Skip to main content
SpringerLink
Log in

PARIS: Psoralen Analysis of RNA Interactions and Structures with High Throughput and Resolution

  • Protocol
RNA Detection

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1649))

Abstract

RNA has the intrinsic propensity to form base pairs, leading to complex intramolecular and intermolecular helices. Direct measurement of base pairing interactions in living cells is critical to solving transcriptome structure and interactions, and investigating their functions (Lu and Chang, Curr Opin Struct Biol 36:142–148, 2016). Toward this goal, we developed an experimental method, PARIS (Psoralen Analysis of RNA Interactions and Structures), to directly determine transcriptome-wide base pairing interactions (Lu et al., Cell 165(5):1267–1279, 2016). PARIS combines four critical steps, in vivo cross-linking, 2D gel purification, proximity ligation, and high-throughput sequencing to achieve high-throughput and near-base pair resolution determination of the RNA structurome and interactome in living cells. In this chapter, we aim to provide a comprehensive discussion on the principles behind the experimental and computational strategies, and a step-by-step description of the experiment and analysis.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 139.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 179.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 179.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Cech TR, Steitz JA (2014) The noncoding RNA revolution-trashing old rules to forge new ones. Cell 157(1):77–94. doi:10.1016/j.cell.2014.03.008

    Article  CAS  PubMed  Google Scholar 

  2. Mattick JS, Makunin IV (2006) Non-coding RNA. Human molecular genetics 15 Spec No 1:R17-29. doi:10.1093/hmg/ddl046

  3. Rinn JL, Chang HY (2012) Genome regulation by long noncoding RNAs. Annu Rev Biochem 81:145–166. doi:10.1146/annurev-biochem-051410-092902

    Article  CAS  PubMed  Google Scholar 

  4. Wang KC, Chang HY (2011) Molecular mechanisms of long noncoding RNAs. Mol Cell 43(6):904–914. doi:10.1016/j.molcel.2011.08.018

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Lu Z, Matera AG (2014) Developmental analysis of spliceosomal snRNA isoform expression. G3 5(1):103–110. doi:10.1534/g3.114.015735

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. O’Reilly D, Dienstbier M, Cowley SA, Vazquez P, Drozdz M, Taylor S, James WS, Murphy S (2013) Differentially expressed, variant U1 snRNAs regulate gene expression in human cells. Genome Res 23(2):281–291. doi:10.1101/gr.142968.112

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Jia Y, JC M, Ackerman SL (2012) Mutation of a U2 snRNA gene causes global disruption of alternative splicing and neurodegeneration. Cell 148(1–2):296–308. doi:10.1016/j.cell.2011.11.057

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Lu Z, Guan X, Schmidt CA, Matera AG (2014) RIP-seq analysis of eukaryotic Sm proteins identifies three major categories of Sm-containing ribonucleoproteins. Genome Biol 15(1):R7. doi:10.1186/gb-2014-15-1-r7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Kim SH, Quigley GJ, Suddath FL, McPherson A, Sneden D, Kim JJ, Weinzierl J, Rich A (1973) Three-dimensional structure of yeast phenylalanine transfer RNA: folding of the polynucleotide chain. Science 179(4070):285–288

    Article  CAS  Google Scholar 

  10. Robertus JD, Ladner JE, Finch JT, Rhodes D, Brown RS, Clark BF, Klug A (1974) Structure of yeast phenylalanine tRNA at 3 A resolution. Nature 250(467):546–551

    Article  CAS  Google Scholar 

  11. Rouskin S, Zubradt M, Washietl S, Kellis M, Weissman JS (2014) Genome-wide probing of RNA structure reveals active unfolding of mRNA structures in vivo. Nature 505(7485):701–705. doi:10.1038/nature12894

    Article  CAS  PubMed  Google Scholar 

  12. Ding Y, Tang Y, Kwok CK, Zhang Y, Bevilacqua PC, Assmann SM (2014) In vivo genome-wide profiling of RNA secondary structure reveals novel regulatory features. Nature 505(7485):696–700. doi:10.1038/nature12756

    Article  CAS  PubMed  Google Scholar 

  13. Spitale RC, Flynn RA, Zhang QC, Crisalli P, Lee B, Jung JW, Kuchelmeister HY, Batista PJ, Torre EA, Kool ET, Chang HY (2015) Structural imprints in vivo decode RNA regulatory mechanisms. Nature 519(7544):486–490. doi:10.1038/nature14263

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Lu Z, Chang HY (2016) Decoding the RNA structurome. Curr Opin Struct Biol 36:142–148. doi:10.1016/j.sbi.2016.01.007

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Cimino GD, Gamper HB, Isaacs ST, Hearst JE (1985) Psoralens as photoactive probes of nucleic acid structure and function: organic chemistry, photochemistry, and biochemistry. Annu Rev Biochem 54:1151–1193. doi:10.1146/annurev.bi.54.070185.005443

    Article  CAS  PubMed  Google Scholar 

  16. Thompson JF, Hearst JE (1983) Structure of E. coli 16S RNA elucidated by psoralen crosslinking. Cell 32(4):1355–1365

    Article  CAS  Google Scholar 

  17. Ross A, Brimacombe R (1979) Experimental determination of interacting sequences in ribosomal RNA. Nature 281(5729):271–276

    Article  CAS  Google Scholar 

  18. Calvet JP, Pederson T (1979) Heterogeneous nuclear RNA double-stranded regions probed in living HeLa cells by crosslinking with the psoralen derivative aminomethyltrioxsalen. Proc Natl Acad Sci U S A 76(2):755–759

    Article  CAS  Google Scholar 

  19. Calvet JP, Pederson T (1981) Base-pairing interactions between small nuclear RNAs and nuclear RNA precursors as revealed by psoralen cross-linking in vivo. Cell 26(3 Pt 1):363–370

    Article  CAS  Google Scholar 

  20. Hausner TP, Giglio LM, Weiner AM (1990) Evidence for base-pairing between mammalian U2 and U6 small nuclear ribonucleoprotein particles. Genes Dev 4(12A):2146–2156

    Article  CAS  Google Scholar 

  21. Lu Z, Zhang QC, Lee B, Flynn RA, Smith MA, Robinson JT, Davidovich C, Gooding AR, Goodrich KJ, Mattick JS, Mesirov JP, Cech TR, Chang HY (2016) RNA duplex map in living cells reveals higher-order transcriptome structure. Cell 165(5):1267–1279. doi:10.1016/j.cell.2016.04.028

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30(15):2114–2120. doi:10.1093/bioinformatics/btu170

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Dobin A, Davis CA, Schlesinger F, Drenkow J, Zaleski C, Jha S, Batut P, Chaisson M, Gingeras TR (2013) STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29(1):15–21. doi:10.1093/bioinformatics/bts635

    Article  CAS  PubMed  Google Scholar 

  24. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R, Genome Project Data Processing S (2009) The Sequence Alignment/Map format and SAMtools. Bioinformatics 25(16):2078–2079. doi:10.1093/bioinformatics/btp352

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Zuker M (2003) Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 31(13):3406–3415

    Article  CAS  Google Scholar 

  26. Robinson JT, Thorvaldsdottir H, Winckler W, Guttman M, Lander ES, Getz G, Mesirov JP (2011) Integrative genomics viewer. Nat Biotechnol 29(1):24–26. doi:10.1038/nbt.1754

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Lorenz R, Bernhart SH, Honer Z, Siederdissen C, Tafer H, Flamm C, Stadler PF, Hofacker IL (2011) ViennaRNA Package 2.0. Algorithms Mol Biol 6:26. doi:10.1186/1748-7188-6-26

    Article  PubMed  PubMed Central  Google Scholar 

  28. Gesell T, von Haeseler A (2006) In silico sequence evolution with site-specific interactions along phylogenetic trees. Bioinformatics 22(6):716–722. doi:10.1093/bioinformatics/bti812

    Article  CAS  PubMed  Google Scholar 

  29. Flynn RA, Zhang QC, Spitale RC, Lee B, Mumbach MR, Chang HY (2016) Transcriptome-wide interrogation of RNA secondary structure in living cells with icSHAPE. Nat Protoc 11(2):273–290. doi:10.1038/nprot.2016.011

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Kiss-Laszlo Z, Henry Y, Bachellerie JP, Caizergues-Ferrer M, Kiss T (1996) Site-specific ribose methylation of preribosomal RNA: a novel function for small nucleolar RNAs. Cell 85(7):1077–1088

    Article  CAS  Google Scholar 

  31. Tycowski KT, You ZH, Graham PJ, Steitz JA (1998) Modification of U6 spliceosomal RNA is guided by other small RNAs. Mol Cell 2(5):629–638

    Article  CAS  Google Scholar 

  32. Hearst JE (1981) Psoralen photochemistry. Annu Rev Biophys Bioeng 10:69–86. doi:10.1146/annurev.bb.10.060181.000441

    Article  CAS  PubMed  Google Scholar 

  33. Grass JA, Hei DJ, Metchette K, Cimino GD, Wiesehahn GP, Corash L, Lin L (1998) Inactivation of leukocytes in platelet concentrates by photochemical treatment with psoralen plus UVA. Blood 91(6):2180–2188

    CAS  PubMed  Google Scholar 

  34. Sastry SS, Ross BM, P’Arraga A (1997) Cross-linking of DNA-binding proteins to DNA with psoralen and psoralen furan-side monoadducts. Comparison of action spectra with DNA-DNA cross-linking. J Biol Chem 272(6):3715–3723

    Article  CAS  Google Scholar 

  35. Sastry SS, Spielmann HP, Hoang QS, Phillips AM, Sancar A, Hearst JE (1993) Laser-induced protein-DNA cross-links via psoralen furanside monoadducts. Biochemistry 32(21):5526–5538

    Article  CAS  Google Scholar 

  36. Zechel K, Weber K (1977) Degradation of nucleic acids in cell lysates by S1 nuclease in the presence of 9 M urea and sodium dodecylsulfate. Eur J Biochem 77(1):133–139

    Article  CAS  Google Scholar 

Download references

Acknowledgment

The authors wish to thank Dr. Howard Chang at Stanford University for his generous support and encouragement. Z.L. is a Layton Family Fellow of the Damon Runyon-Sohn Foundation Pediatric Cancer Fellowship Award (DRSG-14-15). This work was supported by the National Natural Science Foundation of China (31671355) and the National Thousand Young Talents Program of China to Q.C.Z., and by the Jump Start Award for Excellence in Research to Z.L. (Stanford University, award No. 106).

Author information

Authors and Affiliations

  1. Department of Dermatology, Center for Personal Dynamic Regulomes, Stanford University, Stanford, CA, 94305, USA

    Zhipeng Lu

  2. MOE Key Laboratory of Bioinformatics, Beijing Advanced Innovation Center for Structural Biology, Center for Synthetic and Systems Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Haidian, Beijing, 100084, China

    Jing Gong & Qiangfeng Cliff Zhang

Authors
  1. Zhipeng Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jing Gong
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Qiangfeng Cliff Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Qiangfeng Cliff Zhang .

Editor information

Editors and Affiliations

  1. Developmental Biology Unit, EMBL Heidelberg, Heidelberg, Germany

    Imre Gaspar

Rights and permissions

Reprints and permissions

Copyright information

© 2018 Springer Science+Business Media LLC

About this protocol

Cite this protocol

Lu, Z., Gong, J., Zhang, Q.C. (2018). PARIS: Psoralen Analysis of RNA Interactions and Structures with High Throughput and Resolution. In: Gaspar, I. (eds) RNA Detection. Methods in Molecular Biology, vol 1649. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7213-5_4

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-7213-5_4

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7212-8

  • Online ISBN: 978-1-4939-7213-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation