Biology:HITS-CLIP

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High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP) is a variant of CLIP[1] for genome-wide mapping protein–RNA binding sites or RNA modification sites in vivo.[2][3][4] HITS-CLIP was originally used to generate genome-wide protein-RNA interaction maps for the neuron-specific RNA-binding protein and splicing factor NOVA1 and NOVA2;[3] since then a number of other splicing factor maps have been generated, including those for PTB,[5] RbFox2,[6] SFRS1,[7] hnRNP C,[8] and even N6-Methyladenosine (m6A) mRNA modifications.[4][9] HITS-CLIP of the RNA-binding protein Argonaute has been performed for the identification of microRNA targets[10] by decoding microRNA-mRNA and protein-RNA interaction maps in mouse brain,[11][12] and subsequently in Caenorhabditis elegans,[13] embryonic stem cells[14] and tissue culture cells.[15]

As a novel modification of HITS-CLIP, m6A-CLIP was developed to precisely map N6-Methyladenosine(m6A) locations in mRNA by UV-crosslinking m6A antibody to the target RNA.[4][9] Recently, improved bioinformatics applied to Argonaute HITS-CLIP enables identification of binding sites with single nucleotide resolution.[16]

Similar methods

  • PAR-CLIP, for identifying the binding sites of cellular RNA-binding proteins (RBPs) and microRNA-containing ribonucleoprotein complexes (miRNPs) in tissue culture cells.
  • iCLIP, for a thorough amplification of the cDNA library, including truncated cDNAs, thus also enabling an additional means to identify crosslink sites.

References

  1. ↑ Ule, Jernej; Jensen, Kirk B.; Ruggiu, Matteo; Mele, Aldo; Ule, Aljaz; Darnell, Robert B. (2003-11-14). "CLIP identifies Nova-regulated RNA networks in the brain". Science 302 (5648): 1212–1215. doi:10.1126/science.1090095. ISSN 1095-9203. PMID 14615540. Bibcode: 2003Sci...302.1212U. https://pubmed.ncbi.nlm.nih.gov/14615540. 
  2. ↑ Darnell RB (2010). "HITS-CLIP: panoramic views of protein-RNA regulation in living cells". Wiley Interdiscip Rev RNA 1 (2): 266–86. doi:10.1002/wrna.31. PMID 21935890. 
  3. ↑ 3.0 3.1 "HITS-CLIP yields genome-wide insights into brain alternative RNA processing". Nature 456 (7221): 464–9. November 2008. doi:10.1038/nature07488. PMID 18978773. Bibcode: 2008Natur.456..464L. 
  4. ↑ 4.0 4.1 4.2 Ke, S; Alemu, EA; Mertens, C; Gantman, EC; Fak, JJ; Mele, A; Haripal, B; Zucker-Scharff, I et al. (24 September 2015). "A majority of m6A residues are in the last exons, allowing the potential for 3′ UTR regulation.". Genes & Development 29 (19): 2037–53. doi:10.1101/gad.269415.115. PMID 26404942. 
  5. ↑ "Genome-wide analysis of PTB-RNA interactions reveals a strategy used by the general splicing repressor to modulate exon inclusion or skipping", Molecular Cell 36 (6): 996–1006, 2009, doi:10.1016/j.molcel.2009.12.003, PMID 20064465 
  6. ↑ "An RNA code for the FOX2 splicing regulator revealed by mapping RNA-protein interactions in stem cells". Nat Struct Mol Biol 16 (2): 130–137. 2009. doi:10.1038/nsmb.1545. PMID 19136955. 
  7. ↑ "Splicing factor SFRS1 recognizes a functionally diverse landscape of RNA transcripts". Genome Research 19 (3): 381–394. 2009. doi:10.1101/gr.082503.108. PMID 19116412. 
  8. ↑ "iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution", Nat Struct Mol Biol 17 (7): 909–915, 2010, doi:10.1038/nsmb.1838, PMID 20601959 
  9. ↑ 9.0 9.1 Ke, S; Pandya-Jones, A; Saito, Y; Fak, JJ; Vågbø, CB; Geula, S; Hanna, JH; Black, DL et al. (15 May 2017). "m6A mRNA modifications are deposited in nascent pre-mRNA and are not required for splicing but do specify cytoplasmic turnover.". Genes & Development 31 (10): 990–1006. doi:10.1101/gad.301036.117. PMID 28637692. 
  10. ↑ Thomson, DW; Bracken, CP; Goodall, GJ (2011-06-07). "Experimental strategies for microRNA target identification.". Nucleic Acids Research 39 (16): 6845–6853. doi:10.1093/nar/gkr330. PMID 21652644. 
  11. ↑ Chi, S.W.; Zang, J.B.; Mele, A.; Darnell,R.B. (2009), "Argonaute HITS-CLIP decodes microRNA-mRNA interaction maps", Nature 460 (7254): 479–486, doi:10.1038/nature08170, PMID 19536157, Bibcode: 2009Natur.460..479C 
  12. ↑ "starBase: a database for exploring microRNA–mRNA interaction maps from Argonaute CLIP-Seq and Degradome-Seq data.". Nucleic Acids Res. 39 (Database issue): D202–D209. 2011. doi:10.1093/nar/gkq1056. PMID 21037263. 
  13. ↑ "Comprehensive discovery of endogenous Argonaute binding sites in Caenorhabditis elegans", Nat Struct Mol Biol 17 (2): 173–179, 2010, doi:10.1038/nsmb.1745, PMID 20062054 
  14. ↑ "Genome-wide identification of Ago2 binding sites from mouse embryonic stem cells with and without mature microRNAs", Nat Struct Mol Biol 19 (9): 1084, 2011, doi:10.1038/nsmb0911-1084a 
  15. ↑ "Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP", Cell 141 (1): 129–141, 2010, doi:10.1016/j.cell.2010.03.009, PMID 20371350 
  16. ↑ Zhang, C.; Darnell,R.B. (2011). "Mapping in vivo protein-RNA interactions at single-nucleotide resolution from HITS-CLIP data.". Nature Biotechnology 29 (7): 607–614. doi:10.1038/nbt.1873. PMID 21633356. 

External links

  • CLIPSim-MC: CLIPSim-MC is a tool that uses CLIP-seq data to find miRNA/MRE pairings using a Monte-Carlo-based approach.[1]
  • starBase database: a database for exploring miRNA-mRNA, miRNA-lncRNA, miRNA-sncRNA, miRNA-circRNA, miRNA-pseudogene, protein-lncRNA, protein-RNA interactions and ceRNA networks from HITS-CLIP (CLIP-Seq, PAR-CLIP, iCLIP, CLASH) data, and TargetScan[2], PicTar, RNA22, miRanda and PITA microRNA target sites.
  • clipz: a pipeline to analyze short RNA reads from HITS-CLIP experiments.
  • dCLIP: dCLIP is a Perl program for discovering differential binding regions in two comparative CLIP-Seq (HITS-CLIP, PAR-CLIP or iCLIP) experiments.



  1. ↑ Peter M. Clark; Phillipe Loher; Kevin Quann; Jonathan Brody; Eric R. Londin; Isidore Rigoutsos (2014), "Argonaute CLIP-Seq reveals miRNA targetome diversity across tissue types", Scientific Reports 4 (5947): 5947, doi:10.1038/srep05947, PMID 25103560, Bibcode: 2014NatSR...4E5947C 
  2. ↑ Agarwal, Vikram; Bell, George W.; Nam, Jin-Wu; Bartel, David P. (2015-08-12). "Predicting effective microRNA target sites in mammalian mRNAs" (in en). eLife 4: e05005. doi:10.7554/eLife.05005. ISSN 2050-084X. PMID 26267216. 
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