Beta Cell 5′-Shifted isomiRs Are Candidate Regulatory Hubs in Type 2 DiabetesReportar como inadecuado

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Next-generation deep sequencing of small RNAs has unveiled the complexity of the microRNA miRNA transcriptome, which is in large part due to the diversity of miRNA sequence variants -isomiRs-. Changes to a miRNA’s seed sequence nucleotides 2–8, including shifted start positions, can redirect targeting to a dramatically different set of RNAs and alter biological function. We performed deep sequencing of small RNA from mouse insulinoma MIN6 cells widely used as a surrogate for the study of pancreatic beta cells and developed a bioinformatic analysis pipeline to profile isomiR diversity. Additionally, we applied the pipeline to recently published small RNA-seq data from primary human beta cells and whole islets and compared the miRNA profiles with that of MIN6. We found that: 1 the miRNA expression profile in MIN6 cells is highly correlated with those of primary human beta cells and whole islets; 2 miRNA loci can generate multiple highly expressed isomiRs with different 5′-start positions 5′-isomiRs; 3 isomiRs with shifted start positions 5′-shifted isomiRs are highly expressed, and can be as abundant as their unshifted counterparts 5′-reference miRNAs. Finally, we identified 10 beta cell miRNA families as candidate regulatory hubs in a type 2 diabetes T2D gene network. The most significant candidate hub was miR-29, which we demonstrated regulates the mRNA levels of several genes critical to beta cell function and implicated in T2D. Three of the candidate miRNA hubs were novel 5′-shifted isomiRs: miR-375+1, miR-375-1 and miR-183-5p+1. We showed by in silico target prediction and in vitro transfection studies that both miR-375+1 and miR-375-1 are likely to target an overlapping, but distinct suite of beta cell genes compared to canonical miR-375. In summary, this study characterizes the isomiR profile in beta cells for the first time, and also highlights the potential functional relevance of 5′-shifted isomiRs to T2D.

Autor: Jeanette Baran-Gale, Emily E. Fannin, C. Lisa Kurtz, Praveen Sethupathy



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