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| Status |
Public on May 04, 2020 |
| Title |
RIC-seq for global in situ profiling of RNA-RNA spatial interactions |
| Organisms |
Drosophila melanogaster; Homo sapiens |
| Experiment type |
Other
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| Summary |
Highly structured RNA molecules usually interact with each other, and associate with various RNA-binding proteins, to regulate critical biological processes. However, RNA structures and interactions in intact cells remain largely unknown. Here, by coupling proximity ligation mediated by RNA-binding proteins with deep sequencing, we report an RNA in situ conformation sequencing (RIC-seq) technology for the global profiling of intra- and intermolecular RNA–RNA interactions. This technique not only recapitulates known RNA secondary structures and tertiary interactions, but also facilitates the generation of three-dimensional (3D) interaction maps of RNA in human cells. Using these maps, we identify noncoding RNA targets globally, and discern RNA topological domains and trans-interacting hubs. We reveal that the functional connectivity of enhancers and promoters can be assigned using their pairwise-interacting RNAs. Furthermore, we show that CCAT1-5L—a super-enhancer hub RNA—interacts with RNA-binding protein hnRNPK, as well as RNA derived from the MYC promoter and enhancer, to boost MYC transcription by modulating chromatin looping. Our study demonstrates the power and applicability of RIC-seq in discovering the 3D structures, interactions and regulatory roles of RNA.
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| Overall design |
We first developed an RNA in situ conformation sequencing technology, named RIC-seq, for global mapping RNA-RNA interactions in HeLa cells. Two biological replicates were generated for rRNA+ and rRNA- samples. To figure out the cell-to-cell random ligation rate, we mixed HeLa cells and Drosophila cells and then created a RIC-seq library for sequencing. As we found that super-enhancer lncRNA CCAT1-5L interacted with RNA-binding protein hnRNPK to regulate MYC transcription, so we further performed CLIP-seq to map hnRNPK binding sites globally. Both monomeric and dimeric hnRNPK-associated RNAs were recovered and converted into libraries for sequencing.
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| Contributor(s) |
Cai Z, Cao C, Ji L, Xue Y |
| Citation(s) |
32499643 |
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| Submission date |
Feb 26, 2019 |
| Last update date |
Aug 03, 2020 |
| Contact name |
Yuanchao Xue |
| E-mail(s) |
ycxue@ibp.ac.cn
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| Organization name |
Chinese Academy of Sciences
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| Department |
Institute of Biophysics
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| Lab |
Key Laboratory of RNA Biology
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| Street address |
Datun Road #15
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| City |
Beijing |
| ZIP/Postal code |
100101 |
| Country |
China |
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| Platforms (2) |
| GPL20795 |
HiSeq X Ten (Homo sapiens) |
| GPL26226 |
HiSeq X Ten (Drosophila melanogaster; Homo sapiens) |
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| Samples (18)
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| Relations |
| BioProject |
PRJNA524320 |
| SRA |
SRP186883 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE127188_RAW.tar |
1.6 Gb |
(http)(custom) |
TAR (of BW, HIC) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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