Ribo-proteomics approach to profile RNA–protein and protein–protein interaction networks

Characterizing protein–protein and protein–RNA interaction networks is a fundamental step to understanding the function of an RNA-binding protein. In many cases, these interactions are transient and highly dynamic. Therefore, capturing stable as well as transient interactions in living cells for the identification of protein-binding partners and the mapping of RNA-binding sequences is key to a successful establishment of the molecular interaction network. In this chapter, we will describe a method for capturing the molecular interactions in living cells using formaldehyde as a crosslinker and enriching a specific RNA–protein complex from cell extracts followed by mass spectrometry and Next-Gen sequencing analyses.