Danica Galonic Fujimori, PhD
Professor
Cellular Molecular Pharmacology
School of Medicine

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.

• First-in-Class Allosteric Inhibitors of DNMT3A Disrupt Protein-Protein Interactions and Induce Acute Myeloid Leukemia Cell Differentiation.
  Journal of medicinal chemistry 2022 Sandoval JE, Ramabadran R, Stillson N, Sarah L, Fujimori DG, Goodell MA, Reich N
• Viral E protein neutralizes BET protein-mediated post-entry antagonism of SARS-CoV-2.
  Cell reports 2022 Chen IP, Longbotham JE, McMahon S, Suryawanshi RK, Khalid MM, Taha TY, Tabata T, Hayashi JM, Soveg FW, Carlson-Stevermer J, Gupta M, Zhang MY, Lam VL, Li Y, Yu Z, Titus EW, Diallo A, Oki J, Holden K, …
• Covalent labeling of a chromatin reader domain using proximity-reactive cyclic peptides.
  Chemical science 2022 Zhang MY, Yang H, Ortiz G, Trnka MJ, Petronikolou N, Burlingame AL, DeGrado WF, Fujimori DG
• Structural basis for context-specific inhibition of translation by oxazolidinone antibiotics.
  Nature structural & molecular biology 2022 Tsai K, Stojkovic V, Lee DJ, Young ID, Szal T, Klepacki D, Vázquez-Laslop N, Mankin AS, Fraser JS, Fujimori DG
• Directed evolution of the rRNA methylating enzyme Cfr reveals molecular basis of antibiotic resistance.
  eLife 2022 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
• Recognition of Histone H3 Methylation States by the PHD1 Domain of Histone Demethylase KDM5A.
  ACS chemical biology 2021 Longbotham JE, Kelly MJS, Fujimori DG
• miCLIP-MaPseq Identifies Substrates of Radical SAM RNA-Methylating Enzyme Using Mechanistic Cross-Linking and Mismatch Profiling.
  Methods in molecular biology (Clifton, N.J.) 2021 Stojkovic V, Weinberg DE, Fujimori DG
• Exploring the Ligand Preferences of the PHD1 Domain of Histone Demethylase KDM5A Reveals Tolerance for Modifications of the Q5 Residue of Histone 3.
  ACS chemical biology 2020 Anderson SE, Longbotham JE, O'Kane PT, Ugur FS, Fujimori DG, Mrksich M
• Domain cross-talk in regulation of histone modifications: Molecular mechanisms and targeting opportunities.
  Current opinion in chemical biology 2020 Longbotham JE, Zhang MY, Fujimori DG
• A SARS-CoV-2 protein interaction map reveals targets for drug repurposing.
  Nature 2020 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, …
• Dissecting contributions of catalytic and reader domains in regulation of histone demethylation.
  Methods in enzymology 2020 Petronikolou N, Longbotham JE, Fujimori DG
• A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug-Repurposing.
  bioRxiv : the preprint server for biology 2020 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, …
• Assessment of the nucleotide modifications in the high-resolution cryo-electron microscopy structure of the Escherichia coli 50S subunit.
  Nucleic acids research 2020 Stojkovic V, Myasnikov AG, Young ID, Frost A, Fraser JS, Fujimori DG
• Extended Recognition of the Histone H3 Tail by Histone Demethylase KDM5A.
  Biochemistry 2020 Petronikolou N, Longbotham JE, Fujimori DG
• cfr(B), cfr(C), and a New cfr-Like Gene, cfr(E), in Clostridium difficile Strains Recovered across Latin America.
  Antimicrobial agents and chemotherapy 2019 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
• Histone H3 binding to the PHD1 domain of histone demethylase KDM5A enables active site remodeling.
  Nature communications 2019 Longbotham JE, Chio CM, Dharmarajan V, Trnka MJ, Torres IO, Goswami D, Ruiz K, Burlingame AL, Griffin PR, Fujimori DG
• miCLIP-MaPseq, a Substrate Identification Approach for Radical SAM RNA Methylating Enzymes.
  Journal of the American Chemical Society 2018 Stojkovic V, Chu T, Therizols G, Weinberg DE, Fujimori DG
• Mutations in RNA methylating enzymes in disease.
  Current opinion in chemical biology 2017 Stojkovic V, Fujimori DG
• Determinants of tRNA Recognition by the Radical SAM Enzyme RlmN.
  PloS one 2016 Fitzsimmons CM, Fujimori DG
• Antibiotic resistance evolved via inactivation of a ribosomal RNA methylating enzyme.
  Nucleic acids research 2016 Stojkovic V, Noda-Garcia L, Tawfik DS, Fujimori DG
• Editorial overview: Chemical genetics and epigenetics.
  Current opinion in chemical biology 2016 Fujimori DG, Conway SJ
• Htm1p-Pdi1p is a folding-sensitive mannosidase that marks N-glycoproteins for ER-associated protein degradation.
  Proceedings of the National Academy of Sciences of the United States of America 2016 Liu YC, Fujimori DG, Weissman JS
• Opposing Chromatin Signals Direct and Regulate the Activity of Lysine Demethylase 4C (KDM4C).
  The Journal of biological chemistry 2016 Pack LR, Yamamoto KR, Fujimori DG
• Docking and Linking of Fragments To Discover Jumonji Histone Demethylase Inhibitors.
  Journal of medicinal chemistry 2015 Korczynska M, Le DD, Younger N, Gregori-Puigjané E, Tumber A, Krojer T, Velupillai S, Gileadi C, Nowak RP, Iwasa E, Pollock SB, Ortiz Torres I, Oppermann U, Shoichet BK, Fujimori DG
• Functional coupling between writers, erasers and readers of histone and DNA methylation.
  Current opinion in structural biology 2015 Torres IO, Fujimori DG
• Improved Peak Detection and Deconvolution of Native Electrospray Mass Spectra from Large Protein Complexes.
  Journal of the American Society for Mass Spectrometry 2015 Lu J, Trnka MJ, Roh SH, Robinson PJ, Shiau C, Fujimori DG, Chiu W, Burlingame AL, Guan S
• Radical SAM-Mediated Methylation of Ribosomal RNA.
  Methods in enzymology 2015 Stojkovic V, Fujimori DG
• Histone demethylase KDM5A is regulated by its reader domain through a positive-feedback mechanism.
  Nature communications 2015 Torres IO, Kuchenbecker KM, Nnadi CI, Fletterick RJ, Kelly MJ, Fujimori DG
• Product binding enforces the genomic specificity of a yeast polycomb repressive complex.
  Cell 2014 Dumesic PA, Homer CM, Moresco JJ, Pack LR, Shanle EK, Coyle SM, Strahl BD, Fujimori DG, Yates JR, Madhani HD
• Radical SAM-mediated methylation reactions.
  Current opinion in chemical biology 2013 Fujimori DG
• Reconstitution of nucleosome demethylation and catalytic properties of a Jumonji histone demethylase.
  Chemistry & biology 2013 Shiau C, Trnka MJ, Bozicevic A, Ortiz Torres I, Al-Sady B, Burlingame AL, Narlikar GJ, Fujimori DG
• Site-specific and regiospecific installation of methylarginine analogues into recombinant histones and insights into effector protein binding.
  Journal of the American Chemical Society 2013 Le DD, Cortesi AT, Myers SA, Burlingame AL, Fujimori DG
• Covalent Intermediate in the Catalytic Mechanism of the Radical S-Adenosyl-l-methionine Methyl Synthase RlmN Trapped by Mutagenesis
  J Am Chem Soc 2012 McCusker KP, Medzihradszky KF, Shiver AL, Nichols RJ, Yan F, Maltby DA, Gross CA, Fujimori DG.
• Covalent intermediate in the catalytic mechanism of the radical S-adenosyl-L-methionine methyl synthase RlmN trapped by mutagenesis.
  Journal of the American Chemical Society 2012 McCusker KP, Medzihradszky KF, Shiver AL, Nichols RJ, Yan F, Maltby DA, Gross CA, Fujimori DG
• Protein and nucleic acid methylating enzymes: mechanisms and regulation.
  Current opinion in chemical biology 2012 Le DD, Fujimori DG
• The chemistry of peptidyltransferase center-targeted antibiotics: enzymatic resistance and approaches to countering resistance.
  ACS chemical biology 2011 McCusker KP, Fujimori DG
• RNA methylation by radical SAM enzymes RlmN and Cfr proceeds via methylene transfer and hydride shift.
  Proceedings of the National Academy of Sciences of the United States of America 2011 Yan F, Fujimori DG
• A novel enzymatic rearrangement.
  Chemistry & biology 2010 Fujimori DG
• RlmN and Cfr are radical SAM enzymes involved in methylation of ribosomal RNA.
  Journal of the American Chemical Society 2010 Yan F, LaMarre JM, Röhrich R, Wiesner J, Jomaa H, Mankin AS, Fujimori DG
• Structural analysis of an open active site conformation of nonheme iron halogenase CytC3.
  Journal of the American Chemical Society 2009 Wong C, Fujimori DG, Walsh CT, Drennan CL
• Hypoxia sensing goes gauche.
  Nature chemical biology 2009 Fujimori DG
• Halogenation strategies in natural product biosynthesis.
  Chemistry & biology 2008 Neumann CS, Fujimori DG, Walsh CT
• CD and MCD of CytC3 and taurine dioxygenase: role of the facial triad in alpha-KG-dependent oxygenases.
  Journal of the American Chemical Society 2007 Neidig ML, Brown CD, Light KM, Fujimori DG, Nolan EM, Price JC, Barr EW, Bollinger JM, Krebs C, Walsh CT, Solomon EI
• Spectroscopic evidence for a high-spin Br-Fe(IV)-oxo intermediate in the alpha-ketoglutarate-dependent halogenase CytC3 from Streptomyces.
  Journal of the American Chemical Society 2007 Fujimori DG, Barr EW, Matthews ML, Koch GM, Yonce JR, Walsh CT, Bollinger JM, Krebs C, Riggs-Gelasco PJ
• Cloning and characterization of the biosynthetic gene cluster for kutznerides.
  Proceedings of the National Academy of Sciences of the United States of America 2007 Fujimori DG, Hrvatin S, Neumann CS, Strieker M, Marahiel MA, Walsh CT
• What's new in enzymatic halogenations.
  Current opinion in chemical biology 2007 Fujimori DG, Walsh CT
• Chemical glycosylation in the synthesis of glycoconjugate antitumour vaccines.
  Nature 2007 Galonic DP, Gin DY
• Characterization of the aminocarboxycyclopropane-forming enzyme CmaC.
  Biochemistry 2007 Kelly WL, Boyne MT, Yeh E, Vosburg DA, Galonic DP, Kelleher NL, Walsh CT
• Two interconverting Fe(IV) intermediates in aliphatic chlorination by the halogenase CytC3.
  Nature chemical biology 2007 Galonic DP, Barr EW, Walsh CT, Bollinger JM, Krebs C
• Enzymatic generation of the antimetabolite gamma,gamma-dichloroaminobutyrate by NRPS and mononuclear iron halogenase action in a streptomycete.
  Chemistry & biology 2006 Ueki M, Galonic DP, Vaillancourt FH, Garneau-Tsodikova S, Yeh E, Vosburg DA, Schroeder FC, Osada H, Walsh CT
• Halogenation of unactivated carbon centers in natural product biosynthesis: trichlorination of leucine during barbamide biosynthesis.
  Journal of the American Chemical Society 2006 Galonic DP, Vaillancourt FH, Walsh CT
• Aziridine-2-carboxylic acid-containing peptides: application to solution- and solid-phase convergent site-selective peptide modification.
  Journal of the American Chemical Society 2005 Galonic DP, Ide ND, van der Donk WA, Gin DY
• Site-selective conjugation of thiols with aziridine-2-carboxylic acid-containing peptides.
  Journal of the American Chemical Society 2004 Galonic DP, van der Donk WA, Gin DY
• Oligosaccharide-peptide ligation of glycosyl thiolates with dehydropeptides: synthesis of S-linked mucin-related glycopeptide conjugates.
  Chemistry (Weinheim an der Bergstrasse, Germany) 2003 Galonic DP, Van Der Donk WA, Gin DY
• Selenocysteine derivatives for chemoselective ligations.
  Chembiochem : a European journal of chemical biology 2002 Gieselman MD, Zhu Y, Zhou H, Galonic D, van der Donk WA
• J Serb Chem Soc
  Model Study of Epothilone Synthesis: An Alternative Synthetic Approach to the C1-C7 fragment 2002 Lalic G, Galonic D, Matovic R, Saicic RN
• Tetrahedron
  Alkylation of Carbonyl Compounds in the TiCl4-promoted Reaction of Trimethylsilyl Enol Ethers with Epoxides 2001 Lalic G, Petrovski Z, Galonic DP, Matovic R, Saicic RN