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Posttranscriptional regulation of gene expression shapes the B cell transcriptome and controls messenger RNA (mRNA) translation into protein. Recent reports have highlighted the importance of RNA binding proteins (RBPs) for mRNA splicing, subcellular location, stability, and translation during B lymphocyte development, activation, and differentiation. Here we describe individual-nucleotide resolution UV cross-linking and immunoprecipitation (iCLIP) in primary lymphocytes, a method that maps RNA-protein interactions in a genome-wide scale allowing mechanistic analysis of RBP function. We discuss the latest improvements in iCLIP technology and provide some examples of how integration of the RNA-protein interactome with other high-throughput mRNA sequencing methodologies uncovers the important role of RBP-mediated RNA regulation in key biological cell processes.

Original publication

DOI

10.1007/978-1-4939-7095-7_14

Type

Journal article

Journal

Methods Mol Biol

Publication Date

2017

Volume

1623

Pages

159 - 179

Keywords

3′ Untranslated regions (UTRs), Individual-nucleotide resolution UV cross-linking and immunoprecipitation (iCLIP), Intron, Primary B cells, RNA binding proteins, mRNA maturation, mRNA stability, mRNA translation, Animals, B-Lymphocytes, Binding Sites, Cell Separation, Computational Biology, Gene Expression Profiling, Gene Library, High-Throughput Nucleotide Sequencing, Humans, Immunoprecipitation, Introns, Lymphocyte Activation, Protein Binding, RNA Stability, RNA-Binding Proteins, Transcriptome, Ultraviolet Rays