title: Ribo-Proteomics Approach to Profile RNA-Protein and Protein-Protein Interaction Networks.
abstract: 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.
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota Twin Cities, 6-155 Jackson Hall, 321 Church Street SE, Minneapolis, MN, 55455, USA.
authors.forename: Hsin-Sung
authors.initials: HS
authors.lastname: Yeh
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota Twin Cities, 6-155 Jackson Hall, 321 Church Street SE, Minneapolis, MN, 55455, USA.
authors.forename: Jae-Woong
authors.initials: JW
authors.lastname: Chang
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota Twin Cities, 6-155 Jackson Hall, 321 Church Street SE, Minneapolis, MN, 55455, USA. jyong@umn.edu.
authors.forename: Jeongsik
authors.initials: J
authors.lastname: Yong
mesh_terms: D000818:Animals; D003432:Cross-Linking Reagents / Q000737:chemistry; D005557:Formaldehyde / Q000737:chemistry; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D060066:Protein Interaction Maps; D040901:Proteomics / Q000379:methods*; D012313:RNA / Q000032:analysis / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000032:analysis / Q000378:metabolism*
chemical_list: D003432:Cross-Linking Reagents; D016601:RNA-Binding Proteins; D005557:Formaldehyde; D012313:RNA
keywords: Crosslinking immunoprecipitation; Formaldehyde crosslinking; Mass spectrometry; Next-Gen sequencing; RNA-binding proteins