genomic binding sites of a noncoding RNA

Long noncoding RNAs (lncRNAs) have important regulatory roles and can function at the level of chromatin. To determine where lncRNAs bind to chromatin, we developed capture hybridization analysis of RNA targets (CHART), a hybridization-based technique that specifically enriches endogenous RNAs along...

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Published inProceedings of the National Academy of Sciences - PNAS Vol. 108; no. 51; pp. 20497 - 20502
Main Authors Simon, Matthew D, Wang, Charlotte I, Kharchenko, Peter V, West, Jason A, Chapman, Brad A, Alekseyenko, Artyom A, Borowsky, Mark L, Kuroda, Mitzi I, Kingston, Robert E
Format Journal Article
LanguageEnglish
Published United States National Academy of Sciences 20.12.2011
National Acad Sciences
Subjects
Amino Acid Motifs
Animals
Binding Sites
Biological Sciences
Charts
Chromatin
Chromatin - chemistry
Chromatin - genetics
Chromatin Immunoprecipitation
Compensation
crosslinking
DNA
Dosage compensation
Dosage Compensation, Genetic
Drosophila
Drosophila - genetics
Drosophila Proteins - genetics
Gene mapping
Genes
Genetic loci
Genomes
Genomics
humans
Hybridization
Hybridization analysis
Immunoprecipitation
Insects
Male
Models, Genetic
non-coding RNA
Nucleic Acid Hybridization
Peptide mapping
precipitin tests
Proteins
Ribonuclease H - chemistry
Ribonucleic acid
RNA
RNA, Untranslated - genetics
RNA-Binding Proteins - genetics
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Summary:Long noncoding RNAs (lncRNAs) have important regulatory roles and can function at the level of chromatin. To determine where lncRNAs bind to chromatin, we developed capture hybridization analysis of RNA targets (CHART), a hybridization-based technique that specifically enriches endogenous RNAs along with their targets from reversibly cross-linked chromatin extracts. CHART was used to enrich the DNA and protein targets of endogenous lncRNAs from flies and humans. This analysis was extended to genome-wide mapping of roX2, a well-studied ncRNA involved in dosage compensation in DROSOPHILA: CHART revealed that roX2 binds at specific genomic sites that coincide with the binding sites of proteins from the male-specific lethal complex that affects dosage compensation. These results reveal the genomic targets of roX2 and demonstrate how CHART can be used to study RNAs in a manner analogous to chromatin immunoprecipitation for proteins.
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Edited by* Keith R. Yamamoto, University of California, San Francisco, CA, and approved October 19, 2011 (received for review August 17, 2011)
Author contributions: M.D.S., C.I.W., J.A.W., A.A.A., M.I.K., and R.E.K. designed research; M.D.S., C.I.W., and J.A.W. performed research; M.D.S. and A.A.A. contributed new reagents/analytic tools; M.D.S., P.V.K., B.A.C., and M.L.B. analyzed data; and M.D.S., J.A.W., M.I.K., and R.E.K. wrote the paper.
ISSN:0027-8424
1091-6490
1091-6490
DOI:10.1073/pnas.1113536108