Beers K W, Chini E N, Lee H C, Dousa T P
Department of Physiology, Mayo Clinic and Foundation, Rochester 55905.
Am J Physiol. 1995 Mar;268(3 Pt 1):C741-6. doi: 10.1152/ajpcell.1995.268.3.C741.
We have previously shown that NAD+ inhibits renal Na(+)-Pi symport; however, the biochemical mechanism of NAD+ in this action is not clarified. We now propose that NAD+ acts indirectly by first being converted to cyclic ADP-ribose (cADPR), a potent stimulator of intracellular Ca2+ mobilization. In permeabilized opossum kidney (OK) cells, a cell line often employed as a model for study of proximal tubular epithelial transport, cADPR is synthesized from beta-NAD+ in a substrate concentration (0.01-1 mM) and time-dependent manner. That cADPR was generated from beta-NAD+ by OK cells was verified by coelution with authentic cADPR on anion exchange high-performance liquid chromatography and by homologous desensitization of the Ca2+ release bioassay to authentic cADPR. cADPR synthesized by permeabilized OK cells was not influenced by the addition of parathyroid hormone. The OK cell also contains the enzyme activity necessary to catalyze catabolism of cADPR. Identification of these two key enzyme activities of cADPR metabolism in OK cells is consistent with a possible role of cADPR in regulation of the Na(+)-Pi symporter by NAD+ in response to metabolic stimuli.
我们之前已经表明，NAD+抑制肾脏的Na(+)-Pi协同转运；然而，NAD+在此作用中的生化机制尚不清楚。我们现在提出，NAD+首先被转化为环ADP-核糖（cADPR），一种细胞内Ca2+动员的有效刺激物，从而间接发挥作用。在通透的负鼠肾（OK）细胞中，该细胞系常被用作近端肾小管上皮转运研究的模型，cADPR以底物浓度（0.01-1 mM）和时间依赖性方式从β-NAD+合成。通过在阴离子交换高效液相色谱上与 authentic cADPR 共洗脱以及Ca2+释放生物测定对 authentic cADPR 的同源脱敏，验证了OK细胞从β-NAD+生成了cADPR。通透的OK细胞合成的cADPR不受甲状旁腺激素添加的影响。OK细胞还含有催化cADPR分解代谢所需的酶活性。在OK细胞中鉴定出cADPR代谢的这两种关键酶活性，与cADPR在NAD+响应代谢刺激调节Na(+)-Pi协同转运蛋白中可能发挥的作用一致。
