role has been demonstrated for these molecules.ResultsTo assess the biological relevance of isomiRs, we have performed ultra-deep miRNA-seq on ten adult human tissues, and created an analysis pipeline called miRNA-MATE to align, annotate, and analyze miRNAs and their isomiRs. We find that isomiRs share sequence and expression characteristics with canonical miRNAs, and are generally strongly correlated with canonical miRNA expression. A large proportion of isomiRs potentially derive from AGO2 cleavage independent of Dicer. We isolated polyribosome-associated mRNA, captured the mRNA-bound miRNAs, and found that isomiRs and canonical miRNAs are equally associated with translational machinery. Finally, we transfected cells with biotinylated RNA duplexes encoding isomiRs or their canonical counterparts and directly assayed their mRNA targets. These studies allow us to experimentally determine genome-wide mRNA targets, and these experiments showed substantial overlap in functional mRNA networks suppressed by both canonical miRNAs and their isomiRs.ConclusionsTogether, these results find isomiRs to be biologically relevant and functionally cooperative partners of canonical miRNAs that act coordinately to target pathways of functionally related genes. This work exposes the complexity of the miRNA-transcriptome, and helps explain a major miRNA paradox: how specific regulation of biological processes can occur when the specificity of miRNA targeting is mediated by only 6 to 11 nucleotides.

Bioinformatic analysis of endogenous and exogenous small RNAs on lipoproteins

“…Due to imprecise cleavage of miRNAs from precursor miRNA hairpins [23–25], one miRNA locus can produce multiple isoforms, termed isomiRs, which can differ by one or two nts at the 5ʹ start position. Consequently, the canonical miRNA “seed” sequence is altered, potentially conferring recognition of different mRNA targets [24–26].…”

MECHANISMS IN ENDOCRINOLOGY: MicroRNA in diagnostics and therapy of thyroid cancer

“…This fact is of profound importance for miRNA's function, because the addition or deletion of nucleotides at the isomiR 5 0 end results in a change in the seed region and, consequently, leads to the regulation of a distinct set of target genes compared to the isomiR's canonical counterpart. As functional partners of their reference canonical molecules, isomiRs are tissue specific; therefore, the proper interpretation of microRNA-regulated networks requires comprehensive information on the miRNome for each particular tissue type (15,16,17).…”

miRNAs in cardiac disease: Sitting duck or moving target?

“…It is implausible that the whole phenomenon is simply a manifestation of noise in the system, although this may well be the case for individual examples. Expression of miRNA variants has been hypothesized to facilitate robust repression of a core set of mRNAs they target collectively, whereas potential individual off-targeting effects of each variant are ''diluted out'' by its relatively lower abundance (32). Another possibility is that miRNA sequence variants may be differentially localized and regulated or otherwise may associate with diverse effector proteins to create distinct outcomes (e.g., translational repression vs. mRNA decay).…”

“…start sites are generated by differential Drosha or Dicer processing and have shifted seed sequences relative to the canonical miRNA. 5 0 isomiRs may thus have differential targeting properties and experiments indeed showed that they bind both common and unique targets (32). Additionally, there is experimental evidence that 5 0 isomiRs of the cardiac miR-133a can have selective mRNA repression properties (34).…”

“…The proportion of isomiRs can vary between miRNAs and between cellular contexts for a given miRNA. Importantly, they have been detected in Argonaute complexes and in translating ribosomes, implying that they are functional miRNAs (23,32,33). start sites are generated by differential Drosha or Dicer processing and have shifted seed sequences relative to the canonical miRNA.…”