β-Actin mRNA interactome mapping by proximity biotinylation

Significance Transport of specific mRNAs to defined sites in the cytoplasm allows local protein production and contributes to cell polarity, embryogenesis, and neuronal function. These localized mRNAs contain signals (i.e., zipcodes) that help direct them to their destination site. Zipcodes are recognized by RNA-binding proteins that, with the help of molecular motor proteins and supplementary factors, mediate mRNA trafficking. To identify all proteins assembling with a localized mRNA, we advanced a proximity labeling method, BioID, by tethering a biotin ligase to the 3′ UTR of mRNA encoding the conserved β-actin protein. We demonstrate that this method allows the identification of functionally important proteins required for mRNA localization.