Research Summary

Our group investigates mechanisms, regulation, and biological functions of methyl group addition to proteins and RNA. Methylation, a common post-transcriptional and post-translational modification, has a profound effect on the regulation of fundamental biological processes such as gene expression, cellular localization, and RNA structure and function. Deregulation of methylation is associated with a wide range of diseases. The enzymatic regulation of methyl group addition and removal provides an opportunity for therapeutic intervention. We seek to understand the molecular mechanisms that control methylation, and develop chemical probes to interrogate the pathophysiological function of enzymes that regulate this modification. Specifically, our research focuses on the following areas:

Regulation and Small Molecule Inhibition of Jumonji Histone Demethylases
Jumonji histone demethylases, a family of epigenetic “erasers”, catalyze the removal of methyl marks from lysine residues in proteins. Jumonji demethylases are complex proteins that, in addition to the catalytic domain, often contain one or more chromatin “reader” domains. The reader modules commonly interact with chromatin, and this interaction can be modulated by chromatin modifications. We investigate the functional cross-talk between chromatin recognition and demethylation in the jumonji family to understand how chromatin context impacts methyl mark removal, and consequently transcription. Furthermore, we are interested in understanding how additional regulatory inputs, such as metabolism and cellular signaling cascades, influence chromatin methylation and transcriptional regulation. In addition, our lab is actively involved in the development of small molecule inhibitors of the jumonji demethylases that can be used as cellular probes of their function. We use both rational design and high-throughput screening to identify starting scaffolds, and further optimize these scaffolds through iterative cycles of chemical synthesis and testing their potency and selectivity. Our goal is to use these molecules to inhibit aberrant demethylation caused by misregulation of demethylases in disease models.

Mechanisms and Cellular Roles of RNA Methylation
Methylation of RNA is the abundant post-transcriptional modification identified in various types of RNAs. Despite its prevalence, the functional role of methylation is poorly understood. We are interested in elucidating the mechanisms responsible for RNA methylation, and understanding the role this modification plays in controlling the cellular function of RNA. We are particularly interested in 2-methyl and 8-methyladenosine modifications, catalyzed by related enzymes that utilize an unusual mechanism to achieve methylation. Incorporation of 2-methyladenosine into RNA has been implicated in the regulation of translational fidelity, although the mechanisms by which this is achieved are yet to be elucidated. In contrast, 8-methyladenosine formation is responsible for resistance to five chemically distinct classes of antibiotics that target the peptidyltransferase center of the bacterial ribosome, including linezolid. We investigate catalytic mechanisms, substrate recognition, and evolution of function in enzymes that carry out these methylations. Our goal is to determine the impact of methylation on the cellular function of substrate RNA.

Research Funding

  • May 15, 2022 - April 30, 2025 - Development of Novel Antivirals Targeting Viral RNA Methylation , AViDD Project 4, lead PI . Sponsor: NIAID, Sponsor Award ID: U19AI171110
  • July 1, 2020 - June 30, 2024 - Discovery of Small Molecule Ligands for PHD1 Reader Domain of Histone Demethylase KDM5A , Principal Investigator . Sponsor: NIH, Sponsor Award ID: R01CA250459
  • September 14, 2018 - August 31, 2022 - Radical SAM-dependent methylation in antibiotic resistance , Principal Investigator . Sponsor: NIH/NIAID, Sponsor Award ID: R01AI137270
  • September 15, 2015 - August 31, 2019 - Allosteric Regulation in the KDM5 Family of Histone Demethylases , Principal Investigator . Sponsor: NIH, Sponsor Award ID: R01GM114044

Education

University of Belgrade, Belgrade, Serbia BSc 07/2000 Chemistry
University of Illinois, Urbana, IL PhD 05/2005 Chemistry
Harvard Medical School, Boston, MA Postdoc 06/2008 Biochemistry

Honors & Awards

  • 2000
    Serbian Chemical Society Outstanding Undergraduate Student Award
  • 2001
    University of Illinois Teachers Rated as Excellent
  • 2003
    Procter and Gamble Predoctoral Fellowship
  • 2004
    University of Illinois Pines Predoctoral Fellowship
  • 2005
    Damon Runyon Cancer Research Foundation Postdoctoral Fellowship
  • 2007
    NIH Pathway to Independence Award
  • 2009
    Kimmel Scholar Award
  • 2010
    V Foundation Scholar Award
  • 2011
    Basil O'Connor Starter Scholar Award, March of Dimes
  • 2011
    NSF CAREER Award
  • 2011
    Searle Scholar Award
  • 2014
    UCSF Excellence in Teaching Award

Selected Publications

  1. Zhang MY, Yang H, Ortiz G, Trnka MJ, Petronikolou N, Burlingame AL, DeGrado WF, Fujimori DG. Covalent labeling of a chromatin reader domain using proximity-reactive cyclic peptides. Chem Sci. 2022 Jun 07; 13(22):6599-6609.  View on PubMed
  2. Tsai K, Stojkovic V, Lee DJ, Young ID, Szal T, Klepacki D, Vázquez-Laslop N, Mankin AS, Fraser JS, Fujimori DG. Structural basis for context-specific inhibition of translation by oxazolidinone antibiotics. Nat Struct Mol Biol. 2022 02; 29(2):162-171.  View on PubMed
  3. Tsai K, Stojkovic V, Noda-Garcia L, Young ID, Myasnikov AG, Kleinman J, Palla A, Floor SN, Frost A, Fraser JS, Tawfik DS, Fujimori DG. Directed evolution of the rRNA methylating enzyme Cfr reveals molecular basis of antibiotic resistance. Elife. 2022 01 11; 11.  View on PubMed
  4. Longbotham JE, Kelly MJS, Fujimori DG. Recognition of Histone H3 Methylation States by the PHD1 Domain of Histone Demethylase KDM5A. ACS Chem Biol. 2021 Feb 23.  View on PubMed
  5. Stojkovic V, Weinberg DE, Fujimori DG. miCLIP-MaPseq Identifies Substrates of Radical SAM RNA-Methylating Enzyme Using Mechanistic Cross-Linking and Mismatch Profiling. Methods Mol Biol. 2021; 2298:105-122.  View on PubMed
  6. Anderson SE, Longbotham JE, O'Kane PT, Ugur FS, Fujimori DG, Mrksich M. Exploring the Ligand Preferences of the PHD1 Domain of Histone Demethylase KDM5A Reveals Tolerance for Modifications of the Q5 Residue of Histone 3. ACS Chem Biol. 2021 01 15; 16(1):205-213.  View on PubMed
  7. Longbotham JE, Zhang MY, Fujimori DG. Domain cross-talk in regulation of histone modifications: Molecular mechanisms and targeting opportunities. Curr Opin Chem Biol. 2020 08; 57:105-113.  View on PubMed
  8. Gordon DE, Jang GM, Bouhaddou M, Xu J, Obernier K, White KM, O'Meara MJ, Rezelj VV, Guo JZ, Swaney DL, Tummino TA, Hüttenhain R, Kaake RM, Richards AL, Tutuncuoglu B, Foussard H, Batra J, Haas K, Modak M, Kim M, Haas P, Polacco BJ, Braberg H, Fabius JM, Eckhardt M, Soucheray M, Bennett MJ, Cakir M, McGregor MJ, Li Q, Meyer B, Roesch F, Vallet T, Mac Kain A, Miorin L, Moreno E, Naing ZZC, Zhou Y, Peng S, Shi Y, Zhang Z, Shen W, Kirby IT, Melnyk JE, Chorba JS, Lou K, Dai SA, Barrio-Hernandez I, Memon D, Hernandez-Armenta C, Lyu J, Mathy CJP, Perica T, Pilla KB, Ganesan SJ, Saltzberg DJ, Rakesh R, Liu X, Rosenthal SB, Calviello L, Venkataramanan S, Liboy-Lugo J, Lin Y, Huang XP, Liu Y, Wankowicz SA, Bohn M, Safari M, Ugur FS, Koh C, Savar NS, Tran QD, Shengjuler D, Fletcher SJ, O'Neal MC, Cai Y, Chang JCJ, Broadhurst DJ, Klippsten S, Sharp PP, Wenzell NA, Kuzuoglu-Ozturk D, Wang HY, Trenker R, Young JM, Cavero DA, Hiatt J, Roth TL, Rathore U, Subramanian A, Noack J, Hubert M, Stroud RM, Frankel AD, Rosenberg OS, Verba KA, Agard DA, Ott M, Emerman M, Jura N, von Zastrow M, Verdin E, Ashworth A, Schwartz O, d'Enfert C, Mukherjee S, Jacobson M, Malik HS, Fujimori DG, Ideker T, Craik CS, Floor SN, Fraser JS, Gross JD, Sali A, Roth BL, Ruggero D, Taunton J, Kortemme T, Beltrao P, Vignuzzi M, García-Sastre A, Shokat KM, Shoichet BK, Krogan NJ. A SARS-CoV-2 protein interaction map reveals targets for drug repurposing. Nature. 2020 07; 583(7816):459-468.  View on PubMed
  9. Petronikolou N, Longbotham JE, Fujimori DG. Dissecting contributions of catalytic and reader domains in regulation of histone demethylation. Methods Enzymol. 2020; 639:217-236.  View on PubMed
  10. Gordon DE, Jang GM, Bouhaddou M, Xu J, Obernier K, O'Meara MJ, Guo JZ, Swaney DL, Tummino TA, Hüttenhain R, Kaake RM, Richards AL, Tutuncuoglu B, Foussard H, Batra J, Haas K, Modak M, Kim M, Haas P, Polacco BJ, Braberg H, Fabius JM, Eckhardt M, Soucheray M, Bennett MJ, Cakir M, McGregor MJ, Li Q, Naing ZZC, Zhou Y, Peng S, Kirby IT, Melnyk JE, Chorba JS, Lou K, Dai SA, Shen W, Shi Y, Zhang Z, Barrio-Hernandez I, Memon D, Hernandez-Armenta C, Mathy CJP, Perica T, Pilla KB, Ganesan SJ, Saltzberg DJ, Ramachandran R, Liu X, Rosenthal SB, Calviello L, Venkataramanan S, Lin Y, Wankowicz SA, Bohn M, Trenker R, Young JM, Cavero D, Hiatt J, Roth T, Rathore U, Subramanian A, Noack J, Hubert M, Roesch F, Vallet T, Meyer B, White KM, Miorin L, Agard D, Emerman M, Ruggero D, García-Sastre A, Jura N, von Zastrow M, Taunton J, Schwartz O, Vignuzzi M, d'Enfert C, Mukherjee S, Jacobson M, Malik HS, Fujimori DG, Ideker T, Craik CS, Floor S, Fraser JS, Gross J, Sali A, Kortemme T, Beltrao P, Shokat K, Shoichet BK, Krogan NJ. A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug-Repurposing. bioRxiv. 2020 Mar 22.  View on PubMed
  11. Stojkovic V, Myasnikov AG, Young ID, Frost A, Fraser JS, Fujimori DG. Assessment of the nucleotide modifications in the high-resolution cryo-electron microscopy structure of the Escherichia coli 50S subunit. Nucleic Acids Res. 2020 03 18; 48(5):2723-2732.  View on PubMed
  12. Petronikolou N, Longbotham JE, Fujimori DG. Extended Recognition of the Histone H3 Tail by Histone Demethylase KDM5A. Biochemistry. 2020 02 11; 59(5):647-651.  View on PubMed
  13. Stojkovic V, Ulate MF, Hidalgo-Villeda F, Aguilar E, Monge-Cascante C, Pizarro-Guajardo M, Tsai K, Tzoc E, Camorlinga M, Paredes-Sabja D, Quesada-Gómez C, Fujimori DG, Rodríguez C. cfr(B), cfr(C), and a New cfr-Like Gene, cfr(E), in Clostridium difficile Strains Recovered across Latin America. Antimicrob Agents Chemother. 2019 12 20; 64(1).  View on PubMed
  14. Longbotham JE, Chio CM, Dharmarajan V, Trnka MJ, Torres IO, Goswami D, Ruiz K, Burlingame AL, Griffin PR, Fujimori DG. Histone H3 binding to the PHD1 domain of histone demethylase KDM5A enables active site remodeling. Nat Commun. 2019 01 09; 10(1):94.  View on PubMed
  15. Stojkovic V, Chu T, Therizols G, Weinberg DE, Fujimori DG. miCLIP-MaPseq, a Substrate Identification Approach for Radical SAM RNA Methylating Enzymes. J Am Chem Soc. 2018 06 13; 140(23):7135-7143.  View on PubMed
  16. Stojkovic V, Fujimori DG. Mutations in RNA methylating enzymes in disease. Curr Opin Chem Biol. 2017 Dec; 41:20-27.  View on PubMed
  17. Fitzsimmons CM, Fujimori DG. Determinants of tRNA Recognition by the Radical SAM Enzyme RlmN. PLoS One. 2016; 11(11):e0167298.  View on PubMed
  18. Stojkovic V, Noda-Garcia L, Tawfik DS, Fujimori DG. Antibiotic resistance evolved via inactivation of a ribosomal RNA methylating enzyme. Nucleic Acids Res. 2016 Oct 14; 44(18):8897-8907.  View on PubMed
  19. Fujimori DG, Conway SJ. Editorial overview: Chemical genetics and epigenetics. Curr Opin Chem Biol. 2016 08; 33:vi-vii.  View on PubMed
  20. Liu YC, Fujimori DG, Weissman JS. Htm1p-Pdi1p is a folding-sensitive mannosidase that marks N-glycoproteins for ER-associated protein degradation. Proc Natl Acad Sci U S A. 2016 07 12; 113(28):E4015-24.  View on PubMed

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