Analysis of opossum kidney NaPi-IIc sodium-dependent phosphate transporter to understand Pi handling in human kidney
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The role of Na+-dependent inorganic phosphate (Pi) transporters in the human kidney is not fully clarified. Hereditary hypophosphatemic rickets with hypercalciuria (HHRH) is caused by loss-of-function mutations in the IIc Na+-dependent Pi transporter (NPT2c/Npt2c/NaPi-IIc) gene. Another Na+-dependent type II transporter, (NPT2A/Npt2a/NaPi-IIa), is also important for renal Pi reabsorption in humans. In mice, Npt2c deletion does not lead to hypophosphatemia and rickets because Npt2a compensates for the impaired Pi reabsorption. To clarify the differences between mouse and human, we investigated the relation between NaPi-IIa and NaPi-IIc functions in opossum kidney (OK) cells.
We cloned NaPi-IIc from OK cells and created opossum NaPi-IIc (oNaPi-IIc) antibodies. We used oNaPi-IIc small interference (si)RNA and investigated the role of NaPi-IIc in Pi transport in OK cells.
We cloned opossum kidney NaPi-IIc cDNAs encoding 622 amino acid proteins (variant1) and examined their pH- and sodium-dependency. The antibodies reacted specifically with 75-kDa and 150-kDa protein bands, and the siRNA of NaPi-IIc markedly suppressed endogenous oNaPi-IIc in OK cells. Treatment with siRNA significantly suppressed the expression of NaPi-4 (NaPi-IIa) protein and mRNA. oNaPi-IIc siRNA also suppressed Na+/H+ exchanger regulatory factor 1 expression in OK cells.
These findings suggest that NaPi-IIc is important for the expression of NaPi-IIa (NaPi-4) protein in OK cells. Suppression of Npt2c may downregulate Npt2a function in HHRH patients.
KeywordsPhosphate Transporter Proximal tubule Hereditary hypophosphatemic rickets with hypercalciuria
We would like to thank the National Disease Research Interchange (NDRI, Philadelphia, PA) for procurement of the kidney samples used in this study. Research reported in this publication was supported by the Ministry of Education, Culture, Sports, Science, and Technology of Japan (No. 23689045 to H. Segawa, No. 26293204 to K. Miyamoto), and The Salt Science Research Foundation (Japan).
Compliance with ethical standards
Conflict of interest
Authors have declared that no conflict of interest exists.
Human and animal rights statement
Mice were handled in accordance with the Guidelines for Animal Experimentation of Tokushima University School of Medicine (T29-3). This study was conducted according to the guidelines laid down in the Declaration of Helsinki, and the protocol was approved by the Ethics Committee of the Tokushima University Hospital (1343-1).
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