Targeted Disruption of CD38 Accelerates Autoimmune Diabetes in NOD/Lt Mice by Enhancing Autoimmunity in an ADP-Ribosyltransferase 2-Dependent Fashion 1 Jing Chen,* Yi-Guang Chen,* Peter C. Reifsnyder,* William H. Schott,* Chul-Ho Lee, † Melissa Osborne,* Felix Scheuplein, ‡ Friedrich Haag, ‡ Friedrich Koch-Nolte, ‡ David V. Serreze,* and Edward H. Leiter 2 * Ubiquitously expressed CD38 and T cell-expressed ADP-ribosyltransferase 2 (ART2) are ectoenzymes competing for NAD sub- strate. CD38 exerts pleiotropic actions in hemopoietic and nonhemopoietic compartments via effects on calcium mobilization. ART2 is an ADP-ribosyltransferase on naive CD4  and CD8  T cells. ART2-catalyzed ADP-ribosylation of the P2X7 purinore- ceptor elicits apoptosis. Transfer of a genetically disrupted CD38 allele into the autoimmune diabetes-prone NOD/Lt background accelerated diabetes onset in both sexes, whereas transfer of a disrupted ART2 complex had no effect. However, the fact that the accelerated pathogenesis mediated by CD38 deficiency required ART2 activity was demonstrated by combining both ART2 and CD38 deficiencies. Reciprocal bone marrow reconstitution studies demonstrated accelerated diabetes only when CD38-deficient bone marrow was transferred into CD38-deficient recipients. Neither decreases in  cell function nor viability were indicated. Rather, the balance between T-effectors and T-regulatory cells was disturbed in CD38-deficient but ART2-intact NOD mice. In these mice, significant reductions in total viable CD8  T cells were observed. This was accompanied by an age-dependent increase in a diabetogenic CD8 clonotype. This in turn correlated with impaired T-regulatory development (10-fold reduction in Foxp3 mRNA expression). These changes were corrected when CD38 deficiency was combined with ART2 deficiency. Both ART2- deficient and CD38/ART2 combined deficient T cells were resistant to NAD-induced killing in vitro, whereas CD38-deficient but ART2-intact T cells showed increased sensitivity, particularly the CD4  CD25  subset. Unexpectedly, diabetes development in the combined CD38/ART2 stock was strongly suppressed, possibly through epistatic interactions between genes linked to the targeted CD38 on Chromosome 5 and the ART2 complex on Chromosome 7. The Journal of Immunology, 2006, 176: 4590 – 4599. C D38 (ADP ribosyl cyclase/cADPR hydrolase) is a NAD using ectoenzyme ubiquitously expressed to varying de- grees on both hemopoietic and nonhemopoietic tissues. The cyclase activity of this enzyme converts NAD to cyclic ADP- ribose (cADPR), 3 which is further hydrolyzed to ADPR by the enzyme’s hydrolase activity (1). CD38 can also catalyze the ex- change of the nicotinamide group of NADP  with nicotinic acid, producing nicotinic acid adenine dinucleotide phosphate (NAADP) (2). Both cADPR and NAADP mobilize Ca 2 from internal calcium stores through the ryanodine receptor, which is a different mobilization mechanism than that mediated through the inositol triphosphate pathway (3). CD38 reportedly binds CD31/ PECAM-1 to promote human monocyte-derived dendritic cell (DC) maturation, proliferation, and Ag-presenting function (4). Genetic disruption of CD38 in mice on a mixed genetic back- ground reportedly produced severe impairment of glucose toler- ance (5, 6). Impaired  cell insulin secretion was associated with absence of cADPR, a CD38 product to mobilize calcium in re- sponse to glucose (6). However, independent targeting of the CD38 gene (7) followed by 10 backcrosses to the defined C57BL/6J (B6) background failed to impair glucose tolerance (8). Thus, the reported impairment of glucose tolerance and  cell function very likely reflected the influence of genetic background rather than CD38 deficiency alone (8). In the immune system, CD38 deficiency has been associated with defects in B lymphocyte receptor-stimulated Ig production (7), neutrophil migration (9), DC migration (10), and T cell signaling via direct interaction with the Lck Src homology 2 domain (11). High CD38 expression on a CD4 T cell subset has also been associated with immunoregulatory function (12). Another NAD using ectoenzyme, ADP-ribosyltransferase 2 (ART2), competes with CD38 for NAD substrate (13). ART2 is a GPI-anchored enzyme on resting/naive mouse T cells that transfers the ADP-ribose moiety from NAD to target proteins, including CD8 and LFA-1 (14, 15). P2X7, a purinoceptor on T cell surfaces that serve as another ART2 target protein, is activated by ADP- ribosylation (16). This generates a nonselective pore on the T cell surface, triggering calcium influx, and finally leading to apoptosis if extracellular NAD concentrations are sufficiently high (16, 17). *The Jackson Laboratory, Bar Harbor, ME 04609; † Korea Research Institute of Bio- science and Biotechnology, Daejeon, Korea; and ‡ Institute of Immunology, Univer- sity Hospital, Hamburg, Germany Received for publication October 21, 2005. Accepted for publication January 24, 2006. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 This work was supported by National Institutes of Health Grants DK27722 and DK36175 (to E.H.L.), DK46266 and DK51090 (to D.V.S.), and Deutsche For- schungsgemeinschaft Grant No310/6 (to F.K.-N. and F.H.). J.C. is a recipient of the Francis M. Sherwin Endowed Fellowship at The Jackson Laboratory. Y.-G.C. is a recipient of the Juvenile Diabetes Research Foundation postdoctoral fellowship. The stay of F.S. in the E.H.L.’s laboratory was supported by a stipend from the Boehringer Ingelheim Foundation. Institutional shared services were supported by National Can- cer Institute Center Support Grant CA-34196. 2 Address correspondence and reprint requests Dr. Edward H. Leiter, The Jackson Laboratory, Bar Harbor, ME 04609. E-mail address: ed.leiter@jax.org 3 Abbreviations used in this paper: cADPR, cyclic ADP-ribose; NAADP, nicotinic acid adenine dinucleotide phosphate; DC, dendritic cell; ART2, ADP-ribosyltrans- ferase 2; NICD,NAD-induced cell death; PLN, pancreatic lymph node; MLN, mes- enteric lymph node; PI, propidium iodide; Treg, regulatory T cell. The Journal of Immunology Copyright © 2006 by The American Association of Immunologists, Inc. 0022-1767/06/$02.00