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P2X7 receptor-dependent and -independent T cell death is induced by nicotinamide adenine dinucleotide
Hiroki Kawamura1, Fred Aswad, Masahiro Minagawa
1Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, CA 90033, USA.
Abstract:
Adding NAD to murine T lymphocytes inhibits their functions and induces annexin V binding. This report shows that NAD induces cell death in a subset of T cells within seconds whereas others do not die until many hours later. Low NAD concentrations (<10 microM) suffice to trigger rapid cell death, which is associated with annexin V binding and membrane pore formation, is not blocked by the caspase inhibitor Z-VADfmk, and requires functional P2X7 receptors. The slower induction of death requires higher NAD concentrations (>100 microM), is blocked by caspase inhibitor Z-VADfmk, is associated with DNA fragmentation, and does not require P2X7 receptors. T cells degrade NAD to ADP-ribose (ADPR), and adding ADPR to T cells leads to slow but not rapid cell death. NAD but not ADPR provides the substrate for ADP-ribosyltransferase (ART-2)-mediated attachment of ADP-ribosyl groups to cell surface proteins; expression of ART-2 is required for NAD to trigger rapid but not slow cell death. These results support the hypothesis that cell surface ART-2 uses NAD but not ADPR to attach ADP-ribosyl groups to the cell surface, and that these groups act as ligands for P2X7 receptors that then induce rapid cell death. Adding either NAD or ADPR also triggers a different set of mechanisms, not requiring ART-2 or P2X7 receptors that more slowly induce cell death.
Insights
Nicotinamide adenine dinucleotide (NAD) rapidly kills T cells via P2X7 receptors and ART-2, while higher concentrations induce slower, caspase-dependent cell death. This reveals distinct NAD-mediated cell death pathways in T lymphocytes.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Nicotinamide adenine dinucleotide (NAD) plays crucial roles in cellular metabolism and signaling.
- T lymphocytes are key components of the adaptive immune system.
- Mechanisms of NAD-induced cell death in immune cells are not fully elucidated.
Purpose of the Study:
- To investigate the distinct mechanisms by which NAD induces cell death in murine T lymphocytes.
- To identify the roles of P2X7 receptors, caspase activation, and ADP-ribosyltransferase-2 (ART-2) in NAD-mediated T cell death.
Main Methods:
- Treatment of murine T lymphocytes with varying concentrations of NAD and ADP-ribose (ADPR).
- Assessment of cell viability, annexin V binding, membrane integrity, and DNA fragmentation.
- Use of caspase inhibitor Z-VADfmk and evaluation of P2X7 receptor and ART-2 dependency.
Main Results:
- Low NAD concentrations (<10 microM) induced rapid, caspase-independent cell death via P2X7 receptors and ART-2.
- Higher NAD concentrations (>100 microM) triggered slower, caspase-dependent cell death with DNA fragmentation, independent of P2X7.
- NAD, but not ADPR, served as a substrate for ART-2 in rapid cell death induction.
Conclusions:
- NAD induces T cell death through at least two distinct pathways: rapid P2X7/ART-2-mediated and slower caspase-dependent mechanisms.
- ART-2 acts as a cell surface receptor utilizing NAD to initiate rapid T cell death.
- These findings provide novel insights into the immunomodulatory functions of NAD and its metabolites.
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