1. The mRNA-Bound Proteome and Its Global Occupancy Profile on Protein-Coding Transcripts
- Author
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Kevin Drew, Noah Youngs, Markus Schueler, Markus Landthaler, Alexandra Vasile, Matthias Selbach, Mathias Munschauer, Emanuel Wyler, Alexander G. Baltz, Björn Schwanhäusser, Christoph Dieterich, Duncan Penfold-Brown, Miha Milek, Richard Bonneau, and Yasuhiro Murakawa
- Subjects
Genetics ,Proteomics ,Messenger RNA ,Binding Sites ,Sequence analysis ,Sequence Analysis, RNA ,Quantitative proteomics ,RNA ,RNA-Binding Proteins ,Computational biology ,Cell Biology ,Biology ,Mass Spectrometry ,Cell Line ,RNA splicing ,Proteome ,Humans ,RNA, Messenger ,Binding site ,ICLIP ,Molecular Biology - Abstract
Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation, and translation. We developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing. Application to a human embryonic kidney cell line identified close to 800 proteins. To our knowledge, nearly one-third were not previously annotated as RNA binding, and about 15% were not predictable by computational methods to interact with RNA. Protein occupancy profiling provides a transcriptome-wide catalog of potential cis-regulatory regions on mammalian mRNAs and showed that large stretches in 3' UTRs can be contacted by the mRNA-bound proteome, with numerous putative binding sites in regions harboring disease-associated nucleotide polymorphisms. Our observations indicate the presence of a large number of mRNA binders with diverse molecular functions participating in combinatorial posttranscriptional gene-expression networks.
- Published
- 2012
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