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]
- 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.
Licatalosi DD, Mele A, Fak JJ, Ule J, Kayikci M, Chi SW, Clark TA, Schweitzer AC, Blume JE, Wang X, Darnell JC, Darnell RB (November 2008). "HITS-CLIP yields genome-wide insights into brain alternative RNA processing". Nature. 456 (7221): 464–9. Bibcode:2008Natur.456..464L. doi:10.1038/nature07488. PMC 2597294. PMID 18978773. Ke, S; Alemu, EA; Mertens, C; Gantman, EC; Fak, JJ; Mele, A; Haripal, B; Zucker-Scharff, I; Moore, MJ; Park, CY; Vågbø, CB; Kusnierczyk, A; Klungland, A; Darnell, JE; Darnell, RB (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. PMC 4604345. PMID 26404942. Xue Y, Zhou Y, Wu T, Zhu T, Ju X, Kwon YS, Zhang C, Yeo G, Black DL, Sun H, Fu XD, Zhang Y (2009), "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, doi:10.1016/j.molcel.2009.12.003, PMC 2807993, PMID 20064465 Konig J, Zarnack K, Rot G, Curk T, Kayikci M, Zupan B, Turner DJ, Luscombe NM, Ule J (2010), "iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution", Nat Struct Mol Biol, 17 (7): 909–915, doi:10.1038/nsmb.1838, PMC 3000544, PMID 20601959 Ke, S; Pandya-Jones, A; Saito, Y; Fak, JJ; Vågbø, CB; Geula, S; Hanna, JH; Black, DL; Darnell, JE; Darnell, RB (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. PMC 5495127. PMID 28637692. 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, Bibcode:2009Natur.460..479C, doi:10.1038/nature08170, PMC 2733940, PMID 19536157 Zisoulis DG, Lovci MT, Wilbert ML, Hutt KR, Liang TY, Pasquinelli AE, Yeo GW (2010), "Comprehensive discovery of endogenous Argonaute binding sites in Caenorhabditis elegans", Nat Struct Mol Biol, 17 (2): 173–179, doi:10.1038/nsmb.1745, PMC 2834287, PMID 20062054 Leung AK, Young AG, Bhutkar A, Zheng GX, Bosson AD, Nielsen CB, Sharp PA (2011), "Genome-wide identification of Ago2 binding sites from mouse embryonic stem cells with and without mature microRNAs", Nat Struct Mol Biol, 19 (9): 1084, doi:10.1038/nsmb0911-1084a, PMC 3078052 Hafner M, Landthaler M, Burger L, Khorshid M, Hausser J, Berninger P, Rothballer A, Ascano M Jr, Jungkamp AC, Munschauer M, Ulrich A, Wardle GS, Dewell S, Zavolan M, Tuschl T (2010), "Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP", Cell, 141 (1): 129–141, doi:10.1016/j.cell.2010.03.009, PMC 2861495, PMID 20371350
- 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.