Effect of two tyrosine mutations on the activity and regulation of the renal type II Na/Pi-cotransporter expressed in oocytes.
The rat renal type II Na/Pi-cotransporter (NaPi2), which is regulated by mechanisms involving endocytosis and lysosomal degradation, contains two sequences that show high homology with two tyrosine (Y)-based consensus motifs previously reported to be involved in such intracellular trafficking: GY402FAM matching the consensus sequence GYXXZ, and Y509RWF matching the motif YXXO. Mutations of any of these two Y nearly abolished the NaPi2 mediated 32Pi-uptake after cRNA-injection into oocytes. The mechanisms underlying these defects are however different. Mutation of the Y402 results in a lack of glycosylation and reduced surface expression of the cotransporter, that are specific for the Y402 mutation since substitution of the neighboring F403 did not have any effect. The inhibitory effect of the Y509 mutation is related to a functional inactivation of the protein expressed in the plasma membrane; mutation of the neighboring R510 also led to a decrease in the cotransporter activity. Pharmacological activation of the protein kinase C cascade by DOG induced the retrieval of both wild-type (WT) as well as Y509 cotransporters from the oocyte plasma membrane. These data suggest that the Y402 is important for the surface expression whereas Y509 for the function of the type II Na/Pi-cotransporter expressed in oocytes. Y509 seems not to be involved in the membrane retrieval of the cotransporter.
Hernando
N
N
Institute of Physiology, University of Zürich, Winterthurerstrasse 190, CH-8057, Switzerland.
Traebert
M
M
Forster
I
I
Biber
J
J
Murer
H
H
D019943:Amino Acid Substitution; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics* / Q000502:physiology; D016384:Consensus Sequence; D004075:Diglycerides / Q000494:pharmacology; D017136:Ion Transport / Q000187:drug effects; D008845:Microinjections; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D010710:Phosphates / Q000378:metabolism*; D017354:Point Mutation; D011493:Protein Kinase C / Q000378:metabolism; D018075:RNA, Complementary / Q000235:genetics; D051381:Rats; D012964:Sodium / Q000378:metabolism*; D050608:Sodium-Phosphate Cotransporter Proteins; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D027981:Symporters; D014443:Tyrosine / Q000502:physiology*; D014982:Xenopus laevis
D002352:Carrier Proteins; D004075:Diglycerides; D010710:Phosphates; D018075:RNA, Complementary; D050608:Sodium-Phosphate Cotransporter Proteins; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D027981:Symporters; D014443:Tyrosine; D012964:Sodium; D011493:Protein Kinase C; C013965:diolein