Arg89Cys Substitution Results in Very Low Membrane Expression of the Duffy Antigen/Receptor for Chemokines in Fyx Individuals

Abstract

The Duffy (FY) blood group antigens are carried by the DARC glycoprotein, a widely expressed chemokine receptor. The molecular basis of the Fya/Fyb and Fy(a-b-) polymorphisms has been clarified, but little is known about the Fyxantigen and the FY*X allele associated with weak expression of Fyb, Fy3, Fy5, and Fy6 antigens. We analyzed here the structure and expression of the FY gene in 4 Fy(a-bweak) individuals. As compared with Fy(a-b+) controls, the Fy(a-bweak) red blood cell membranes contained residual amount of DARC polypeptide and these cells were poorly bound by anti-Fy antibodies and chemokines. The FY gene from Fy(a-b+) and Fy(a-bweak) individuals differed by one substitution, C286T. The resulting Arg89Cys amino acid change reduced the binding of anti-Fy antibodies and chemokines to DARC transfectants. We concluded that the Fybweak donors carried theFY*X allele at the FY locus and that the Fyxantigen corresponds to highly reduced expression of a grossly normal Fyb polypeptide caused by the Arg89Cys substitution. Because FY is a single copy gene, this defect should also affect DARC expression in nonerythroid cells. Because the Fyx phenotype is not associated with apparent clinical consequences, we discussed these findings in the light of the putative roles of DARC in various tissues. Finally, we developed a Fyx DNA typing assay that should be useful for genetic studies and clinical transfusion medicine. © 1998 by The American Society of Hematology.

Structural characterization of the epitope recognized by the new anti‐Fy6 monoclonal antibody NaM185‐2C3

“…Flow cytometry analysis indicated that K562 cells expressing the Fy a or Fy b allelic form of Duffy antigen bound 2C3 MoAb, whereas no binding was detected on mock transfected cells (not shown). Calculation of the MoAb 2C3 apparent binding sites suggested that about 200 000 copies of Duffy antigen were expressed at the surface of K562/Duffy cell, which correlates well with results previously obtained by using the i3A anti‐Fy6 MoAb (Tournamille et al ., 1998). Finally, we found that the binding of 125 I‐IL8 to red cells can be inhibited by 2C3 MoAb in a dose‐dependent fashion, but not by a control MoAb specific for RhD (Fig.…”

Characterization of Jk(a+weak): a new blood group phenotype associated with an altered JK*01 allele

“…Perturbation of expression and function in multipass membrane proteins induced by intracellular amino acid changes has been observed previously. A similar example to that described here is the amino acid exchange of Arg89Cys in the cytoplasmic domain of DARC, the transmembrane protein that carries the FY blood group antigens 24,25 . The presence of Cys89 can weaken the expression of the Fy b antigen (and of the whole DARC protein) to apparently undetectable levels, 26,27 the so‐called Fy x phenotype.…”

“…The presence of Cys89 can weaken the expression of the Fy b antigen (and of the whole DARC protein) to apparently undetectable levels, 26,27 the so‐called Fy x phenotype. This was clearly shown by in vitro expression studies, 24,27 where transcript levels were normal but protein expression at the cell surface was markedly reduced. There are also multiple examples in RhD of single amino acid changes in the intracellular or transmembrane regions which reduce the expression of the protein 28 …”

High‐throughput multiplex PCR genotyping for 35 red blood cell antigens in blood donors

“…As a result of our assessment of antibody prevalence, clinical significance of the antibody and antigen frequency, the following 35 alleles were included into the multiplex screening panel: in the MNS blood group system, the genotypes for MNS1 (‘M’), MNS2 (‘N’), MNS3 (‘S’) and MNS4 (‘s’) [19, 20]; in the Lutheran system LU1 (‘Lu a ’), LU2 (‘Lu b ’), LU8 and LU14 [21]; in the Kell system KEL1 (‘K’), KEL2 (‘k’), KEL3 (‘Kp a ’), KEL4 (‘Kp b ’), KEL6 (‘Js a ’), KEL7 (‘Js b ’), KEL11 , KEL17 and KEL21 [22, 23]; in the Duffy system FY1 (‘Fy a ’), FY2 (‘Fy b ’) [24], one FyX allele [25, 26] and a Fy0 allele (−33 promoter silencing polymorphism) [27, 28]; in the Kidd system JK1 (‘Jk a ’) and JK2 (‘Jk b ’) [29]; in the Diego system DI1 (‘Di a ’), DI2 (‘Di b ’), DI3 (‘Wr a ’) and DI4 (‘Wr b ’) [30, 31]; in the YT (‘Cartwright’) system YT1 (‘Yt a ’) and YT2 (‘Yt b ’) [32]; in the Dombrock system DO1 (‘Do a ’) and DO2 (‘Do b ’) [33]; in the Colton system CO1 (‘Co a ’) and CO2 (‘Co b ’) [34] and the IN1 (‘In a ’) and IN2 (‘In b ’) polymorphism of the Indian blood group system [35].…”