Relative contributions of Na+-dependent phosphate co-transporters to phosphate transport in mouse kidney: RNase H-mediated hybrid depletion analysis.
Reabsorption of Pi in the proximal tubule of the kidney is an important determinant of Pi homoeostasis. At least three types (types I-III) of high-affinity Na+-dependent Pi co-transporters have been identified in mammalian kidneys. The relative roles of these three types of Na+/Pi co-transporters in Pi transport in mouse kidney cortex have now been investigated by RNase H-mediated hybrid depletion. Whereas isolated brush-border membrane vesicles showed the presence of two kinetically distinct Na+/Pi co-transport systems (high Km-low Vmax and low Km-high Vmax), Xenopus oocytes, microinjected with polyadenylated [poly(A)+] RNA from mouse kidney cortex, showed only the high-affinity Pi uptake system. Kidney poly(A)+ RNA was incubated in vitro with antisense oligonucleotides corresponding to Npt-1 (type I), NaPi -7 (type II) or Glvr-1 (type III) Na+/Pi co-transporter mRNAs, and then with RNase H. Injection of such treated RNA preparations into Xenopus oocytes revealed that an NaPi-7 antisense oligonucleotide that resulted in complete degradation of NaPi-7 mRNA (as revealed by Northern blot analysis), also induced complete inhibition of Pi uptake. Degradation of Npt-1 or Glvr-1 mRNAs induced by corresponding antisense oligonucleotides had no effect on Pi transport, which was subsequently measured in oocytes. These results indicate that the type II Na+/Pi co-transporter NaPi-7 mediated most Na+-dependent Pi transport in mouse kidney cortex.
Miyamoto
K
K
Department of Clinical Nutrition, School of Medicine, Tokushima University, Kuramoto-Cho 3, Tokushima 770, Japan.
Segawa
H
H
Morita
K
K
Nii
T
T
Tatsumi
S
S
Taketani
Y
Y
Takeda
E
E
D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D007672:Kidney Cortex / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008813:Mice, Inbred ICR; D008845:Microinjections; D008871:Microvilli / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000378:metabolism; D009865:Oocytes; D010710:Phosphates / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism / Q000494:pharmacology; D016914:Ribonuclease H / Q000378:metabolism*; D050608:Sodium-Phosphate Cotransporter Proteins; D050609:Sodium-Phosphate Cotransporter Proteins, Type I; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D050611:Sodium-Phosphate Cotransporter Proteins, Type III; D027981:Symporters; D014982:Xenopus laevis
D002352:Carrier Proteins; D016376:Oligonucleotides, Antisense; D010710:Phosphates; D012333:RNA, Messenger; C493547:Slc17a1 protein, mouse; C493587:Slc20a1 protein, mouse; D050608:Sodium-Phosphate Cotransporter Proteins; D050609:Sodium-Phosphate Cotransporter Proteins, Type I; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D050611:Sodium-Phosphate Cotransporter Proteins, Type III; D027981:Symporters; D016914:Ribonuclease H