Description of Research Expertise:
Our laboratory broadly focuses on DNA modifying enzymes and pathways, particularly those that contribute to genomic plasticity. We utilize a broad array of approaches, including biochemical characterization of enzyme mechanisms, chemical synthesis of enzyme probes, and biological assays spanning immunology and virology to study the fundamental question of how a genomic diversity arises in nature.
Mutation and modification of the genome play an important role in several physiologically relevant areas and our areas of interest include:
1. Decipher the molecular basis for deamination by AID/APOBEC enzymes and perturb deaminase immunological functions
From the host immune perspective, the generation of genomic diversity is used as both a defensive and an offensive weapon. Host mutator enzymes such as Activation-Induced Cytidine Deaminase (AID) seed diversity in the adaptive immune system by introducing targeted mutations into the immunoglobulin locus that result in antibody maturation. Related deaminases of the innate immune system can directly attack retroviral threats by garbling the pathogen genome through mutation, as accomplished by the deaminase APOBEC3G, which restricts infection with HIV. Immune mutator enzymes, however, also pose a risk to the host, as overexpression or dysregulation have been associated with oncogenesis.
2. Explore the interplay of cytosine modifying enzymes on DNA demethylation
The singular genome is responsible for a wealth of different cell types, each of which can respond and adapt to environmental cues. In part, these epigenetic differences are linked to DNA modification. These modifications center around cytosine, where DNA deamination (AID/APOBEC enzymes) , oxidation (TET family enzymes) and methylation (DNMTs) can all interplay and tune the genome's potential. We are interested in the enzymatic activities of these cytosine modifying enzymes, particularly in the process of DNA demethylation which plays a role in embryogenesis, gene regulation and a potential pathological role in cancer.
3. Target pathogen pathways that promote evolution and resistance.
From the pathogen perspective, alteration in key antigenic determinants at a rate that outpaces immune responses is a potent means for evasion. Further, rapid mutation may allow for the development of resistance to antimicrobials. In bacteria, adaptation and evolution are closely linked to the stress response of SOS pathway. The SOS pathway can be triggered by numerous stressors, including antibiotics, and the net result is accelerated acquisition of drug resistance. We aim to characterize the key regulatory and effector enzymes from the SOS pathway and to target the pathway as a means to combat antibiotic resistance.
Our research program aims to understand these pathways of purposeful DNA modification and mutation. Additionally, we apply chemical biology to decipher and target these pathways, to impede the development of multidrug-resistance in pathogens or prevent the neoplastic transformations that can result from genomic mutation.
Schutsky EK, DeNizio JE, Hu P, Liu MY, Nabel CS, Fabyanic EB, Hwang Y, Bushman FD, Wu H, Kohli RM: Nondestructive, base-resolution sequencing of 5-hydroxymethylcytosine using a DNA deaminase Nature Biotech 36 : 1083–1090,2018.
DeNizio JE, Liu MY, Leddin EM, Cisneros GA, Kohli RM: Selectivity and Promiscuity in TET-Mediated Oxidation of 5-Methylcytosine in DNA and RNA. Biochemistry 58 : 411-421,2019.
Samuels AN, Roggiani M, Zhu J, Goulian M, Kohli RM: The SOS Response Mediates Sustained Colonization of the Mammalian Gut. Infect Immun 87 : pii: e00711-18,2019.
Ghanty U, DeNizio JE, Liu MY, Kohli RM.: Exploiting Substrate Promiscuity to Develop Activity-Based Probes for TET Family Enzymes. J Am Chem Soc : doi: 10.1021/jacs.8b04722,2018.
Selwood T, Larsen BJ, Mo CY, Culyba MJ, Hostetler ZM, Kohli RM, Reitz AB, Baugh SDP.: Advancement of the 5-Amino-1-(Carbamoylmethyl)-1H-1,2,3-Triazole-4-Carboxamide Scaffold to Disarm the Bacterial SOS Response. Front Microbiol 9 : 2961,2018.
Hrit J, Goodrich L, Li C, Wang BA, Nie J, Cui X, Martin EA, Simental E, Fernandez J, Liu MY, Nery JR, Castanon R, Kohli RM, Tretyakova N, He C, Ecker JR, Goll M, Panning B: OGT binds a conserved C-terminal domain of TET1 to regulate TET1 activity and function in development. Elife 7 : pii: e34870,2018.
Hostetler ZM, Ferrie JJ, Bornstein MR, Sungwienwong I, Petersson EJ, Kohli RM: Systematic Evaluation of Soluble Protein Expression Using a Fluorescent Unnatural Amino Acid Reveals No Reliable Predictors of Tolerability ACS Chem Bio 13 (10): 2855-2861,2018.
Sungwienwong I, Ferrie JJ, Jun JV, Liu C, Barrett TM, Hostetler ZM, Ieda N, Hendricks A, Muthusamy AK, Kohli RM, Chenoweth DM, Petersson GA, Petersson EJ: Improving the fluorescent probe acridonylalanine through a combination of theory and experiment J Phys Org Chen : DOI: 10.1002/poc.3813,2018.
Fraietta JA, Nobles CL, Sammons MA, Lundh S, Carty SA, Reich TJ, Cogdill AP, Morrissette JJD, DeNizio JE, Reddy S, Hwang Y, Gohil M, Kulikovskaya I, Nazimuddin F, Gupta M, Chen F, Everett JK, Alexander KA, Lin-Shiao E, Gee MH, Liu X, Young RM, Ambrose D, Wang Y, Xu J, Jordan MS, Marcucci KT, Levine BL, Garcia KC, Zhao Y, Kalos M, Porter DL, Kohli RM, Lacey SF, Berger SL, Bushman FD, June CH, Melenhorst JJ: Disruption of TET2 Promotes the Therapeutic Efficacy of CD19-targeted T cells Nature 558 (7709): 307-312,2018.
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