The regulatory mechanism of phosphate transport activity mediated by sodium-dependent phosphate transporter

无机磷酸盐（磷酸盐）是糖酵解、糖异生、能量代谢和骨骼矿化过程中必需的营养素。肾刷状边界膜（BBM）上表达的II型钠依赖性磷酸盐转运蛋白（NPT2）在生理和病理生理上调节磷酸盐稳态。NPT2的表达受转录水平、细胞内易位和转录后水平的调控。本研究旨在探讨NPT2细胞内易位的分子机制。PTH在细胞水平上抑制肾磷酸运输，涉及通过激活蛋白激酶A和C介导的肾近端小管BBM选择性内吞去除NPT2。甲状旁腺激素也刺激100或77 kDa蛋白的磷酸化，但不刺激NPT2。在降钙素中也发现了类似的现象。内吞的NPT2被重新分配到细胞质中，但如果PTH治疗延长，它们似乎是快速溶酶体降解的目标，而不是再循环到BBM。在另一种机制中，研究了从大鼠肾脏中分离的独特的npt2相关蛋白（NPT2α， NPT2β和NPT2γ）的参与。NPT2α和NPT2γ被糖基化，分别为45和35 kda蛋白。在分离的BBM囊泡中，n端抗体与45和40-kDa蛋白反应，c端抗体与37-kDa蛋白反应。这些蛋白质的大小与糖基化形式的蛋白质一致。功能分析表明，NPT2γ显著抑制爪蟾卵母细胞中NPT2的活性。研究结果表明，这种短异构体可能是全长转运蛋白的显性阴性抑制剂。因此，PTH和NPT2短异构体刺激NPT2在刷状边界膜的快速内吞内化，导致近端小管磷酸盐重吸收受到抑制。

Inorganic phosphate (phosphate) is an essential nutrient in the processes of glycolysis, gluconeogenesis, energy metabolism and skeletal mineralization. Type II sodium-dependent phosphate transporter (NPT2) expressed on renal brush border membrane (BBM) serves to physiologically and pathophysiologically regulate phosphate homeostasis. The NPT2 expression is regulated by transcriptional level, intracellular translocation of NPT2, and post-transcriptional level. The present study was performed to investigate the molecular mechanism of intracellular translocation of NPT2.PTH inhibits renal phosphate transport at the cellular level involves selective endocytic removal of NPT2 from the BBM of the renal proximal tubule mediated by the activation of both protein kinases A and C activated. PTH also stimulates phosphorylation of 100 or 77 kDa proteins, but not NPT2. Similar observation was found in calcitonin. The endocytosed NPT2 are redistributed to the cytoplasm but they appear to be targeted for rapid lysosomal degradation rather than recycling to the BBM if PTH treatment is prolonged.In another mechanism, the involvement of unique NPT2-related proteins (NPT2α, NPT2β, and NPT2γ) isolated from a rat kidney were investigated. NPT2α and NPT2γ were glycosylated and revealed to be 45- and 35-kDa proteins, respectively. In isolated BBM vesicles, an N-terminal antibody was reacted with the 45- and 40-kDa proteins, and a C-terminal antibody was reacted with the 37-kDa protein. The sizes of these proteins corresponded to those in glycosylated forms. A functional analysis demonstrated that NPT2γ markedly inhibited NPT2 activity in Xenopus oocytes. The findings suggest that this short isoform may function as a dominant negative inhibitor of the full-length transporter.Therefore, it is concluded PTH and short isoform of NPT2 stimulate rapid endocytic internalization of NPT2 at the brush border membrane, leads to an inhibition of phosphate reabsorption in the proximal tubules.