David Boone, Ph.D.

Research Summary

Research in my laboratory is focused on the regulation of inflammation, especially as it relates to gastrointestinal disorders like Crohn's disease, ulcerative colitis and inflammatory liver diseases. We are interested in the role of innate and adaptive immune cells and their interaction with commensal microbes, pathogens and non-immune cells of the gastrointestinal tract. Our approach to study these diseases involves the generation of mouse models through targeted genetic mutations ("knockout mice"). These mouse models are used for experiments ranging from classic cellular immunology to physiology to investigate the roles of specific gene products in the regulation of inflammation and gastrointestinal function. In addition, cells derived from these mice are used to elucidate the fundamental mechanisms involved in the regulation of inflammation. These studies focus on the role of a specific protein in the regulation of cellular functions like proliferation, death or signal transduction and the underlying biochemical processes that mediate these events. Once the cellular and biochemical mechanisms involved in the regulation of inflammation by a specific gene product are understood we then complement stem cells with vectors carrying mutant or normal versions of that gene. These studies serve to demonstrate the relevance of a cellular or biochemical mechanism to the regulation inflammatory disease and allow for the examination of the roles of specific subsets of immune and non-immune cells to the disease process.

As an example of this approach, we are studying the role of the gene A20 in inflammatory diseases. A20 knockout (A20-/-) mice develop inflammation in the intestine and liver. Bone marrow chimera (A20-/- immune cells in a normal mouse) also develop inflammation in the gut and liver. Immune and non-immune cells derived from A20-/- mice display exaggerated inflammatory responses to TNF. These cells also display excessive inflammatory responses to microbial products like LPS, demonstrating the role of A20 in the regulation of Toll-like receptor (TLR) signaling. Examination of the signal transduction pathways in these cells reveals that A20 is required for the termination of at least two pro-inflammatory kinases activated by TNF and TLR signals. Biochemical studies with recombinant A20 protein demonstrate that A20 is a de-ubiquitylating enzyme and that its enzymatic activity depends on a critical cysteine residue at position 103 (C103) of the protein. The de-ubiquitylation, by A20, of key signal transduction proteins upstream of the pro-inflammatory kinases is required for terminating responses to TNF and TLR ligands. Complementation of A20, but not a C103A mutant A20, restores A20-/- cells to a normal inflammatory response to TNF and LPS. We are currently complementing A20-/- hematopoietic stem cells with A20, and the C103A mutant A20, to determine the role of A20's de-ubiquitylating activity in immune cells in the prevention of inflammation in the liver and intestine. Finally, other A20-like proteins that have domains consistent with a de-ubiquitylating enzyme activity are proposed, based on the mouse genome. These genes will also be targeted to assess the overall role of A20-like proteins and de-ubiquitylation in other models of inflammation and disease.

Selected Papers

Lodolce JP, Boone DL, Chai S, Swain RE, Dassopoulos T, Trettin S and Ma A (1998). IL-15 receptor maintains lymphoid homeostasis by supporting lymphocyte homing and proliferation. Immunity. 9:669-676

Lee EG*, Boone DL*, Chai S, Libby SL, Chien M, Lodolce JP, Ma A (2000). Failure to regulate TNF-induced NF-kappaB and cell death responses in A20-deficient mice. Science 289:2350-2354 (co-first authors).

Burkett PR, Koka RM, Lodolce JP, Chien MC, Chan F, Ma A and Boone DL. (2003). IL-15Ra expression on CD8 T cells is dispensable for T cell memory. Proceedings of the National Academy of Sciences USA. 100:4724-4729

Wertz IE, O'Rourke KM, Zhou H, Eby M, Aravind L, Seshagiri S, Wu P, Wiesmann C, Baker R, Boone DL, Ma A, Koonin EV and Dixit VM. (2004). De-ubiquitination and ubiquitin ligase domains of A20 downregulate NF-kappaB signaling . Nature. 430:694-699.

Boone DL*, Turer EE*, Lee EG*, Ahmad RC, Wheeler MT, Tsui C, Hurley P, Chien M, Chai S, Hitotsumatsu O, McNally E, Pickart C, and Ma A (2004). The ubiquitin modifying enzyme A20 is required for terminating Toll-like receptor responses. Nature Immunology. 5:1052-1050 (co-first authors)