Gerbich Blood Group System Research Articles

Simultaneous high throughput genotyping of 36 blood group systems using NGS based on probe capture technology

Human blood group antigen has important biological functions, and transfusion of incompatible blood can cause alloimmunization and may lead to serious hemolytic reactions. Currently, serological methods are most commonly used in blood group typing. However, this technique has certain limitations and cannot fully meet the increasing demand for the identification of blood group antigens. This study describes a next-generation sequencing (NGS) technology platform based on exon and flanking region capture probes to detect full coding exon and flanking intron regions of the 36 blood group systems, providing a new high-throughput method for the identification of blood group antigens. The 871 capture probes were designed for the exon and flanking intron sequences of 36 blood group system genes, and synchronization analysis for 36 blood groups was developed. The library for NGS was tested using the MiSeq Sequencing Reagent Kit (v2, 300 cycles) by Illumina NovaSeq, and the data were analyzed by the CLC Genomics Workbench 21.0 software. A total of 199 blood specimens have been sequenced for the 41 genes from 36 blood groups. Among them, heterozygote genotypes were found in the ABO, Rh, MNS, Lewis, Duffy, Kidd, Diego, Gerbich, Dombrock, Globoside, JR, LAN, and Landsteiner-Wiene blood group systems. Only the homozygous genotype was found in the remaining 22 blood group systems. The obtained data in the NGS method shows a good correlation (99.98 %) with those of the polymerase chain reaction-sequence-based typing. An NGS technology platform for 36 blood group systems genotyping was successfully established, which has the characteristics of high accuracy, high throughput, and wide coverage.

Genetic diversity of Gerbich alleles in Brazilians reveals an unexpected prevalence of the GE:-2,-3,4 phenotype.

Gerbich (GE) blood group system carries high-frequency antigens and the absence of them leads to rare phenotypes: GE:-2,3,4, GE:-2,-3,4 and GE:-2,-3,-4. Their serological differentiation is limited and misclassification of Gerbich phenotypes may occur, but this can be avoided by molecular characterization. This study aimed to characterize the molecular background responsible for rare Gerbich phenotypes in Brazilian population. We selected eight samples from patients with anti-Ge, six from their relatives and nine samples with normal expression of Gerbich antigens. Serological tests were performed in gel and red blood cells (RBCs) were tested with anti-Ge2 and anti-Ge3. Monocyte monolayer assay (MMA) was performed. Molecular investigation was performed with allele-specific polymerase chain reaction and DNA sequencing. Patient plasma samples reacted with all commercial RBCs. Patient RBCs showed negative results with anti-Ge2 and anti-Ge3. Using MMA two of eight antibodies were clinically significant. Exon 3 was not amplified in any of the patient samples and in two samples from relatives, suggesting the presence of GE*01.-03/GE*01.-03. By sequencing, we identified the genetic variability that interferes with the definition of deletion breakpoints, thus two options of genetic structure were suggested to be responsible for the GE:-2,-3,4 phenotype. This study showed for the first time the genetic diversity of GYPC alleles for carriers of Gerbich-negative phenotypes in a Brazilian population and showed an unexpected prevalence of the GE:-2,-3,4 phenotype. It also demonstrated the importance of using molecular tools to correctly classify Gerbich phenotypes for selection of variants in antigen-matched transfusions.

Occurrence of Rare Deletional Yus and Gerbich Alleles in Syria and Neighbouring Countries

Background: Gerbich-negative phenotypes of the Gerbich Blood Group System (ISBT 020) are very rare (with the exception of Papua New Guinea). The Gerbich-negative phenotypes Yus and Gerbich are negative for the antigens Ge2, and Ge2 and Ge3, respectively. In antigen-negative individuals, anti-Ge2 and anti-Ge3 antibodies can be naturally occurring, or are triggered during pregnancies and after transfusions. Previous studies suggested an elevated frequency of Gerbich-negative phenotypes for the Middle East. In the summer of 2015, a large-scale migration of people from the Middle East to Europe occurred raising the issue of question how to guarantee blood supply for patients and manage antenatal care for pregnant women from these countries. Materials and Methods: To investigate the frequency of rare Gerbich-negative phenotypes, 1,665 immigrants to Germany originating from the Middle East were genetically tested for the presence of rare Yus, i.e., GE*01.-02, and Gerbich, i.e., GE*01-03, alleles and compared to results obtained from 507 Germans. Results: Seven Yus GE*01.-02.01 and one Gerbich GE*01.-03.02 alleles were exclusively observed among people from the Middle East, with five of them clustering among 797 Syrians. No such alleles were observed in Germans. A cumulative Yus- and GE*01.-03-type allele frequency of 0.00314 and resultant overall Gerbich-negative phenotype frequency of one among 101,633 Syrians were calculated. Conclusion: This manuscript describes for the first time an exclusively genetic screening for carriers of Gerbich-negative alleles. In conclusion, the Gerbich blood group system should be considered as one causative agent of unusual antibodies to red cell antigens, in routine patients and pregnant women, especially when originating from the Middle East.

The Gerbich blood group system: old knowledge, new importance.

Antigens of the Gerbich blood group system are expressed on glycophorin C (GPC) and glycophorin D (GPD), minor sialoglycoproteins of human erythrocytes. GPC and GPD help maintain erythrocyte shape of and contributes to the stability of its membrane. There are six high-prevalence Gerbich antigens: Ge2, Ge3, Ge4, GEPL (GE10), GEAT (GE11), GETI (GE12) and five low-prevalence Gerbich antigens: Wb (GE5), Lsa (GE6), Ana (GE7), Dha (GE8), GEIS (GE9). Some Gerbich antigens (Ge4, Wb, Dha, GEAT) are expressed only on GPC, two (Ge2, Ana) are expressed only on GPD, while others (Ge3, Lsa, GEIS, GEPL, GETI) are expressed on both GPC and GPD. Antibodies recognizing GPC/GPD may arise naturally (so-called "naturally-occurring RBC antibodies") or as the result of alloimmunization, and some of them may be clinically relevant. Gerbich antibodies usually do not cause serious hemolytic transfusion reactions (HTR); autoantibodies of anti-Ge2- or anti-Ge3 specificity can cause autoimmune hemolytic anemia (AIHA).

Clinical significance of antibodies to antigens in the Scianna, Dombrock, Colton, Landsteiner-Weiner, Chido/Rodgers, H, Kx, Cromer, Gerbich, Knops, Indian, and Ok blood group systems

This article reviews information regarding the clinical significance of antibodies to antigens in the Scianna, Dombrock, Colton, Landsteiner-Wiener, Chido/Rodgers, H, Kx, Cromer, Gerbich, Knops, Indian, and Ok blood group systems. Like most blood group systems, antibodies to many of the antigens in these groups are rarely encountered because of the high prevalence of the associated antigens in most populations. For many, the clinical significance-that is, the potential to cause reduced survival of transfused antigen-positive red blood cells or a transfusion reaction (e.g., anti-Ge2, anti-H) and/or hemolytic disease of the fetus and newborn (e.g., anti-Coa, anti-Ge3)-has been documented. Some of these antibodies are not always clinically significant, and because of the high prevalence of the antigen, antigen-negative blood may be extremely difficult to find (e.g., anti-LW, anti-Inb). The use of a monocyte monolayer assay may be helpful when making transfusion decisions for patients with these antibodies. For others, their prevalence is so rare that information on the clinical significance of their antibodies is not available (e.g., anti-Co4, anti-Ok).

The use of next-generation sequencing for the determination of rare blood group genotypes.

Next-generation sequencing (NGS) for the determination of rare blood group genotypes was tested in 72 individuals from different ethnicities. Traditional serological-based antigen detection methods, as well as genotyping based on specific single nucleotide polymorphisms (SNPs) or single nucleotide variants (SNVs), are limited to detecting only a limited number of known antigens or alleles. NGS methods do not have this limitation. NGS using Ion torrent Personal Genome Machine (PGM) was performed with a customised Ampliseq panel targeting 15 different blood group systems on 72 blood donors of various ethnicities (Caucasian, Hispanic, Asian, Middle Eastern and Black). Blood group genotypes for 70 of 72 samples could be obtained for 15 blood group systems in one step using the NGS assay and, for common SNPs, are consistent with previously determined genotypes using commercial SNP assays. However, particularly for the Kidd, Duffy and Lutheran blood group systems, several SNVs were detected by the NGS assay that revealed additional coding information compared to other methods. Furthermore, the NGS assay allowed for the detection of genotypes related to VEL, Knops, Gerbich, Globoside, P1PK and Landsteiner-Wiener blood group systems. The NGS assay enables a comprehensive genotype analysis of many blood group systems and is capable of detecting common and rare alleles, including alleles not currently detected by commercial assays.

Genetic background of the rare Yus and Gerbich blood group phenotypes: homologous regions of the GYPC gene contribute to deletion alleles.

The GYPC gene encodes the glycophorins C and D. The two moieties express 12 known antigens of the Gerbich blood group system and functionally stabilize red blood cell membranes through their intracellular interaction with protein 4.1 and p55. Three GYPC exon deletions are responsible for the lack of the high-frequency antigens Ge2 (Yus type, exon 2 deletion), Ge2 and Ge3 (Gerbich type, exon 3 deletion), and Ge2 to 4 (Leach type, exons 3 and 4 deletion), but lack exact molecular description.A total of 29 rare blood samples with Yus (GE:-2,3,4) and Gerbich (GE:-2,-3,4) phenotypes, including individuals of Middle-Eastern, North-African or Balkan ancestry were examined genetically. All phenotypes could be explained by 4 different Yus alleles, characterized by deletions of exon 2 and adjacent introns, and 3 different Gerbich alleles, with deletions of exon 3 and adjacent introns. A 3600 base pair GYPC region, encompassing exon 2 and flanking region, shares a high degree of sequence homology with a region flanking exon 3, probably representing an evolutionary duplication event. Defining the expression of Gerbich variants presently relies on rare serological reagents. Our approach substitutes the serological characterization with a precise genotype approach to identify the rare Yus and Gerbich alleles.

Antibody‐mediated glycophorin C coligation on K562 cells induces phosphatidylserine exposure and cell death in an atypical apoptotic process

Glycophorin C (GPC) is necessary in the maintenance of red blood cell structure. Severe autoimmune hemolytic anemia and hemolytic disease of the fetus and newborn (HDFN) have been associated with Gerbich (Ge) blood group system antigens expressed on GPC. Previous in vitro studies with cord blood progenitor cells have shown that anti-Ge suppresses erythropoiesis. Here, we evaluated the K562 erythroleukemic cell line to study the cellular effects of a murine anti-GPC. Cell proliferation was evaluated after treatment with anti-GPC. Flow cytometry was used to evaluate exofacial phosphatidylserine (PS) expression and cell viability (propidium iodide binding). Cell morphology was evaluated under light microscopy with cytospin preparations stained with May-Grünwald Giemsa. Anti-GPC dramatically inhibited K562 proliferation and increased PS expression, consistent with cytoplasmic blebbing, suggesting evidence of apoptosis. Z-VAD-FMK, an inhibitor of classical apoptosis, was unable to reverse the suppressive effect of anti-GPC. However, hemin was able to attenuate growth suppression. Together, the data suggest that anti-GPC suppresses erythroid proliferation through the induction of nonclassical apoptosis.

Anti‐Ge3 causes late‐onset hemolytic disease of the newborn: the fourth case in three Hispanic families

The Gerbich (Ge) blood group system consists of 11 antigens carried on red blood cell (RBC) membrane glycophorins C and D; of these, Ge:3 antigen is of high prevalence, and the anti-Ge3 is found to be clinically significant. A 34-week neonate born to a Hispanic mother with anti-Ge3 developed late-onset hemolysis with hyperbilirubinemia and was successfully treated with transfusions from her mother. Relevant clinical findings and laboratory results for this case are summarized and compared to three other previously reported cases; all babies were born from a mother of Hispanic ethnicity. Hemolytic disease of the fetus and new born associated with anti-Ge3 is rare but should be considered when working up a broadly reactive RBC antibody screen in women of Hispanic ethnicity. Early identification of pregnant women with anti-Ge3 is recommended for prenatal transfusion planning and close monitoring of the newborn infant for evidence of late-onset anemia.

The Gerbich blood group system: a review

Abstract Antigens in the Gerbich blood group system are expressed on glycophorin C (GPC) and glycophorin D (GPD), which are both encoded by a single gene, GYPC. The GYPC gene is located on the long arm of chromosome 2, and Gerbich antigens are inherited as autosomal dominant traits. There are 11 antigens in the Gerbich blood group system, six of high prevalence (Ge2, Ge3, Ge4, GEPL [Ge10*], GEAT [Ge11*], GETI [Ge12*]) and five of low prevalence (Wb [Ge5], Lsa [Ge6], Ana [Ge7], Dha [Ge8], GEIS [Ge9]). GPC and GPD interact with protein 4.1R, contributing stability to the RBC membrane. Reduced levels of GPC and GPD are associated with hereditary elliptocytosis, and Gerbich antigens act as receptors for the malarial parasite Plasmodium falciparum. Anti-Ge2 and anti-Ge3 have caused hemolytic transfusion reactions, and anti-Ge3 has produced hemolytic disease of the fetus and newborn (HDFN). Immunohematology 2010;26:60–65.

Antibodies to low-incidence antigens

Low-incidence antigens can be defined as those which are known to occur in less than 1% of individuals in most populations. The red cells of almost all blood recipients or prenatal patients therefore lack these rare antigens meaning a chance exposure to blood which does carry a lowincidence antigen almost always results in potential alloimmunization. However, since the antigens are so rare, the chance of exposure is very low. Incompatibility because of an antibody to a low-incidence antigen therefore does not happen very often, but when it does, can cause the same clinical problems, transfusion reactions, hemolytic disease of the fetus and newborn (HDFN), as antibodies to more common specificities. Table 1 lists the low-incidence antigens currently recognized by the International Society of Blood Transfusion Committee on Terminology for Red Cell Surface Antigens. Most have been shown by biochemical and/or molecular methods to be rare variations in established blood group systems as shown. The antigens listed in the 700 series are recognized as distinct low-incidence antigens but there is no biochemical or genetic data that proves they belong to any of the blood group systems. Molecular studies have shown that most of the antigens are the result of a single nucleotide mutation in the gene which codes for a blood group protein, which in turn leads to a change of only one amino acid in the protein. In some cases the antigenic basis is more difficult to determine, particularly in the Rh, MNS and Gerbich blood group systems, where hybrid genes may be involved. For further information on the molecular basis of antigens the interested reader should consult a comprehensive textbook [1–3] Low-incidence antigens are categorized by their low frequency in most populations. Usually this means they are found to be rare or absent in all populations studied but in some cases the frequency can be much higher in some races or ethnic groups. One of the most striking examples of this is the Di antigen. It is rare on red blood cells from Caucasians and Africans but occurs in approxi-

Recombinant glycophorins C and D as tools for studying Gerbich blood group antigens.

The Gerbich blood group system antigens are carried on glycophorin C (GPC) and glycophorin D (GPD) and variants thereof. These glycoproteins have been expressed in a heterologous system to study the individual antigens and to determine whether Ana is antithetical to Ge2. cDNAs encoding GPC, GPD, GPC.Yus, GPC.Ge, GPC.Lsa, and GPD.Lsa were transfected and stably expressed in a human embryonic kidney cell line (293T). Individual Gerbich antigens were analyzed with MoAbs and human polyclonal antibodies by flow cytometry and immunoblotting. Recombinant GPD and GPD.Ana were expressed transiently and analyzed for expression of Ge2 and Ana antigens. All recombinant variants were detected with sialidase-resistant and -sensitive anti-Ge2, anti-Ge3, and anti-Ge4. Ge4 antigen expression was depressed in GPC.Ls(a) transfectants as well as on Ls(a+) RBCs. GPD.An(a) recombinant protein expressed Ana and Ge2 antigens. Cell lines stably expressing glycosylated Gerbich proteins were developed in a heterologous system by transfecting individual variant forms of GPC and GPD. Unexpectedly, it was found that Ge4 antigen is reduced in both the GPC.Ls(a) recombinant and the Ls(a+) RBCs. It was also shown that Ana and Ge2 antigens were expressed on a single GPD.An(a) protein and, therefore, they cannot be antithetical.

Plasmodium falciparum erythrocyte invasion through glycophorin C and selection for Gerbich negativity in human populations.

Geographic overlap between malaria and the occurrence of mutant hemoglobin and erythrocyte surface proteins has indicated that polymorphisms in human genes have been selected by severe malaria. Deletion of exon 3 in the glycophorin C gene (called GYPCDeltaex3 here) has been found in Melanesians; this alteration changes the serologic phenotype of the Gerbich (Ge) blood group system, resulting in Ge negativity. The GYPCDeltaex3 allele reaches a high frequency (46.5%) in coastal areas of Papua New Guinea where malaria is hyperendemic. The Plasmodium falciparum erythrocyte-binding antigen 140 (EBA140, also known as BAEBL) binds with high affinity to the surface of human erythrocytes. Here we show that the receptor for EBA140 is glycophorin C (GYPC) and that this interaction mediates a principal P. falciparum invasion pathway into human erythrocytes. EBA140 does not bind to GYPC in Ge-negative erythrocytes, nor can P. falciparum invade such cells using this invasion pathway. This provides compelling evidence that Ge negativity has arisen in Melanesian populations through natural selection by severe malaria.

Recombinant forms of glycophorin C as a tool for characterization of epitopes for new murine monoclonal antibodies with anti-glycophorin C specificity.

Glycophorin C (GPC) and glycophorin D (GPD) are minor but important components of human RBC membranes. They carry the high-frequency antigens Ge2, Ge3 and Ge4 of the Gerbich blood group system. The epitopes for five new monoclonal antibodies (MoAbs) with anti-GPC specificity were characterized. Two antibodies (4G11 and 5B11) reacted with glycosylated N-terminal epitopes, and three reacted with internal epitopes of GPC. Pepscan analysis showed that the MoAb RB11 required for binding the EPDP sequence, occurring twice in GPC polypeptide chain. The MoAb 7F11 recognized the sequence 13PLSLEPDP20, and the MoAb RB8 did not react with synthetic peptides. Further characterization of the internal epitopes was performed in fluorescence-activated cell sorter (FACS) with the use of recombinant GPC and its variant forms transiently expressed on COS-7 cells. The results indicated that the MoAb RB11 recognized distinctly its target sequence EPDP only in a normal GPC molecule. The reactivity of the MoAb 7F11 with the PLSLEPDP sequence was confirmed and found to be enhanced by the O-glycan at the Ser15 residue. The MoAb RB8 recognized the glycopeptidic epitope in proximity to the Ser15 residue, requiring the presence of O-glycan. The combination of immunochemical techniques with the use of the recombinant forms of GPC has made it possible to define the role of sugar chains in the recognition of peptidic epitopes in glycosylated antigen and sheds new light on the Gerbich system antigens.

MIMA-9, a valuable antibody for screening for rare donors

Abstract Since monoclonal antibodies (Mabs) are potentially available in an unlimited volume, they can be used to screen numerous donor blood samples to identify antigen-negative donors. We have used a Mab (MIMA-9) with characteristics that allow for the simultaneous screening of RBCs of any ABO group for high-incidence antigennegativity in the Kell and Gerbich blood group systems. MIMA-9, a murine IgG2a antibody, previously shown to facilitate the identification of K+k–, Kp(a+b–), K0, McLeod, or Ge:–3 red blood cells (RBCs), was used in MTS gel cards containing anti-mouse IgG as the second antibody to test 1134 K– donors. Among the 1134 donors tested, we found one Kp(a+b–) and one Ge:–2,–3,4 donor. If random donor samples had been used instead of preselecting for K–, we would have expected to identify two K+k– donors. One reagent (MIMA-9) can be used to simultaneously screen for K+k–, Kp(a+b–), K0, McLeod, and Ge:–3 RBCs and thereby conserve rare antisera. Inclusion of anti-mouse IgG in gel cards allowed for rapid screening. MIMA-9 is also a useful reagent to type RBCs with a positive direct antiglobulin test. This antibody is available to donor screening laboratories at no cost for this specific use. Immunohematology 2002:18:43–45.

New Murine Monoclonal Antibodies Directed against Glycophorins C and D, Have Anti‐Ge2 Specificity

The Gerbich blood group system comprises three high-incidence antigens which have been localized on glycophorins C (GPC) and D (GPD) molecules. Murine monoclonal antibodies (mAbs) directed against Ge4 and Ge3 epitopes have already been obtained. We describe two murine mAbs (ZI51.17 and J6) with Ge2 specificity. The mAbs were investigated by serological methods and immunoblotting, with common, Ge-negative and enzyme-modified RBCs. Both mAbs behaved like anti-Ge2 in serological tests, but bound to GPC and GPD on immunoblots. ZI51.17 did not recognize the GPCYus or GPCGe variants, whereas J6 revealed a 25-kD component from Ge:-2,-3,4 membranes, which is identical to the GPCGe molecule. These two mAbs are directed to slightly different epitopes. The ZI51.17 and J6 epitopes are common to GPC and GPD, thus distinct from the Ge2 antigen defined by human alloantibodies.

New Murine Monoclonal Antibodies Directed against Glycophorins C and D, Have Anti-Ge2 Specificity

Background and Objectives: The Gerbich blood group system comprises three high-incidence antigens which have been localized on glycophorins C (GPC) and D (GPD) molecules. Murine monoclonal antibodies (mAbs) directed against Ge4 and Ge3 epitopes have already been obtained. We describe two murine mAbs (ZI51.17 and J6) with Ge2 specificity. Materials and Methods: The mAbs were investigated by serological methods and immunoblotting, with common, Ge-negative and enzyme-modified RBCs. Results: Both mAbs behaved like anti-Ge2 in serological tests, but bound to GPC and GPD on immunoblots. ZI51.17 did not recognize the GPC^Yus or GPC^Ge variants, whereas J6 revealed a 25-kD component from Ge:-2,-3,4 membranes, which is identical to the GPC^Ge molecule. Conclusion: These two mAbs are directed to slightly different epitopes. The ZI51.17 and J6 epitopes are common to GPC and GPD, thus distinct from the Ge2 antigen defined by human alloantibodies.

A Thr552—>Ile substitution in erythroid band 3 gives rise to the Warrior blood group antigen

Recent family studies established that the low-incidence red cell antigen WARR is not part of the MNS, Lutheran, Lewis, Duffy, Kidd, Xg, Chido/ Rodgers, Kx, or Gerbich blood group systems. Continued serologic and genetic studies of WARR suggest that it is carried on erythroid band 3. To test the hypothesis that expression of WARR is controlled by the anion exchanger 1 gene (AE1), AE1 intronic primers that flank the exons encoding the membrane domain of band 3 were prepared. Polymerase chain reaction-amplified products corresponding to exons 11-20 of AE1 were analyzed for single-strand conformational polymorphism (SSCP) in DNA from WARR-positive and WARR-negative individuals. An SSCP was detected in exon 14. Subsequent sequencing revealed a C-->T mutation in codon 552 that leads to a Thr-->Ile substitution. Because the C-->T mutation eliminates a Bbs I restriction site, it was possible to confirm the phenotypes of all family members. To study the possible effect of the Thr552-->Ile substitution on the expression and function of band 3, polymerase chain reaction-amplified reverse-transcribed reticulocyte mRNA was digested with Bbs I. Both alleles of band 3 mRNA were detected in similar quantities, which suggests that the substitution underlying WARR did not interfere with mRNA stability. Comparison of sodium dodecyl sulfate-polyacrylamide gel electrophoretic mobility and size patterns revealed no difference between proteins isolated from WARR-positive and WARR-negative red cells. Further, the presence of WARR did not alter the di-isothiocyano-dihydrostilbene disulfonate (DIDS)-inhibitable influx of radiolabeled sulfate. Although it appears inconsequential to the function of band 3, the red cell polymorphism known as WARR is controlled by AE1. WARR should be therefore included in the Diego blood group system.

A “new” low‐incidence red cell antigen, LOCR, associated with altered expression of Rh antigens

Mild cases of hemolytic disease of the newborn were being studied when it was recognized that the propositi's red cells carried a novel antigen. Serologic and genetic studies of a new low-incidence antigen, LOCR (International Society of Blood Transfusion series number 700.53), were performed. The antigen is associated with altered expression of the Rh antigen c in two unrelated families and in a third proposita and with an altered expression of e in a fourth proposita. The lods for the gene controlling LOCR relative to Rh are 2.107 at theta = 0.00; that is, they fall short of the requirement for inclusion of the antigen in the Rh blood group system. LOCR is excluded from the ABO, MNS, Lutheran, Kell, Duffy, Kidd, Xg, Chido/Rodgers, Kx, and Gerbich blood group systems. Genetic evidence suggests it is not part of the Yt, Colton, or LW systems. Serologic evidence that LOCR is associated with altered expression of c or e strongly suggests that LOCR is part of the Rh blood group system. However, the genetic evidence falls short of the level required for system assignment.

Molecular basis of glycophorin C variants and their associated blood group antigens.

The normal and variant forms of GPC and GPD molecules carry antigens of the Gerbich blood group system. This blood group system comprises three high-incidence antigens (Ge2, Ge3 and Ge4) and four low-incidence antigens (Wb, Lsa, Dha and Ana). Erythrocytes of the Ge and Yus phenotypes lack normal GPC and GPD molecules but express variant molecules (denoted GPC.Ge, GPC.Yus, respectively) that functionally substitute for normal GPC and GPD in the membrane. Leach phenotype cells lack GPC and GPD molecules and are elliptocytic in shape with a membrane that is less deformable than that of normal cells. The Lsa antigen is expressed on higher molecular-weight variants of GPC (GPC.Lsa) and GPD (GPD.Lsa). Wb, Dha and Ana antigens arise from point mutations in the GYPC gene and are expressed on GPC.Wb, GPC.Dha and GPD.Ana, respectively. The structure of each of the variant GPC and GPD molecules and the location of the Gerbich blood group system antigens is discussed. The GYPC gene, located on chromosome 2q14-q21, is 13.5 kb long and comprises four exons. Exons 1, 2 and most of exon 3 encode the N-terminal extracellular domain while the remainder of exon 3 and exon 4 encode transmembrane and cytoplasmic domains of GPC. Exons 2 and 3 are highly homologous, with less than 5% nucleotide divergence. The molecular basis of generation of variation GPC and GPD molecules, and the structure of the GYPC gene from different Leach phenotype individuals, is discussed.