Lebedeva, S. | Search bioRxiv

bioRxiv2018

Global identification of functional microRNA::mRNA interactions in Drosophila

MicroRNAs (miRNAs) are key mediators of post-transcriptional gene expression silencing. Although Drosophila has been of critical importance for miRNA discovery, biogenesis and function, there has been no comprehensive experimental annotation of functional miRNA target sites. To close this gap, we generated the first in vivo map of miRNA::mRNA interactions in Drosophila melanogaster, making use of crosslinked nucleotides in Argonaute (AGO) crosslinking and immunoprecipitation (CLIP) experiments that enable an unambiguous assignment of miRNAs to AGO binding sites at much higher signal-to-noise ratio than computational predictions alone.\n\nAbsolute quantification of cellular miRNA levels showed the miRNA pool in Drosophila cell lines to be more diverse than previously reported. Benchmarking two different CLIP approaches, we identified a similar predictive potential to unambiguously assign thousands of miRNA::mRNA pairs from AGO1 interaction data at unprecedented depth. Quantitative RNA-Seq and subcodon-resolution ribosomal footprinting data upon AGO1 depletion enabled the determination of miRNA-mediated effects on target expression and translation. We thus provide the first comprehensive resource of miRNA target sites as well as their quantitative functional impact in Drosophila.

systems biology↗

bioRxiv2018

Deciphering human ribonucleoprotein regulatory networks

RNA-binding proteins (RBPs) control and coordinate each stage in the life cycle of RNAs. Although in vivo binding sites of RBPs can now be determined genome-wide, most studies typically focused on individual RBPs. Here, we examined a large compendium of 114 high-quality transcriptome-wide in vivo RBP-RNA cross-linking interaction datasets generated by the same protocol in the same cell line and representing 64 distinct RBPs. Comparative analysis of categories of target RNA binding preference, sequence preference, and transcript region specificity was performed, and identified potential posttranscriptional regulatory modules, i.e. specific combinations of RBPs that bind to specific sets of RNAs and targeted regions. These regulatory modules encoded functionally related proteins and exhibited distinct differences in RNA metabolism, expression variance, as well as subcellular localization. This integrative investigation of experimental RBP-RNA interaction evidence and RBP regulatory function in a human cell line will be a valuable resource for understanding the complexity of post-transcriptional regulation.

genomics↗