People involved: Alexander Kanitz, Alessia Galgano, Felix Schnarwiler (Gerber Lab)
- It is becoming increasingly clear that the microRNA system and classical post-transcriptional regulatory pathways interact on several levels. In addition to post-transcriptional modifications of the microRNA itself, mRNA binding of regulatory RNA-binding proteins (rRBPs) may influence the efficiency of microRNA-mediated knockdown. Recent studies on the interplay between specific miRNA:mRNA:rRBP pairs suggest the existence of different regulatory mechanisms, such as competition for cis-acting regulatory elements in the 3’ UTR. Moreover, we have previously found that miRNA seeds are enriched in the vicinity of the binding motifs of the rRBPs PUM1 and PUM2.
- To gain insight into the nature and extent of this mRNA-dependent crosstalk in human cells, we aim to determine the target sets of several rRBPs on a genome-wide level and compare them with those of the Argonaute proteins (huAGO1-4). In order to achieve this we will employ an improved RBP immunopurification protocol, followed by microarray analysis and/or deep sequencing (‘RIP-Chip’ / ‘RIP-Seq’). Furthermore, interaction networks of the RBPs will be determined by mass spectrometric analysis. Common targets will then be verified and subjected to in-depth bioinformatic and biochemical analysis.