IDEAS home Printed from https://ideas.repec.org/a/nat/natcom/v14y2023i1d10.1038_s41467-023-41844-z.html
   My bibliography  Save this article

Chemoproteomic capture of RNA binding activity in living cells

Author

Listed:
  • Andrew J. Heindel

    (University of Virginia School of Medicine)

  • Jeffrey W. Brulet

    (University of Virginia)

  • Xiantao Wang

    (RNA Molecular Biology Laboratory, National Institute of Arthritis and Musculoskeletal and Skin Disease)

  • Michael W. Founds

    (University of Virginia)

  • Adam H. Libby

    (University of Virginia
    University of Virginia Cancer Center, University of Virginia)

  • Dina L. Bai

    (University of Virginia)

  • Michael C. Lemke

    (University of Virginia School of Medicine)

  • David M. Leace

    (University of Virginia School of Medicine)

  • Thurl E. Harris

    (University of Virginia School of Medicine)

  • Markus Hafner

    (RNA Molecular Biology Laboratory, National Institute of Arthritis and Musculoskeletal and Skin Disease)

  • Ku-Lung Hsu

    (University of Virginia School of Medicine
    University of Virginia
    University of Virginia Cancer Center, University of Virginia
    University of Virginia)

Abstract

Proteomic methods for RNA interactome capture (RIC) rely principally on crosslinking native or labeled cellular RNA to enrich and investigate RNA-binding protein (RBP) composition and function in cells. The ability to measure RBP activity at individual binding sites by RIC, however, has been more challenging due to the heterogenous nature of peptide adducts derived from the RNA-protein crosslinked site. Here, we present an orthogonal strategy that utilizes clickable electrophilic purines to directly quantify protein-RNA interactions on proteins through photoaffinity competition with 4-thiouridine (4SU)-labeled RNA in cells. Our photo-activatable-competition and chemoproteomic enrichment (PACCE) method facilitated detection of >5500 cysteine sites across ~3000 proteins displaying RNA-sensitive alterations in probe binding. Importantly, PACCE enabled functional profiling of canonical RNA-binding domains as well as discovery of moonlighting RNA binding activity in the human proteome. Collectively, we present a chemoproteomic platform for global quantification of protein-RNA binding activity in living cells.

Suggested Citation

  • Andrew J. Heindel & Jeffrey W. Brulet & Xiantao Wang & Michael W. Founds & Adam H. Libby & Dina L. Bai & Michael C. Lemke & David M. Leace & Thurl E. Harris & Markus Hafner & Ku-Lung Hsu, 2023. "Chemoproteomic capture of RNA binding activity in living cells," Nature Communications, Nature, vol. 14(1), pages 1-14, December.
  • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-41844-z
    DOI: 10.1038/s41467-023-41844-z
    as

    Download full text from publisher

    File URL: https://www.nature.com/articles/s41467-023-41844-z
    File Function: Abstract
    Download Restriction: no

    File URL: https://libkey.io/10.1038/s41467-023-41844-z?utm_source=ideas
    LibKey link: if access is restricted and if your library uses this service, LibKey will redirect you to where you can use your library subscription to access this item
    ---><---

    References listed on IDEAS

    as
    1. Keriann M. Backus & Bruno E. Correia & Kenneth M. Lum & Stefano Forli & Benjamin D. Horning & Gonzalo E. González-Páez & Sandip Chatterjee & Bryan R. Lanning & John R. Teijaro & Arthur J. Olson & Denn, 2016. "Proteome-wide covalent ligand discovery in native biological systems," Nature, Nature, vol. 534(7608), pages 570-574, June.
    2. Joel I. Perez-Perri & Birgit Rogell & Thomas Schwarzl & Frank Stein & Yang Zhou & Mandy Rettel & Annika Brosig & Matthias W. Hentze, 2018. "Discovery of RNA-binding proteins and characterization of their dynamic responses by enhanced RNA interactome capture," Nature Communications, Nature, vol. 9(1), pages 1-13, December.
    3. Benedikt M. Beckmann & Rastislav Horos & Bernd Fischer & Alfredo Castello & Katrin Eichelbaum & Anne-Marie Alleaume & Thomas Schwarzl & Tomaž Curk & Sophia Foehr & Wolfgang Huber & Jeroen Krijgsveld &, 2015. "The RNA-binding proteomes from yeast to man harbour conserved enigmRBPs," Nature Communications, Nature, vol. 6(1), pages 1-9, December.
    4. Florian Erhard & Marisa A. P. Baptista & Tobias Krammer & Thomas Hennig & Marius Lange & Panagiota Arampatzi & Christopher S. Jürges & Fabian J. Theis & Antoine-Emmanuel Saliba & Lars Dölken, 2019. "scSLAM-seq reveals core features of transcription dynamics in single cells," Nature, Nature, vol. 571(7765), pages 419-423, July.
    5. Thomas Conrad & Anne-Susann Albrecht & Veronica Rodrigues de Melo Costa & Sascha Sauer & David Meierhofer & Ulf Andersson Ørom, 2016. "Serial interactome capture of the human cell nucleus," Nature Communications, Nature, vol. 7(1), pages 1-11, September.
    Full references (including those not matched with items on IDEAS)

    Most related items

    These are the items that most often cite the same works as this one and are cited by the same works as this one.
    1. Haofan Sun & Bin Fu & Xiaohong Qian & Ping Xu & Weijie Qin, 2024. "Nuclear and cytoplasmic specific RNA binding proteome enrichment and its changes upon ferroptosis induction," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    2. Joel I. Perez-Perri & Dunja Ferring-Appel & Ina Huppertz & Thomas Schwarzl & Sudeep Sahadevan & Frank Stein & Mandy Rettel & Bruno Galy & Matthias W. Hentze, 2023. "The RNA-binding protein landscapes differ between mammalian organs and cultured cells," Nature Communications, Nature, vol. 14(1), pages 1-20, December.
    3. Johanna Luige & Alexandros Armaos & Gian Gaetano Tartaglia & Ulf Andersson Vang Ørom, 2024. "Predicting nuclear G-quadruplex RNA-binding proteins with roles in transcription and phase separation," Nature Communications, Nature, vol. 15(1), pages 1-13, December.
    4. JohnCarlo Kristofich & Christopher V. Nicchitta, 2023. "Signal-noise metrics for RNA binding protein identification reveal broad spectrum protein-RNA interaction frequencies and dynamics," Nature Communications, Nature, vol. 14(1), pages 1-20, December.
    5. Teresa Rummel & Lygeri Sakellaridi & Florian Erhard, 2023. "grandR: a comprehensive package for nucleotide conversion RNA-seq data analysis," Nature Communications, Nature, vol. 14(1), pages 1-17, December.
    6. Bibiana Costa & Jennifer Becker & Tobias Krammer & Felix Mulenge & Verónica Durán & Andreas Pavlou & Olivia Luise Gern & Xiaojing Chu & Yang Li & Luka Čičin-Šain & Britta Eiz-Vesper & Martin Messerle , 2024. "Human cytomegalovirus exploits STING signaling and counteracts IFN/ISG induction to facilitate infection of dendritic cells," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    7. Liang-Cui Chu & Pedro Arede & Wei Li & Erika C. Urdaneta & Ivayla Ivanova & Stuart W. McKellar & Jimi C. Wills & Theresa Fröhlich & Alexander Kriegsheim & Benedikt M. Beckmann & Sander Granneman, 2022. "The RNA-bound proteome of MRSA reveals post-transcriptional roles for helix-turn-helix DNA-binding and Rossmann-fold proteins," Nature Communications, Nature, vol. 13(1), pages 1-18, December.
    8. Anna Rodina & Chao Xu & Chander S. Digwal & Suhasini Joshi & Yogita Patel & Anand R. Santhaseela & Sadik Bay & Swathi Merugu & Aftab Alam & Pengrong Yan & Chenghua Yang & Tanaya Roychowdhury & Palak P, 2023. "Systems-level analyses of protein-protein interaction network dysfunctions via epichaperomics identify cancer-specific mechanisms of stress adaptation," Nature Communications, Nature, vol. 14(1), pages 1-26, December.
    9. Ebru Aydin & Silke Schreiner & Jacqueline Böhme & Birte Keil & Jan Weber & Bojan Žunar & Timo Glatter & Cornelia Kilchert, 2024. "DEAD-box ATPase Dbp2 is the key enzyme in an mRNP assembly checkpoint at the 3’-end of genes and involved in the recycling of cleavage factors," Nature Communications, Nature, vol. 15(1), pages 1-20, December.
    10. Lukas Bartonek & Bojan Zagrovic, 2017. "mRNA/protein sequence complementarity and its determinants: The impact of affinity scales," PLOS Computational Biology, Public Library of Science, vol. 13(7), pages 1-16, July.
    11. Tin-Yan Koo & Hinyuk Lai & Daniel K. Nomura & Clive Yik-Sham Chung, 2023. "N-Acryloylindole-alkyne (NAIA) enables imaging and profiling new ligandable cysteines and oxidized thiols by chemoproteomics," Nature Communications, Nature, vol. 14(1), pages 1-19, December.
    12. Meeli Mullari & Nicolas Fossat & Niels H. Skotte & Andrea Asenjo-Martinez & David T. Humphreys & Jens Bukh & Agnete Kirkeby & Troels K. H. Scheel & Michael L. Nielsen, 2023. "Characterising the RNA-binding protein atlas of the mammalian brain uncovers RBM5 misregulation in mouse models of Huntington’s disease," Nature Communications, Nature, vol. 14(1), pages 1-20, December.
    13. Jasper Panten & Tobias Heinen & Christina Ernst & Nils Eling & Rebecca E. Wagner & Maja Satorius & John C. Marioni & Oliver Stegle & Duncan T. Odom, 2024. "The dynamic genetic determinants of increased transcriptional divergence in spermatids," Nature Communications, Nature, vol. 15(1), pages 1-13, December.
    14. Yang Zhou & Partho Sarothi Ray & Jianguo Zhu & Frank Stein & Mandy Rettel & Thileepan Sekaran & Sudeep Sahadevan & Joel I. Perez-Perri & Eva K. Roth & Ola Myklebost & Leonardo A. Meza-Zepeda & Andreas, 2024. "Systematic analysis of RNA-binding proteins identifies targetable therapeutic vulnerabilities in osteosarcoma," Nature Communications, Nature, vol. 15(1), pages 1-22, December.
    15. Carmen Aguilar & Mindaugas Pauzuolis & Malvika Pompaiah & Ehsan Vafadarnejad & Panagiota Arampatzi & Mara Fischer & Dominik Narres & Mastura Neyazi & Özge Kayisoglu & Thomas Sell & Nils Blüthgen & Mar, 2022. "Helicobacter pylori shows tropism to gastric differentiated pit cells dependent on urea chemotaxis," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    16. Shichao Lin & Kun Yin & Yingkun Zhang & Fanghe Lin & Xiaoyong Chen & Xi Zeng & Xiaoxu Guo & Huimin Zhang & Jia Song & Chaoyong Yang, 2023. "Well-TEMP-seq as a microwell-based strategy for massively parallel profiling of single-cell temporal RNA dynamics," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    17. Lior Fishman & Avani Modak & Gal Nechooshtan & Talya Razin & Florian Erhard & Aviv Regev & Jeffrey A. Farrell & Michal Rabani, 2024. "Cell-type-specific mRNA transcription and degradation kinetics in zebrafish embryogenesis from metabolically labeled single-cell RNA-seq," Nature Communications, Nature, vol. 15(1), pages 1-20, December.
    18. Jing Gao & Bo Hou & Qiwen Zhu & Lei Yang & Xingyu Jiang & Zhifeng Zou & Xutong Li & Tianfeng Xu & Mingyue Zheng & Yi-Hung Chen & Zhiai Xu & Huixiong Xu & Haijun Yu, 2022. "Engineered bioorthogonal POLY-PROTAC nanoparticles for tumour-specific protein degradation and precise cancer therapy," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    19. Wai Cheung Chan & Xiaoxi Liu & Robert S. Magin & Nicholas M. Girardi & Scott B. Ficarro & Wanyi Hu & Maria I. Tarazona Guzman & Cara A. Starnbach & Alejandra Felix & Guillaume Adelmant & Anthony C. Va, 2023. "Accelerating inhibitor discovery for deubiquitinating enzymes," Nature Communications, Nature, vol. 14(1), pages 1-13, December.

    More about this item

    Statistics

    Access and download statistics

    Corrections

    All material on this site has been provided by the respective publishers and authors. You can help correct errors and omissions. When requesting a correction, please mention this item's handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-41844-z. See general information about how to correct material in RePEc.

    If you have authored this item and are not yet registered with RePEc, we encourage you to do it here. This allows to link your profile to this item. It also allows you to accept potential citations to this item that we are uncertain about.

    If CitEc recognized a bibliographic reference but did not link an item in RePEc to it, you can help with this form .

    If you know of missing items citing this one, you can help us creating those links by adding the relevant references in the same way as above, for each refering item. If you are a registered author of this item, you may also want to check the "citations" tab in your RePEc Author Service profile, as there may be some citations waiting for confirmation.

    For technical questions regarding this item, or to correct its authors, title, abstract, bibliographic or download information, contact: Sonal Shukla or Springer Nature Abstracting and Indexing (email available below). General contact details of provider: http://www.nature.com .

    Please note that corrections may take a couple of weeks to filter through the various RePEc services.

    IDEAS is a RePEc service. RePEc uses bibliographic data supplied by the respective publishers.