Cloning and expression of cDNA for a Na/Pi cotransport system of kidney cortex.
A cDNA library from rabbit kidney cortex was screened for expression of Na-dependent transport of phosphate (Pi) using Xenopus laevis oocytes as an expression system. A single clone was eventually isolated (designated NaPi-1) that stimulated expression of Na/Pi cotransport approximately 700-fold compared to total mRNA. The predicted sequence of the Na/Pi cotransporter consists of 465 amino acids (relative molecular mass, 51,797); hydropathy profile predictions suggest six (possibly eight) membrane-spanning segments. In vitro translation of NaPi-1/complementary RNA in the presence of pancreatic microsomes indicated NaPi-1 to be a glycosylated protein; four potential N-glycosylation sites are present in the amino acid sequence. Northern blot analysis demonstrated the presence of NaPi-1/mRNA in kidney cortex and liver; no hybridization signal was obtained with mRNA from other tissues (including small intestine). Kinetic analysis of Na/Pi cotransport expressed by NaPi-1/complementary RNA demonstrated characteristics (sodium interaction) similar to those observed in cortical apical membranes. The alignment of 5 amino acid residues (Gly342/Ala381-Xaa-Xaa-Xaa-Xaa-Leu386-Xaa-Xaa-Xaa-P ro390- Arg391) is consistent with a motif proposed for Na-dependent transport systems. We conclude that we have cloned a cDNA for a Na/Pi cotransport system present in rabbit kidney cortex.
Werner
A
A
Department of Physiology, University of Zürich, Switzerland.
Moore
M L
ML
Mantei
N
N
Biber
J
J
Semenza
G
G
Murer
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D015870:Gene Expression; D007672:Kidney Cortex / Q000502:physiology*; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D010710:Phosphates / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D016415:Sequence Alignment; D012964:Sodium / Q000378:metabolism; D050608:Sodium-Phosphate Cotransporter Proteins; D012995:Solubility; D027981:Symporters; D014982:Xenopus laevis
D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D010710:Phosphates; D012333:RNA, Messenger; D050608:Sodium-Phosphate Cotransporter Proteins; D027981:Symporters; D004247:DNA; D012964:Sodium