Identification of a cDNA/protein leading to an increased Pi-uptake in Xenopus laevis oocytes.
In a previous report we documented an increased Na(+)-dependent transport of inorganic phosphate (P(i)) in Xenopus laevis oocytes injected with mRNA isolated from rabbit duodenum (Yagci et al., Pfluegers Arch. 422:211-216, 1992; ref 24). In the present study we have used expression cloning in oocytes to search for the cDNA/mRNA involved in this effect. The identified cDNA (provisionally named PiUS; for P(i)-uptake stimulator) lead to a 3-4-fold stimulation of Na(+)-dependent P(i)-uptake (10ng cRNA injected, 3-5 days of expression). Na(+)-independent uptake of P(i) was also affected but transport of sulphate and L-arginine (in the presence or absence of sodium) remained unchanged. The apparent K(m)-values for the induced Na(+)-dependent uptake were 0.26 +/- 0.04 mM for P(i) and 14.8 +/- 3.0 mM for Na+. The 1796 bp cDNA codes for a protein of 425 amino acids. Hydropathy analysis suggests a lack of transmembrane segments. In vitro translation resulted in a protein of 60 kDa and provided no evidence of glycosylation. In Northern blots a mRNA of approximately 2 kb was recognized in various tissues including different intestinal segments, kidney cortex, kidney medulla, liver and heart. Homology searches showed no similarity to proteins involved in membrane transport and its control. In conclusion, we have cloned from a rabbit small intestinal cDNA library a novel cDNA encoding a protein stimulating P(i)-uptake into Xenopus laevis oocytes, but which is not a P(i)-transporter itself.
Norbis
F
F
Institute of Physiology, University of Zürich, Switzerland.
Boll
M
M
Stange
G
G
Markovich
D
D
Verrey
F
F
Biber
J
J
Murer
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics*; D005260:Female; D066298:In Vitro Techniques; D017136:Ion Transport; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009865:Oocytes / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism*; D014176:Protein Biosynthesis; D011506:Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018075:RNA, Complementary / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D014018:Tissue Distribution; D014982:Xenopus laevis
D018076:DNA, Complementary; D010710:Phosphates; D011506:Proteins; D018075:RNA, Complementary; D012333:RNA, Messenger