HITS-CLIP yields genome-wide insights into brain alternative RNA processing

Donny D. Licatalosi, Aldo Mele, John J. Fak, Jernej Ule, Melis Kayikci, Sung Wook Chi, Tyson A. Clark, Anthony C. Schweitzer, John E. Blume, Xuning Wang, Jennifer C. Darnell and Robert B. Darnell ()
Additional contact information
Donny D. Licatalosi: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,
Aldo Mele: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,
John J. Fak: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,
Jernej Ule: MRC Laboratory of Molecular Biology, Cambridge, CB2 0QH, UK
Melis Kayikci: MRC Laboratory of Molecular Biology, Cambridge, CB2 0QH, UK
Sung Wook Chi: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,
Tyson A. Clark: Expression Research, Affymetrix, Inc., Santa Clara, California 95051, USA
Anthony C. Schweitzer: Expression Research, Affymetrix, Inc., Santa Clara, California 95051, USA
John E. Blume: Expression Research, Affymetrix, Inc., Santa Clara, California 95051, USA
Xuning Wang: Biocomputing, Information Technology, The Rockefeller University, 1230 York Avenue, New York, New York 10021, USA
Jennifer C. Darnell: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,
Robert B. Darnell: Laboratory of Molecular Neuro-Oncology and Howard Hughes Medical Institute,

Nature, 2008, vol. 456, issue 7221, 464-469

Abstract: Abstract Protein–RNA interactions have critical roles in all aspects of gene expression. However, applying biochemical methods to understand such interactions in living tissues has been challenging. Here we develop a genome-wide means of mapping protein–RNA binding sites in vivo, by high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP). HITS-CLIP analysis of the neuron-specific splicing factor Nova revealed extremely reproducible RNA-binding maps in multiple mouse brains. These maps provide genome-wide in vivo biochemical footprints confirming the previous prediction that the position of Nova binding determines the outcome of alternative splicing; moreover, they are sufficiently powerful to predict Nova action de novo. HITS-CLIP revealed a large number of Nova–RNA interactions in 3′ untranslated regions, leading to the discovery that Nova regulates alternative polyadenylation in the brain. HITS-CLIP, therefore, provides a robust, unbiased means to identify functional protein–RNA interactions in vivo.

Date: 2008
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DOI: 10.1038/nature07488

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