NADP+-dependent internalization of recombinant CD38 in CHO cells

CD38 is a 46-kDa type II transmembrane glycoprotein that catalyses the synthesis of cyclic ADP-ribose (cADPR) from NAD(+). cADPR is a second messenger known to regulate intracellular Ca2+-induced Ca2+-release (CICR), A recent study has revealed that CD38 in Namalwa B cells undergoes internalization upon exposure to external NAD(+). In this study, recombinant rat CD38 was expressed in Chinese hamster ovary (CHO) cells and the possibility of the protein to undergo internalization upon exposure to a substrate analog NADP(+) was examined. It was found that such treatment of CHO cells resulted in a decrease of ADP-ribosyl cyclase activity, as well as immunofluorescence of CD38 on the cell surface. The same treatment of CHO cells also resulted in intracellular clustering of CD38 molecules as revealed by confocal. microscopic analysis,The internalized CD38 was purified using a streptavidin/biotin-based method and was found to exhibit both ADP-ribosyl cyclase and cADPR hydrolase activities. On immunoblot, the internalized CD38 appeared as a monomer of 46 kDa under reducing condition of SDS-PAGE, Our data demonstrate that NADP(+) can efficiently induce internalization of CD38, a process that may be important in the production of cADPR intracellularly to regulate CICR. (C) 1999 Academic Press.