Identification of microRNA Precursor-Associated Proteins.

MicroRNA (miRNA) biogenesis is regulated intricately at multiple levels. In addition to transcriptional control of pri-miRNA loci, sequence as well as structural features of the pri-miRNA-stem loop determine its processing efficiency by the endonucleases Drosha and Dicer. On the one hand, general features are necessary to allow a hairpin to be recognized by the processing machinery; on the other hand, specific sequence motifs of individual miRNA precursors can be read by RNA binding proteins (RBPs) that regulate processing, leading to increased or decreased levels of functional miRNAs. In a pulldown experiment using the pri-miRNA hairpin as immobilized bait, cognate RBPs can be isolated and analyzed by immunoblotting or mass spectrometry, allowing for the discovery or analysis of protein regulators of miRNA biogenesis.