Analysis of NAD-cleavage Enzymes Involved in Signal Transduction System

人细胞表面抗原CD38是一种46 kDa的II型糖蛋白，具有单一跨膜区，其氨基酸序列与Aplysia ADP-核糖环化酶同源。我们先前证明CD38的胞外区具有NAD糖水解酶(NADase)活性，并且全反式维甲酸(RA)诱导的HL-60细胞外型NADase活性是由于CD38(Kontani，K.et al.，J.Biol.Chem.268：16895,1993)。CD38不仅催化NAD^的水解，而且还催化环状ADP-核糖的形成和水解，这是一种新的候选分子，介导了细胞内钙离子的释放。在本研究中，我们获得了以下发现。1.除上述酶活性外，CD38还具有结合透明质酸的能力。2.抗CD38单抗刺激RA分化的HL-60细胞后，细胞内蛋白质迅速酪氨酸磷酸化，其相对分子质量分别为120,000，87,000和77,000。其中一个显著的磷酸化蛋白被鉴定为原癌基因产物c-cbl，p120^&lt；c-cbl&gt；在分化的HL-60细胞中，抗CD38单抗能显著增强甲酰-甲硫氨酸-亮氨酸-苯丙氨酸诱导的超氧化物生成。4.锌离子直接与CD38相互作用，使其从NAD酶转化为ADP-核糖环化酶，这可能是由于阻止了水分子进入酶-底物复合体的中间体。5.虽然CD38基因的表达是通过核受体介导的，但CD38基因第一内含子的负性调控元件可能参与了RA诱导的HL-60细胞CD38基因的表达。

The human cell surface antigen CD38, which has an amino acid sequence homologous to Aplysia ADP-ribosyl cyclase, is a 46-kDa type II glycoprotein with a single-transmembrane domain. We previously demonstrated that the extracellular domain of CD38 exhibits NAD^+ glycohydrolase (NADase) activity and that the ecto-form NADase activity induced by all-trans retinoic acid (RA) in HL-60 cells is due to CD38 (Kontani, K.et al., J.Biol.Chem.268 : 16895,1993). CD38 catalyzes not only the hydrolysis of NAD^+, but also the formation and hydrolysis of cyclic ADP-ribose, which is a novel candidate that mediates Ca^<2+> release from intracellular Ca^<2+> stores. In the present study, we obtained the following findings. 1. Besides these enzyme activities, CD38 had the ability to bind hyaluronate. 2. Stimulation of RA-differentiated HL-60 cells with anti-CD38 monoclonal antibody (mAb) induced rapid tyrosine phosphorylation of cellular proteins with the molecular weight of 120,000,87,000 and 77,000. One of the prominent phosphorylated proteins was identified as the c-cbl proto-oncogene product, p120^<c-cbl>.3. Superoxide formation in response to formyl-Met-Leu-Phe was markedly enhanced by the anti-CD38 mAb in the differentiated HL-60 cells. 4. Zn^<2+> directly interacted with CD38 to convert its catalytic properties from NADase to ADP-ribosyl cyclase, probably due to prevention of the access of water molecule to an intermediate of the enzyme-substrate complex. 5. Although the expression of CD38 mRNA was mediated through nuclear RA receptors, a negative regulatory element present in the first intron of CD38 gene appeared to be involved in the RA-induced expression of CD38 mRNA in HL-60 cells.