John M. Denu
Honors & Awards
The laboratory investigates the mechanism and biological function of reversible protein modifications involved in modulating signal transduction, chromatin dynamics and gene activation. Histones are one of the best examples of protein function regulated by modification, and are the target of an array of modifications including acetylation, phosphorylation and methylation. The dynamic and specific nature of these modifications has led to the proposal that there is a “Histone Code”, which when decoded would allow us to understand, for instance, what is required to maintain “silenced” or “active” chromatin. In general, histone acetylation (catalyzed by histone acetyltransferases) correlates with gene transcription, while hypo-acetylation (catalyzed by protein deacetylases) correlates with repression. To begin to understand this signaling code, we must first understand the mechanisms and regulation of the enzymes responsible for these modifications. A large component of our work is to understand the major enzyme families that catalyze these reactions.
Chromatin remodeling enzymes rely on co-enzymes derived from metabolic pathways, suggesting coordination between nuclear events and metabolic networks. A unique example of this link is the recently described NAD+-dependent protein/histone deacetylases. The founding member of this family, yeast Sir2 (silent information regulator 2), is involved in gene silencing, chromosomal stability, and ageing. Sir2-like enzymes catalyze a reaction in which the cleavage of NAD+, and histone/protein deacetylation are coupled to the formation of O-acetyl-ADP-ribose, a novel metabolite. The dependence on NAD+ and the generation of this potential second messenger offer new clues in understanding the function and regulation for nuclear, cytoplasmic and mitochondrial Sir2-like enzymes. Questions we are currently attempting to answer with this project: What is the physiological basis for the NAD+ -dependence? What are the cellular, acetylated-protein substrates for the various Sir2 homologues? What is the function of O-acetyl-ADP-ribose? What is the relationship between Sir2 enzyme function and metabolism? How are Sir2 enzymes regulated? How does Sir2 catalyze this unique reaction? To address these questions we are using a breadth of approaches that involve biochemistry, genetics, proteomics, enzymology, and use of a mammalian tissue culture system and yeast as a genetically tractable system to explore biological function.
Publications since 2007
Garske, A. L.; Oliver, S. S.; Wagner, E. K.; Musselman, C. A.; LeRoy, G.; Garcia, B. A.; Kutateladze, T. G.; Denu, J. M., Combinatorial profiling of chromatin binding modules reveals multisite discrimination. Nat Chem Biol 2010, 6 (4), 283-90.
Tong, L.; Denu, J. M., Function and metabolism of sirtuin metabolite O-acetyl-ADP-ribose. Biochim Biophys Acta 2010, 1804 (8), 1617-1625.
Arun, P.; Madhavarao, C. N.; Moffett, J. R.; Hamilton, K.; Grunberg, N. E.; Ariyannur, P. S.; Gahl, W. A.; Anikster, Y.; Mog, S.; Hallows, W. C.; Denu, J. M.; Namboodiri, A. M., Metabolic acetate therapy improves phenotype in the tremor rat model of Canavan disease. J Inherit Metab Dis 2010, 33 (3), 195-210.
Ariyannur, P. S.; Moffett, J. R.; Madhavarao, C. N.; Arun, P.; Vishnu, N.; Jacobowitz, D. M.; Hallows, W. C.; Denu, J. M.; Namboodiri, A. M., Nuclear-cytoplasmic localization of acetyl coenzyme a synthetase-1 in the rat brain. J Comp Neurol 2010, 518 (15), 2952-77.
Albaugh, B. N.; Kolonko, E. M.; Denu, J. M., Kinetic Mechanism of the Rtt109-Vps75 Histone Acetyltransferase-Chaperone Complex. Biochemistry 2010, Aug 3;49(30):6375-85.
Tong, L.; Lee, S.; Denu, J. M., Hydrolase regulates NAD+ metabolites and modulates cellular redox. J Biol Chem 2009, 284 (17), 11256-66.
Smith, B. C.; Hallows, W. C.; Denu, J. M., A continuous microplate assay for sirtuins and nicotinamide-producing enzymes. Anal Biochem 2009, 394 (1), 101-9.
Smith, B. C.; Denu, J. M., Chemical mechanisms of histone lysine and arginine modifications. Biochim Biophys Acta 2009, 1789 (1), 45-57.
Musselman, C. A.; Mansfield, R. E.; Garske, A. L.; Davrazou, F.; Kwan, A. H.; Oliver, S. S.; O'Leary, H.; Denu, J. M.; Mackay, J. P.; Kutateladze, T. G., Binding of the CHD4 PHD2 finger to histone H3 is modulated by covalent modifications. Biochem J 2009, 423 (2), 179-87.
Hallows, W. C.; Smith, B. C.; Lee, S.; Denu, J. M., Ure(k)a! Sirtuins Regulate Mitochondria. Cell 2009, 137 (3), 404-6.
Smith, B. C.; Hallows, W. C.; Denu, J. M., Mechanisms and molecular probes of sirtuins. Chem Biol 2008, 15 (10), 1002-13.
Milne, J. C.; Denu, J. M., The Sirtuin family: therapeutic targets to treat diseases of aging. Curr Opin Chem Biol 2008, 12 (1), 11-7.
Lee, S.; Tong, L.; Denu, J. M., Quantification of endogenous sirtuin metabolite O-acetyl-ADP-ribose. Anal Biochem 2008, 383 (2), 174-9.
Smith, B. C.; Denu, J. M., Mechanism-based inhibition of Sir2 deacetylases by thioacetyl-lysine peptide. Biochemistry 2007, 46 (50), 14478-86.
Smith, B. C.; Denu, J. M., Acetyl-lysine analog peptides as mechanistic probes of protein deacetylases. J Biol Chem 2007, 282 (51), 37256-65.
Garske, A. L.; Smith, B. C.; Denu, J. M., Linking SIRT2 to Parkinson's disease. ACS Chem Biol 2007, 2 (8), 529-32.
Comstock, L. R.; Denu, J. M., Synthesis and biochemical evaluation of O-acetyl-ADP-ribose and N-acetyl analogs. Org Biomol Chem 2007, 5 (19), 3087-91.
Denu JM. Vitamins and aging: pathways to NAD+ synthesis. Cell. 2007 May 4;129(3):453-4.
Smith BC, Denu JM. Sir2 deacetylases exhibit nucleophilic participation of acetyl-lysine in NAD+ cleavage. J Am Chem Soc. 2007 May 9;129(18):5802-3.
Tsubota T, Berndsen CE, Erkmann JA, Smith CL, Yang L, Freitas MA, Denu JM, Kaufman PD. Histone H3-K56 acetylation is catalyzed by histone chaperone-dependent complexes. Mol Cell. 2007 Mar 9;25(5):703-12.
Berndsen CE, Selleck W, McBryant SJ, Hansen JC, Tan S, Denu JM. Nucleosome recognition by the Piccolo NuA4 histone acetyltransferase complex. Biochemistry. 2007 Feb 27;46(8):2091-9.
Berndsen CE, Albaugh BN, Tan S, Denu JM. Catalytic mechanism of a MYST family histone acetyltransferase. Biochemistry. 2007 Jan 23;46(3):623-9.
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of Wisconsin - Department
of Biomolecular Chemistry