Quantitative identification of proteins that influence miRNA biogenesis by RNA pull-down-SILAC mass spectrometry (RP-SMS).

ABSTRACT: RNA-binding proteins mediate and control gene expression. As some examples, they regulate pre-mRNA synthesis and processing; mRNA localisation, translation and decay; and microRNA (miRNA) biogenesis and function. Here, we present a detailed protocol for RNA pull-down coupled to stable isotope labelling by amino acids in cell culture (SILAC) mass spectrometry (RP-SMS) that enables quantitative, fast and specific detection of RNA-binding proteins that regulate miRNA biogenesis. In general, this method allows for the identification of RNA-protein complexes formed using in vitro or chemically synthesized RNAs and protein extracts derived from cultured cells.

SUBMITTER: Choudhury NR 

PROVIDER: S-EPMC6335501 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature