In vivo cross-linking followed by polyA enrichment to identify yeast mRNA binding proteins

Sarah F Mitchell; Roy Parker

doi:10.1007/978-1-4939-2214-7_3

In vivo cross-linking followed by polyA enrichment to identify yeast mRNA binding proteins

Methods Mol Biol. 2015:1259:35-47. doi: 10.1007/978-1-4939-2214-7_3.

Authors

Sarah F Mitchell¹, Roy Parker

Affiliation

¹ Department of Chemistry and Biochemistry, University of Colorado Boulder, JSCBB Room B450, 3415 Colorado Ave, UCB 596, Boulder, CO, 80303, USA, sarah.f.mitchell@colorado.edu.

PMID: 25579578
DOI: 10.1007/978-1-4939-2214-7_3

Abstract

mRNA binding proteins regulate gene expression by controlling the processing, localization, decay, and translation of messenger RNAs (mRNAs). To fully understand this process, it is necessary to identify the complete set of mRNA binding proteins. This work describes a method for the systematic identification of yeast mRNA binding proteins. This method applies in vivo UV cross-linking, affinity pull-down of polyA(+) mRNAs, and analysis by mass spectrometry to identify proteins that directly bind to mRNAs.

MeSH terms

Fungal Proteins / metabolism
Poly A / chemistry*
Protein Binding
RNA, Messenger / metabolism*
RNA-Binding Proteins / metabolism*

Substances

Fungal Proteins
RNA, Messenger
RNA-Binding Proteins
Poly A