Antigenic and functional properties of the human red blood cell urea transporter hUT-B1.
The Kidd (JK) blood group locus encodes the urea transporter hUT-B1, which is expressed on human red blood cells and other tissues. The common JK*A/JK*B blood group polymorphism is caused by a single nucleotide transition G838A changing Asp-280 to Asn-280 on the polypeptide, and transfection of erythroleukemic K562 cells with hUT-B1 cDNAs carrying either the G838 or the A838 nucleotide substitutions resulted in the isolation of stable clones that expressed the Jk(a) or Jk(b) antigens, respectively, thus providing the first direct demonstration that the hUT-B1 gene encodes the Kidd blood group antigens. In addition, immunochemical analysis of red blood cells demonstrated that hUT-B1 also exhibits ABO determinants attached to the single N-linked sugar chain at Asn-211. Moreover, immunoadsorption studies, using inside-out and right-side-out red cell membrane vesicles as competing antigen, demonstrated that the C- and N-terminal ends of hUT-B1 are oriented intracellularly. Mutagenesis and functional studies by expression in Xenopus oocytes revealed that both cysteines Cys-25 and Cys-30 (but not alone) are essential for plasma membrane addressing. Conversely, the transport function was not affected by the JK*A/JK*B polymorphism, C-terminal deletion (residues 360-389), or mutation of the extracellular N-glycosylation consensus site and remains poorly para-chloromercuribenzene sulfonate (pCMBS)-sensitive. However, transport studies by stopped flow light scattering using Jk-K562 transfectants demonstrated that the hUT-B1-mediated urea transport is pCMBS-sensitive in an erythroid context, as reported previously for the transporter of human red blood cells. Mutagenesis analysis also indicated that Cys-151 and Cys-236, at least alone, are not involved in pCMBS inhibition. Altogether, these antigenic, topologic, and functional properties might have implications into the physiology of hUT-B1 and other members of the urea transporter family.
Lucien
Nicole
N
INSERM U76, Institut National de la Transfusion Sanguine, 6 rue Alexandre Cabanel, 75015 Paris, France.
Sidoux-Walter
Frédéric
F
Roudier
Nathalie
N
Ripoche
Pierre
P
Huet
Martine
M
Trinh-Trang-Tan
Marie-Marcelle
MM
Cartron
Jean-Pierre
JP
Bailly
Pascal
P
D000017:ABO Blood-Group System / Q000276:immunology; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology / Q000502:physiology*; D004912:Erythrocytes / Q000737:chemistry* / Q000378:metabolism; D005260:Female; D006031:Glycosylation; D006801:Humans; D020014:K562 Cells; D007667:Kidd Blood-Group System / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000276:immunology / Q000502:physiology*; D026901:Membrane Transport Proteins; D009865:Oocytes / Q000378:metabolism; D010539:Permeability; D014508:Urea / Q000378:metabolism; D014867:Water / Q000378:metabolism; D014981:Xenopus; D000096982:Urea Transporters
D000017:ABO Blood-Group System; D002352:Carrier Proteins; D007667:Kidd Blood-Group System; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D014508:Urea; D014867:Water; D000096982:Urea Transporters