Robust profiling of microRNAs and isomiRs in human plasma exosomes across 46 individuals

Abstract

microRNAs (miRNAs) are small double stranded RNA molecules consisting of two complementary strands called the 5p and 3p arms. Following imprecise processing and/or addition of nucleotides at the ends, miRNA biogenesis can give rise to variants called isomiRs. Exosomes are small vesicles released by cells. They have attracted attention due to their potential use in biomarker development because of their content of biomolecules, including miRNAs and isomiRs. Exosomes are found in body fluids such as plasma. In this study we used next generation sequencing to investigate the distribution of 5p and 3p arms of both miRNAs and isomiRs in plasma exosomes from 46 individuals. Among the canonical miRNAs there was similar prevalence between 5p and 3p miRNAs. Most of the miRNAs had isomiRs, and in approximately half of the cases an isomiR was more abundant than the corresponding canonical miRNA. Most of the isomiRs were generated from 5p miRNAs. There were very small differences in the concentration of canonical miRNA and isomiR sequences between donors, suggesting tight control of isomiR generation and sorting into exosomes. IsomiRs are abundant in plasma exosomes and should be included in analysis when plasma exosomal miRNAs are investigated as potential biomarkers for disease development.

Extracellular Vesicles Analysis in the COVID-19 Era: Insights on Serum Inactivation Protocols towards Downstream Isolation and Analysis

“…MiR-16-5p is among the most abundant miRNA in EVs [ 22 ], while miR-21-5p is reported to be associated withs lipoproteins and could be involved in lipid metabolism [ 23 ]. We quantitatively detected miRNAs levels within NT, HT and S/D groups by ddPCR analysis; results are summarized in Figure 6 .…”

isomiRs–Hidden Soldiers in the miRNA Regulatory Army, and How to Find Them?

“…In addition, EVs isolated from the urine of prostate cancer patients showed enrichment in miRNAs and isomiRs with potential for biomarkers in minimally invasive diagnosis of prostate cancer patients [ 126 ]. The most common feature among biofluid studies is that isomiRs have a higher abundance than known miRNAs [ 39 , 127 , 128 ]. This suggests miRNA precursors may generate more than one miRNA variants with specific properties.…”

Dynamic mRNP Remodeling in Response to Internal and External Stimuli

“…Nevertheless, there is ample evidence that beyond a technical artifact the prevalence of miRNA isoforms differs among cell types and tissues, where the miRNAs are expected to play different biological activities [ 217 , 218 ]. Studies comparing isomiRs indicate that isomiRs are more frequently seen in 5p than 3p miRNAs, suggesting a prevalent modulation at the step of Dicer processing [ 206 ] ( Figure 4 ). This is in agreement with reports that while both 5′ and 3′ end length variations can be seen in miRNAs, 3′ end modifications are the most frequent [ 207 , 208 ].…”

“…DICER and its binding partners TARBP and PACT subsequently process the stem loop of the pre-miRNAs with many RBPs selectively involved at this step depending on the pre-miRNA sequence [ 205 ]. Cleavage of the 5p arm by the resulting DICER complex will lead to 3′ end miRNA variations, which are the most frequent [ 206 , 207 , 208 ]. Nonetheless, 5′-end variations of the 3p arm can also occur.…”