Ectocellular CD38-catalyzed synthesis and intracellular Ca2+-signalling activity of cyclic ADP-ribose in T-lymphocytes are not functionally related.
Cyclic ADP-ribose (cADPR) is a natural metabolite of beta-NAD+ with a potent Ca2+-mobilizing activity in different cell types, including T-lymphocytes. We investigated (i) whether stimulation of T-lymphocytes with different agonists affects the intracellular concentration of cADPR, and (ii) whether the lymphocyte antigen CD38, through its ectocellular ADP-ribosyl cyclase and cADPR-hydrolase enzymatic activities, can account for the regulation of the intracellular levels of cADPR and the Ca2+-mobilizing effects of this nucleotide in Jurkat and HPB.ALL T-lymphocytes. The anti-CD3 antibody OKT3, the sphingolipid sphingosine and lysophosphatidic acid induced an increase in intracellular cADPR with concomitant increases in the intracellular Ca2+ concentration ([Ca2+]i). In contrast, activation of an ectocellular ADP-ribosyl cyclase by preincubation of cells with beta-NAD+ led to a dose-dependent increase in cADPR, but no changes in [Ca2+]i were observed. However, extensive washing of the cells following preincubation with NAD+ demonstrated that the increases in cADPR were not intracellular but due to cell surface-associated nucleotide. Accordingly, measurements of ADP-ribosyl cyclase activity in intact T-cells showed ectocellular synthesis of cADPR, but no evidence was obtained for a shift of this activity into the cells which could account for intracellular accumulation of cADPR. Taken together, the results indicate no direct involvement of the ADP-ribosyl cyclase activity of CD38 on the regulation of the cADPR-mediated intracellular Ca2+-signalling in T-lymphocytes.
da Silva
C P
CP
University of Hamburg, Institute of Physiological Chemistry, Department of Enzyme Chemistry, Germany. dasilva@uke.uni-hamburg.de
Schweitzer
K
K
Heyer
P
P
Malavasi
F
F
Mayr
G W
GW
Guse
A H
AH
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000378:metabolism / Q000502:physiology; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000378:metabolism / Q000502:physiology*; D001692:Biological Transport; D015153:Blotting, Western; D020013:Calcium Signaling; D002384:Catalysis; D036563:Cyclic ADP-Ribose; D005434:Flow Cytometry; D006801:Humans; D019169:Jurkat Cells; D008562:Membrane Glycoproteins; D009244:NAD+ Nucleosidase / Q000378:metabolism / Q000502:physiology*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism / Q000502:physiology*
D015703:Antigens, CD; D000943:Antigens, Differentiation; D008562:Membrane Glycoproteins; D011948:Receptors, Antigen, T-Cell; D036563:Cyclic ADP-Ribose; D000246:Adenosine Diphosphate Ribose; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1