Systematic evaluation of the microRNAome through miR-CATCHv2.0 identifies positive and negative regulators of BRAF-X1 mRNA.

ABSTRACT: Here we present miR-CATCHv2.0, an implemented experimental method that allows the identification of the microRNA species directly bound to an RNA of interest. After cross-linking of microRNA::RNA::Ago2 complexes using formaldehyde, the RNA is fragmented using sonication and then subjected to affinity purification using two sets of biotinylated tiling probes (ODD and EVEN). Finally, enriched microRNA species are retrieved by means of small RNA sequencing coupled with an ad hoc analytical workflow. In BRAFV600E mutant A375 melanoma cells, miR-CATCHv2.0 allowed us to identify 20 microRNAs that target X1, the most abundant isoform of BRAF mRNA. These microRNAs fall into different functional classes, according to the effect that they exert (decrease/increase in BRAFV600E mRNA and protein levels

SUBMITTER: Marranci A 

PROVIDER: S-EPMC6546394 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature