Rapid induction of naive T cell apoptosis by ecto-nicotinamide adenine dinucleotide: Requirement for mono(ADP-ribosyl)Transferase 2 and a downstream effector

Lymphocytes express a number of NAD-metabolizing ectoenzymes, including mon o(ADP-ribosyl)transferases (ART) and ADP ribosylcyclases. These enzymes may regulate lymphocyte functions following the release of NAD in injured or i nflammatory tissues We report here that extracellular NAD induces apoptosis in BALB/c splenic T cells with an IC50 of 3-5 muM. Annexin V staining of c ells was observed already 10 min after treatment with NAD in the absence of any additional signal. Removal of GPI-anchored cell surface proteins by ph osphatidylinositol-specific phospholipase C treatment rendered cells resist ant to NAD-mediated apoptosis. RT-PCR analyses revealed that resting BALB/c T cells expressed the genes for GPI-anchored ART2.1 and ART2.2 but not ART 1. ART2-specific antisera blocked radiolabeling of cell surface proteins wi th both [P-32]NAD and NAD-mediated apoptosis. Further analyses revealed tha t natural knockout mice for Art2.a (C57BL/6) or Art2.b (NZW) were resistant to NAD-mediated apoptosis. Labeling with [P-32]NAD revealed strong cell su rface ART activity on T cells of C57BL/6 and little if any activity on cell s of NZW mice. T cells of (C57BL/6 x NZW)F-1 animals showed strong cell sur face ART activity and were very sensitive to NAD-induced apoptosis. As in B ALB/c T cells, ART2-specific antisera blocked cell surface ART activity and apoptosis in (C57BL/6 x NZW)F-1 T cells. The fact that T cells of F-1 anim als are sensitive to rapid NAD-induced apoptosis suggests that this effect requires the complementation of (at least) two genetic components. We propo se that one of these is cell surface ART2.2 activity (defective in the NZW parent), the other a downstream effector of ADP-ribosylation (defective in the C57BL/6 parent).