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Structural basis of paralog-specific KDM2A/B nucleosome recognition

Nature Chemical Biology volume 19, pages 624–632 (2023)Cite this article

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Abstract

The nucleosome acidic patch is a major interaction hub for chromatin, providing a platform for enzymes to dock and orient for nucleosome-targeted activities. To define the molecular basis of acidic patch recognition proteome wide, we performed an amino acid resolution acidic patch interactome screen. We discovered that the histone H3 lysine 36 (H3K36) demethylase KDM2A, but not its closely related paralog, KDM2B, requires the acidic patch for nucleosome binding. Despite fundamental roles in transcriptional repression in health and disease, the molecular mechanisms governing nucleosome substrate specificity of KDM2A/B, or any related JumonjiC (JmjC) domain lysine demethylase, remain unclear. We used a covalent conjugate between H3K36 and a demethylase inhibitor to solve cryogenic electron microscopy structures of KDM2A and KDM2B trapped in action on a nucleosome substrate. Our structures show that KDM2–nucleosome binding is paralog specific and facilitated by dynamic nucleosomal DNA unwrapping and histone charge shielding that mobilize the H3K36 sequence for demethylation.

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Fig. 1: KDM2 paralog-specific acidic patch binding identified by amino acid resolution nucleosome affinity proteomics.
Fig. 2: Stabilization of JmjC–nucleosome interaction enables structural analysis of KDM2A/B–nucleosome interaction.
Fig. 3: KDM2A interacts with nucleosome acidic patch.
Fig. 4: Dynamic and multivalent KDM2A–nucleosome interactions.

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Data availability

The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD032791. Cryo-EM maps have been deposited in the Electron Microscopy Data Bank under accession codes EMD-26809 (KDM2A:H3K36C-UNC8015 nucleosome complex) and EMD-26810 (KDM2B:H3K36C-UNC8015 nucleosome complex). Atomic coordinates of the KDM2A:H3K36C-UNC8015 nucleosome and KMD2A:H3(29-41)K36C-UNC8015 complexes have been deposited in the Protein Data Bank with accession codes 7UV9 and 7UVA, respectively. Homology model of KDM2B:H3K36C-UNC8015 nucleosome is included as Supplementary File 3. Other structures used in this study are available through the Protein Data Bank with accession codes 4QX7, 6ESF, 6NZO, 7CRR and 7JO9. All materials in this study are available upon reasonable request. Source data are provided with this paper.

Code availability

In-house R script used for proteomics data analysis is available for download at GitHub (https://github.com/GoldfarbLab/NucleosomeProteomeAnalysis).

References

  1. Hyun, K., Jeon, J., Park, K. & Kim, J. Writing, erasing and reading histone lysine methylations. Exp. Mol. Med. 49, e324 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Tsukada, Y.-I. et al. Histone demethylation by a family of JmjC domain-containing proteins. Nature 439, 811–816 (2006).

    Article  CAS  PubMed  Google Scholar 

  3. Shi, Y. et al. Histone demethylation mediated by the nuclear amine oxidase homolog LSD1. Cell 119, 941–953 (2004).

    Article  CAS  PubMed  Google Scholar 

  4. Klose, R. J. & Zhang, Y. Regulation of histone methylation by demethylimination and demethylation. Nat. Rev. Mol. Cell Biol. 8, 307–318 (2007).

    Article  CAS  PubMed  Google Scholar 

  5. Shi, Y. & Whetstine, J. R. Dynamic regulation of histone lysine methylation by demethylases. Mol. Cell 25, 1–14 (2007).

    Article  CAS  PubMed  Google Scholar 

  6. Black, J. C., Van Rechem, C. & Whetstine, J. R. Histone lysine methylation dynamics: establishment, regulation, and biological impact. Mol. Cell 48, 491–507 (2012).

    Article  CAS  PubMed  Google Scholar 

  7. Punnia-Moorthy, G., Hersey, P., Emran, A. A. & Tiffen, J. Lysine demethylases: promising drug targets in melanoma and other cancers. Front. Genet. 12, 680633 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Borgel, J. et al. KDM2A integrates DNA and histone modification signals through a CXXC/PHD module and direct interaction with HP1. Nucleic Acids Res. 45, 1114–1129 (2017).

    CAS  PubMed  Google Scholar 

  9. Frescas, D. et al. KDM2A represses transcription of centromeric satellite repeats and maintains the heterochromatic state. Cell Cycle 7, 3539–3547 (2008).

    Article  CAS  PubMed  Google Scholar 

  10. Bartke, T. et al. Nucleosome-interacting proteins regulated by DNA and histone methylation. Cell 143, 470–484 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Gao, Z. et al. PCGF homologs, CBX proteins, and RYBP define functionally distinct PRC1 family complexes. Mol. Cell 45, 344–356 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Gearhart, M. D., Corcoran, C. M., Wamstad, J. A. & Bardwell, V. J. Polycomb group and SCF ubiquitin ligases are found in a novel BCOR complex that is recruited to BCL6 targets. Mol. Cell. Biol. 26, 6880–6889 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Lagarou, A. et al. dKDM2 couples histone H2A ubiquitylation to histone H3 demethylation during Polycomb group silencing. Genes Dev. 22, 2799–2810 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Sánchez, C. et al. Proteomics analysis of Ring1B/Rnf2 interactors identifies a novel complex with the Fbxl10/Jhdm1B histone demethylase and the Bcl6 interacting corepressor. Mol. Cell. Proteom. 6, 820–834 (2007).

    Article  Google Scholar 

  15. Wu, X., Johansen, J. V. & Helin, K. Fbxl10/Kdm2b recruits polycomb repressive complex 1 to CpG islands and regulates H2A ubiquitylation. Mol. Cell 49, 1134–1146 (2013).

    Article  CAS  PubMed  Google Scholar 

  16. Farcas, A. M. et al. KDM2B links the Polycomb Repressive Complex 1 (PRC1) to recognition of CpG islands. eLife 1, e00205–e00205 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  17. Blackledge, N. P. et al. CpG islands recruit a histone H3 lysine 36 demethylase. Mol. Cell 38, 179–190 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Vacík, T., Lađinović, D. & Raška, I. KDM2A/B lysine demethylases and their alternative isoforms in development and disease. Nucleus 9, 431–441 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  19. Yan, M., Yang, X., Wang, H. & Shao, Q. The critical role of histone lysine demethylase KDM2B in cancer. Am. J. Transl. Res 10, 2222–2233 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Cheng, Z. et al. A molecular threading mechanism underlies Jumonji lysine demethylase KDM2A regulation of methylated H3K36. Genes Dev. 28, 1758–1771 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Skrajna, A. et al. Comprehensive nucleosome interactome screen establishes fundamental principles of nucleosome binding. Nucleic Acids Res. 48, 9415–9432 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Nacev, B. A. et al. The expanding landscape of ‘oncohistone’ mutations in human cancers. Nature 567, 473–478 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. McGinty, R. K. & Tan, S. Principles of nucleosome recognition by chromatin factors and enzymes. Curr. Opin. Struct. Biol. 71, 16–26 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Bennett, R. L. et al. A mutation in histone H2B represents a new class of oncogenic driver. Cancer Disco. 9, 1438–1451 (2019).

    Article  CAS  Google Scholar 

  25. Wan, Y. C. E. et al. Cancer-associated histone mutation H2BG53D disrupts DNA–histone octamer interaction and promotes oncogenic phenotypes. Signal Transduct. Target Ther. 5, 27–4 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Arimura, Y. et al. Cancer-associated mutations of histones H2B, H3.1 and H2A.Z.1 affect the structure and stability of the nucleosome. Nucleic Acids Res. 46, 10007–10018 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Weinberg, D. N., Allis, C. D. & Lu, C. Oncogenic mechanisms of histone H3 mutations. Cold Spring Harb. Perspect. Med. 7, a026443 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  28. Zhou, J. C., Blackledge, N. P., Farcas, A. M. & Klose, R. J. Recognition of CpG island chromatin by KDM2A requires direct and specific interaction with linker DNA. Mol. Cell. Biol. 32, 479–489 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Hamada, S. et al. Design, synthesis, enzyme-inhibitory activity, and effect on human cancer cells of a novel series of jumonji domain-containing protein 2 histone demethylase inhibitors. J. Med. Chem. 53, 5629–5638 (2010).

    Article  CAS  PubMed  Google Scholar 

  30. Suzuki, T. et al. Identification of the KDM2/7 histone lysine demethylase subfamily inhibitor and its antiproliferative activity. J. Med. Chem. 56, 7222–7231 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Luo, X. et al. A selective inhibitor and probe of the cellular functions of Jumonji C domain-containing histone demethylases. J. Am. Chem. Soc. 133, 9451–9456 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Wesley, N. A. et al. Time resolved-fluorescence resonance energy transfer platform for quantitative nucleosome binding and footprinting. Protein Sci. 31, e4339 (2022).

    Article  CAS  PubMed  Google Scholar 

  33. Li, W. et al. Molecular basis of nucleosomal H3K36 methylation by NSD methyltransferases. Nature 590, 498–503 (2021).

    Article  CAS  PubMed  Google Scholar 

  34. Bilokapic, S. & Halic, M. Nucleosome and ubiquitin position Set2 to methylate H3K36. Nat. Commun. 10, 3795 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  35. Bagert, J. D. et al. Oncohistone mutations enhance chromatin remodeling and alter cell fates. Nat. Chem. Biol. 17, 403–411 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Spangler, C. J. et al. DOT1L activity in leukemia cells requires interaction with ubiquitylated H2B that promotes productive nucleosome binding. Cell Rep. 38, 110369 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Anderson, C. J. et al. Structural basis for recognition of ubiquitylated nucleosome by Dot1L methyltransferase. Cell Rep. 26, 1681–1690 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Tan, S., Kern, R. C. & Selleck, W. The pST44 polycistronic expression system for producing protein complexes in Escherichia coli. Protein Expr. Purif. 40, 385–395 (2005).

    Article  CAS  PubMed  Google Scholar 

  39. Weissmann, F. et al. biGBac enables rapid gene assembly for the expression of large multisubunit protein complexes. Proc. Natl Acad. Sci. USA 113, E2564–E2569 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Lowary, P. T. & Widom, J. New DNA sequence rules for high affinity binding to histone octamer and sequence-directed nucleosome positioning. J. Mol. Biol. 276, 19–42 (1998).

    Article  CAS  PubMed  Google Scholar 

  41. Kim, S.-A., Chatterjee, N., Jennings, M. J., Bartholomew, B. & Tan, S. Extranucleosomal DNA enhances the activity of the LSD1/CoREST histone demethylase complex. Nucleic Acids Res. 43, 4868–4880 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Makde, R. D., England, J. R., Yennawar, H. P. & Tan, S. Structure of RCC1 chromatin factor bound to the nucleosome core particle. Nature 467, 562–566 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Luger, K., Rechsteiner, T. J. & Richmond, T. J. Preparation of nucleosome core particle from recombinant histones. Methods Enzymol. 304, 3–19 (1999).

    Article  CAS  PubMed  Google Scholar 

  44. Luger, K., Rechsteiner, T. J., Flaus, A. J., Waye, M. M. & Richmond, T. J. Characterization of nucleosome core particles containing histone proteins made in bacteria. J. Mol. Biol. 272, 301–311 (1997).

    Article  CAS  PubMed  Google Scholar 

  45. Dignam, J. D., Lebovitz, R. M. & Roeder, R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 11, 1475–1489 (1983).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Pieters, B. J. G. E. et al. Installation of trimethyllysine analogs on intact histones via cysteine alkylation. Bioconjug. Chem. 30, 952–958 (2019).

    Article  CAS  PubMed  Google Scholar 

  47. Kabsch, W. XDS. Acta Crystallogr. D Biol. Crystallogr. 66, 125–132 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Evans, P. R. An introduction to data reduction: space-group determination, scaling and intensity statistics. Acta Crystallogr. D Biol. Crystallogr. 67, 282–292 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Murshudov, G. N., Vagin, A. A. & Dodson, E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).

    Article  CAS  PubMed  Google Scholar 

  52. Zivanov, J., Nakane, T. & Scheres, S. H. W. Estimation of high-order aberrations and anisotropic magnification from cryo-EM data sets in RELION-3.1. IUCrJ 7, 253–267 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Rohou, A. & Grigorieff, N. CTFFIND4: fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  54. Zhong, E. D., Bepler, T., Berger, B. & Davis, J. H. CryoDRGN: reconstruction of heterogeneous cryo-EM structures using neural networks. Nat. Methods 18, 176–185 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Afonine, P. V. et al. New tools for the analysis and validation of cryo-EM maps and atomic models. Acta Crystallogr D Struct. Biol. 74, 814–840 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D Biol. Crystallogr. 66, 12–21 (2010).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

Single-particle cryo-EM data were collected with assistance from J. Peck and J. Strauss at the University of North Carolina at Chapel Hill School of Medicine Cryo-Electron Microscopy Facility, which is partially supported by National Institutes of Health (NIH) grant P30CA016086 and is part of the Molecular Microscopy Consortium of the University of North Carolina at Chapel Hill, Duke University and the National Institute of Environmental Health Sciences. Crystallography data were collected at the Southeast Regional Collaborative Access Team (SER-CAT) 22-ID beamline at the Advanced Photon Source, Argonne National Laboratory. SER-CAT is supported by its member institutions and NIH equipment grants S10_RR25528, S10_RR028976 and S10_OD027000. Plasmids containing 601 nucleosome positioning DNA fragments and NSD2 were gifts from S. Tan. We thank members of the McGinty, Goldfarb, Frye and James laboratories for critical comments on the manuscript. This work was supported by NIH grants R35GM133498 to R.K.M., F99CA253730 to C.J.S., R35GM139514 to S.V.F., R01CA010305 to L.I.J. and R01GM132299 to D.K.; American Cancer Society grant 132609-PF-18-153-01-DMC to A.S.; and National Science Foundation grant DGE-1650116 to N.A.W.

Author information

Author notes

  1. Cathy J. Spangler

    Present address: Vollum Institute, Oregon Health and Science University, Portland, OR, USA

  2. Dmitri Kireev

    Present address: Department of Chemistry, University of Missouri, Columbia, MO, USA

  3. These authors contributed equally: Cathy J. Spangler, Aleksandra Skrajna.

Authors and Affiliations

  1. Department of Biochemistry and Biophysics, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Cathy J. Spangler, Gabrielle R. Budziszewski, Nathaniel A. Wesley & Robert K. McGinty

  2. Center for Integrative Chemical Biology and Drug Discovery, Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Aleksandra Skrajna, Caroline A. Foley, Dalal N. Azzam, Eyla C. Arteaga, Holly C. Simmons, Charlotte B. Smith, Dmitri Kireev, Lindsey I. James, Stephen V. Frye & Robert K. McGinty

  3. Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO, USA

    Anh Nguyen, Emily M. Wilkerson & Dennis Goldfarb

  4. Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC, USA

    Jeanne-Marie E. McPherson

  5. Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Lindsey I. James, Stephen V. Frye & Robert K. McGinty

  6. Institute for Informatics, Washington University School of Medicine, St. Louis, MO, USA

    Dennis Goldfarb

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Contributions

R.K.M., D.G., S.V.F. and L.I.J. supervised the studies. C.J.S., A.S. and R.K.M. conceived the study and designed the experiments. C.J.S. prepared samples, grids, collected and processed the cryo-EM data and performed in vitro functional assays. A.S. prepared samples and performed proteomics experiments and in vitro functional assays. C.A.F. helped design and prepared cofactor analogs. A.N. and D.G. processed proteomics data. G.R.B. and D.N.A. prepared samples and solved the crystal structure. E.C.A., H.C.S., C.B.S. and J.-M.E.M. prepared samples. E.M.W. collected proteomics data. N.A.W. developed the nucleosome binding assay and performed nucleosome interaction studies. D.K. modeled substrate analogs. C.J.S., A.S. and R.K.M. wrote the manuscript, with contributions from all coauthors.

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Correspondence to Robert K. McGinty.

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Extended data

Extended Data Fig. 1 Amino acid resolution nucleosome affinity proteomics screen.

a, Schematic of nucleosome affinity proteomics screen. b, Coomassie-stained SDS-polyacrylamide denaturing gel (top) and ethidium bromide-stained native gel (bottom) of biotinylated nucleosome library. These experiments were repeated independently two times with similar results. c, Triplicate pulldowns run briefly on SDS polyacrylamide gels and stained with Coomassie blue prior to excision and in-gel trypsin digestion. Unused replicates of wild-type nucleosome pulldowns marked with an asterisk. d, Western blots following pulldowns from HEK293T nuclear lysates using indicated biotinylated nucleosomes. These experiments were performed once. WT, wild-type.

Extended Data Fig. 2 Oncohistone mutations have distinct effects on nucleosome binding.

a, Full heat map showing log2-fold change relative to wild-type nucleosome of quantified proteins listed at right across screen, clustered by protein and nucleosome mutant. b, Heat map of oncohistone mutant nucleosomes, clustered as in a. c, Heat map of each oncohistone mutation relative to the corresponding alanine mutation, clustered as in a.

Extended Data Fig. 3 KDM2A/B-nucleosome complex purification and 2D classification.

a, Chromatograms displaying normalized 280 nm absorbance from size exclusion purification of KDM2B/WT nucleosome (left), KDM2B/H3K36C-UNC8015 nucleosome (middle), and KDM2A/H3K36C-UNC8015 nucleosome (right) complex reconstitutions. These experiments were performed once. b, Highest populated 2D classes attained from cryo-EM analysis of indicated samples, with selected top views of KDM2A/B-unbound and bound nucleosomes.

Extended Data Fig. 4 Design and synthesis of H3K36-JmjC inhibitor covalent conjugate.

a, Chemical structures of JmjC inhibitors. b, Inhibition assays using H3K36me2 peptide substrates with KDM2B(2–734). c, Dose response inhibition of KDM2B(2–734) by UNC8016 on H3K36me2 peptide substrates. In b and c, mean ± SD are shown (n = 3 assay technical replicates). d, Synthetic scheme for alkylation reaction of H3K36C mutant with UNC8015. e, LCMS spectra and mass deconvolution (determined using ESIprot) of unreacted H3K36C (left) and reacted H3K36C-UNC8015 (right; 5-hour endpoint).

Extended Data Fig. 5 Crystal structure of KDM2A bound to H3K36-UNC8015 peptide covalent conjugate.

a, Structure of KDM2A catalytic domain in complex with H3K36C-UNC8015. b, Identical view of structure of KDM2A catalytic domain in complex with H3K36me2 peptide and α-ketoglutarate (α-KG). c-e, Zoomed views of c, H3K36C-UNC8015 peptide, d, H3K36me2 peptide and α-KG, and e, overlayed aligned structures. f, 2Fo-Fc electron density map of H3K36C-UNC8015 peptide at 1σ.

Extended Data Fig. 6 Cryo-EM data analysis for KDM2A-nucleosome complex.

a, Representative micrograph (left), CTF estimation (middle), and 2D classes (right) from KDM2A-nucleosome complex dataset. b, Cryo-EM data analysis workflow carried out in RELION-3.1. c, Multibody refinement analysis results from RELION. d, Final postprocessed map colored by local resolution (top), angular distribution of particles (middle), and FSC curve (bottom) for KDM2A-nucleosome complex.

Extended Data Fig. 7 Cryo-EM data analysis for KDM2B-nucleosome complex.

a, Representative micrograph (left), CTF estimation (middle), and 2D classes (right) from KDM2B-nucleosome complex dataset. b, Cryo-EM data analysis workflow carried out in RELION-3.1. c, Multibody refinement analysis results from RELION. d, Final postprocessed map colored by local resolution (top), angular distribution of particles (middle), and FSC curve (bottom) for KDM2B-nucleosome complex.

Extended Data Fig. 8 Unwrapped DNA-binding interfaces of KDM2A/B.

a, Sequence alignment of KDM2A and KDM2B DNA-binding interfaces with closely related KDM7A and PHF8 (KDM7B), with conserved basic amino acids and charge swapped amino acids colored blue and red, respectively. b, Zoomed view of unwrapped DNA-binding interface of KDM2A structure (left) and KDM2B structure (right). c, TR-FRET binding isotherms with KDM2A DNA interface mutants and unmodified wild-type 185 bp nucleosomes. d, TR-FRET nucleosome binding competition experiments using unmodified wildtype 185, 147, or 119 bp nucleosomes. In c and d, mean ± SD are shown (n = 3 assay technical replicates).

Extended Data Fig. 9 Acidic patch-localized density of KDM2A/B.

a, b, Acidic patch-localized density in nucleosome-focused cryo-EM maps from multibody analysis of a KDM2A- and b KDM2B-nucleosome complexes at indicated map thresholds. c, Overlay of acidic patch-localized density from nucleosome-focused map shown in a with equivalent region of modeled KDM2A. d, Sequence alignment of KDM2A and KDM2B N-termini, with basic amino acids colored blue.

Extended Data Fig. 10 Comparison of H3K36-bound enzyme structures.

a, Structures of H3K36 lysine demethylase KDM2A, b, H3K36 lysine methyltransferase Set2 (PDB:6NZO) and c, H3K36 lysine methyltransferase NSD3 (PDB:7CRR) bound to the nucleosome. d, Sequence alignment of KDM2A and KDM2B JmjC-adjacent sequences, with acidic amino acids colored red.

Supplementary information

Supplementary Information

Supplementary Tables 1 and 2, Supplementary Figs. 1–7 and Supplementary Note

Reporting Summary

Supplementary Data 1

Nucleosome affinity proteomics data and statistics

Supplementary Data 2

Profile plots from nucleosome affinity proteomics

Supplementary Data 3

KDM2B–nucleosome model

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Spangler, C.J., Skrajna, A., Foley, C.A. et al. Structural basis of paralog-specific KDM2A/B nucleosome recognition. Nat Chem Biol 19, 624–632 (2023). https://doi.org/10.1038/s41589-023-01256-y

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