Alternative titles; symbols
HGNC Approved Gene Symbol: RHD
Cytogenetic location: 1p36.11 Genomic coordinates (GRCh38): 1:25,272,486-25,330,445 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 1p36.11 | [Blood group, RH system] | 111690 | 3 | |
| {Hemolytic disease of fetus and newborn, RH-induced} | 619462 | Isolated cases | 3 |
The RHD gene encodes the Rhesus (Rh) blood group (111690) D antigen.
Le Van Kim et al. (1992) cloned cDNAs representing the RHD gene. They found that the predicted translation product is a 417-amino acid protein of molecular mass 45,500 with a membrane organization of 13 bipolar-spanning domains similar to that of the polypeptide encoded by the CcEe gene. The D and CeEe polypeptides differ by 36 amino acids (8.4% divergence), but the NH2- and COOH-terminal regions of the 2 proteins are well conserved. The sequence homology supports the concept that the genes evolve by duplication of a common ancestral gene. It is evident that the controversy between Wiener (1944), who espoused the existence of a single gene with multiple epitopic sites, and the Fisher-Race school (Race, 1944), which held to the existence of 2 closely linked genes, has now been resolved with the conclusion that each view was partially right and partially wrong. None of the 3 researchers survived to see the definitive resolution of the issue. Arce et al. (1993) likewise cloned the RHD gene.
Smythe et al. (1996) provided definitive proof that the RHD gene encodes the D and G antigens and the RHCE gene encodes the c and E antigens. They did this by retroviral-mediated gene transfer using cDNA transcripts of the RHD and RHCE genes and isolated clones that expressed one or the other of these pairs of antigens. Both c and E antigens were expressed after transduction of the test cells with a single cDNA, indicating that the c antigen does not arise by alternative splicing (exon skipping) of the product of the RHCE gene, as had been suggested. The G antigen is an Rh antigen that is expressed on red blood cells carrying a D and/or C antigen.
By Southern blot analysis, Colin et al. (1991) showed that the Rh locus is composed of 2 homologous structural genes, one encoding the Rh D polypeptide and the other encoding both the Cc and the Ee polypeptides (RHCE; 111700). Alternative splicing of a primary transcript was considered the likely mechanism of the encoding of the Cc and Ee polypeptides by a single gene (Le Van Kim et al., 1992).
Wagner and Flegel (2000) determined that the open reading frames of the RHD and RHCE genes have opposite orientations. The 3-prime ends of the genes face each other and are separated by about 30,000 bp that contain the SMP1 gene (605348). The RHD gene is flanked by 2 DNA segments, dubbed Rhesus boxes by Wagner and Flegel (2000), with a length of approximately 9,000 bp, 98.6% homology, and identical orientation. The molecular structure of the RH locus explains the mechanisms for both the RHD deletion and the generation of RHD/RHCE hybrid alleles.
By in situ hybridization using an Rh protein probe, Cherif-Zahar et al. (1991) mapped the Rh blood group lucus, including the RHD gene, to chromosome 1p36.1-p34.3.
Individuals are classified as Rh-positive and Rh-negative according to the presence or the absence of the major D antigen on the surface of their erythrocytes, but more than 46 other antigens, including those of the CcEe series, have been identified (Issitt, 1989).
Wagner and Flegel (2000) identified the RHD deletion site in the prevalent D-negative haplotypes in Caucasians. D-negative status in a mother can lead to Rh-induced hemolytic disease of the fetus and newborn (HDFNRH; 619462) in the fetus.
Miyoshi et al. (2001) described 2 individuals who were mosaic for the Rh blood group phenotype, one erythrocyte population being D-positive and the other D-negative. In both individuals, biparental disomic patterns of markers spanning chromosome 1 were present in peripheral blood leukocytes, whereas only paternal alleles were detected in hair or hair roots in 1 patient and in one-fourth of hair roots in the second patient. Miyoshi et al. (2001) emphasized that isodisomy for chromosome 1 is not infrequent and may cause an unusual RhD phenotype.
Individuals with the D-- phenotype lack both C-like and E-like reactivity in the presence of D antigen. Kemp et al. (1996) examined 5 unrelated Rh D-- homozygotes and found that, in 4 of them, RHCE sequences have been replaced by Rh D sequences. The 5-prime end of these rearrangements all occurred within a 4.2-kb interval around exon 2. There was, however, heterogeneity at the 3-prime end of the rearranged genes, indicating that they were not identical by descent, but rather that independent recombination events had occurred within a small genomic interval. Kemp et al. (1996) noted that 1 individual studied (HD) expressed the very-low-frequency Evans red cell antigen and should therefore strictly be classified as D.. (see below).
'Weak D' and Partial D Phenotypes
About 0.2% to 1% of whites have red blood cells with a reduced expression of the D antigen, known as weak D, formerly known as D(u). Wagner et al. (1999) sequenced all 10 RHD exons and their splice sites in 161 samples from southwest Germany that were identified as weak D. A total of 16 different molecular weak D types plus 2 alleles characteristic of partial D were identified. The amino acid substitutions of weak D types were located in intracellular and transmembrane protein segments and clustered in 4 regions of the protein (amino acid positions 2 to 13, around 149, amino acids 179 to 225, and amino acids 267 to 397). Wagner et al. (1999) concluded that most, if not all, weak D phenotypes carry altered RhD proteins, suggesting a causal relationship. They suggested that genotyping of weak D may guide Rhesus-negative transfusion policy for such molecular weak D types that were prone to develop anti-D.
D-positive individuals harboring a 'partial' D antigen may produce an allo-anti-D similar to that generated in D-negative individuals. Among Europeans, the population frequency of all known partial D phenotypes combined is less than 1%. The molecular basis is generally a gene conversion, in which parts of the RHD gene were substituted by the respective segments of the RHCE gene, and single missense mutations. The situation is more intricate in Africans, however, because the occurrence of aberrant RHD alleles and anti-D immunizations in D-positive individuals is much more frequent than in Europeans (du Toit et al., 1989). Wagner et al. (2002) described 5 RHD alleles, designated DAU-0 to DAU-4, that share a thr379-to-met (T379M) substitution. Four of the alleles expressed a partial D phenotype characterized by the lack of distinct D epitopes or by an anti-D immunization event. Wagner et al. (2002) provided a detailed RHD phylogeny in which the variant alleles formed a previously unknown cluster, distinct from weak D.
In Caucasian RhD-negative individuals, the RHD gene has not been found by any investigators except Hyland et al. (1994). In Japanese, Okuda et al. (1997) found a different situation. Whereas 27.7% of RhD-negative donors demonstrated the presence of the gene, others showed gross or partial deletion of the RHD gene. Additionally, the RHD gene detected in the RhD-negative donors seemed to be intact through sequencing of the RhD polypeptide cDNA and the promoter region of the RHD gene. The phenotypes of these donors with the RHD gene were CC or Cc, but not cc. The discrepant data on the RHD gene in RhD-negative donors between Japanese and Caucasians appeared to be derived from the difference in the frequency of RhD-negative and RhC-positive phenotypes. The possibility that the differences might be related to differences in the Rhesus blood group-associated glycoprotein, the Rh50 comolecule, was to be investigated.
To understand the mechanism underlying the acquisition of a new function by duplicated genes, Innan (2003) studied the evolutionary process within a relatively short time after gene duplication. Innan (2003) theorized that the pattern of allelic variation in duplicated genes is determined mainly by the balance between gene conversion, which operates against diversification of the duplicated gene, and selection, which favors diversification. Innan (2003) applied this theory to the human RHCE and RHD genes. The very high level of amino acid divergence between the 2 genes was observed only in a short region around exon 7. This exon encodes amino acids that characterize the difference between the RHCE and RHD antigens. The observed pattern of DNA variation in this region was considered consistent with the selection model, suggesting that strong selection might be working to maintain the RHCE/RHD antigen variation in the 2-locus system.
The duplication of the rhesus gene occurred during primate evolution (Matassi et al., 1999), giving rise to the RHD and RHCE genes in humans. Thus nonprimate mammals, such as mice, may reveal the ancient state of the RH locus. Wagner and Flegel (2002) analyzed the sequence of the Rh region. Based on the gene positions and orientations, RHCE was determined to represent the ancestral state. The close proximity of SMP1 and RH known in humans was also observed in the mouse RH locus. Wagner and Flegel (2002) concluded that RHD arose by duplication of RHCE. The orientation of RHD was probably inverted during this event. The so-called rhesus boxes, two 9,000-bp DNA segments of identical orientation flanking the RHD gene, may have been instrumental for the duplication.
Goossens et al. (2009) found that Rhd -/- and Rhag (180297) -/- single-knockout mice and Rhd -/- Rhag -/- double-knockout mice were indistinguishable from wildtype mice at a gross phenotypic level, with normal growth, development, and fertility, and no differences in basic plasma and urine chemistry. Both Rhd -/- and Rhag -/- mice showed slightly increased iron levels. Ferritin levels exhibited a tendency toward decrease in Rhag -/- mice of both sexes and in female Rhd -/- mice, whereas a statistically significant trend towards a decrease in transferrin levels was seen only in male Rhag -/- mice. However, double-knockout mice showed no significant changes in iron, transferrin, or ferritin levels. Flow cytometric analysis showed a loss of Rh protein expression and approximately 70% reduction of Rhag glycoprotein expression in red blood cells (RBCs) from Rhd -/- mice. RBCs from Rhag -/- mice also lost Rh protein expression. Rhag +/- mice displayed an approximately 50% decrease in Rhag expression, with a corresponding 50% reduction in Rh protein expression in RBCs. Western blot analysis revealed absence of Rh protein and Icam4 (614088) in RBCs from Rhd -/- or Rhag -/- mice, and expression of these proteins in double-knockout mice was the same as in single knockouts. Ammonium and methylammonium transport was reduced in red cell ghosts from Rhag -/- mice, and Icam4-dependent adhesion of RBCs to endothelial cells was defective in Rhd -/- and Rhag -/- mice. However, Rhd -/- and Rhag -/- mice showed no major alterations in erythrocyte parameters, blood cell count, blood cell morphology, or histology of spleen and bone marrow, and stress erythropoiesis was not modified in double-knockout mice.
Colin et al. (1991) showed that Rh-negative (dd) individuals are homozygous for a deletion of the RHD gene.
Rh-induced hemolytic disease of the fetus and newborn (HDFNRH; 619462) occurs in pregnancies of Rh-negative mothers who carry Rh-positive fetuses. Wagner and Flegel (2000) identified the RHD deletion site in the prevalent D-negative haplotypes in Caucasians. The RHD gene is flanked by 2 highly homologous DNA segments called Rhesus boxes; the 9,142-bp upstream Rhesus box ends approximately 4,900 bp 5-prime of the RHD start codon, and the 9,145 downstream box originates 104 bp after the RHD stop codon. The orientation of the Rhesus boxes is identical. In the prevalent RhD-negative haplotypes, the 903-bp breakpoint region in the Rhesus boxes was located in a 1,463-bp stretch of 99.9% homology resembling a transposon-like human element (THE-1B) and an L2 repetitive DNA element. The single Rhesus box detected in RhD-negative individuals has a hybrid structure, with the 5-prime end representing the upstream Rhesus box and the 3-prime end the downstream Rhesus box. The RhD deletion may be explained by unequal crossing-over triggered by the highly homologous Rhesus boxes flanking the RHD gene. Wagner and Flegel (2000) established technical procedures for specifically detecting the RHD gene deletion in the common RHD-negative haplotypes.
Although the presence or absence of the major antigen, D, at the red blood cell surface determines the Rh-positive or Rh-negative phenotypes, respectively, some rare Rh-positive variants that belong to 1 of the 7 D category phenotypes, D(II) to D(VII) and DFR, can develop anti-D antibodies following immunization by pregnancy or transfusion; their RBCs do not express some of the 9 determinants (epD1 through epD9), which normally compose the so-called D mosaic structure. Rouillac et al. (1995) analyzed the modification of the RHD gene associated with the D(VII) category, characterized by the lack of epD8 and the expression of the low frequency antigen Rh40. They showed that Rh40 and the lack of epD8 are associated with a single point mutation, 329T-C, in exon 2 of the RHD gene. This nucleotide polymorphism resulted in a leucine to proline substitution at amino acid position 110 of the RhD polypeptide.
Wagner et al. (1999) identified 16 different mutations in the RHD gene in patients with the weak D phenotype. The most common by far was a T-to-G transversion at nucleotide 809 resulting in a valine-to-glycine substitution at codon 270 in exon 6. This mutation is located in the transmembrane domain and was identified in 70.29% of weak D alleles in a southwest German population for a haplotype frequency of 1 in 277.
Arce, M. A., Thompson, E. S., Wagner, S., Coyne, K. E., Ferdman, B. A., Lublin, D. M. Molecular cloning of RhD cDNA derived from a gene present in RhD-positive, but not RhD-negative individuals. Blood 82: 651-655, 1993. [PubMed: 8329718]
Cherif-Zahar, B., Mattei, M. G., Le Van Kim, C., Bailly, P., Cartron, J.-P., Colin, Y. Localization of the human Rh blood group gene structure to chromosome region 1p34.3-1p36.1 by in situ hybridization. Hum. Genet. 86: 398-400, 1991. [PubMed: 1900257] [Full Text: https://doi.org/10.1007/BF00201843]
Colin, Y., Cherif-Zahar, B., Le Van Kim, C., Raynal, V., Van Huffel, V., Cartron, J.-P. Genetic basis of the RhD-positive and RhD-negative blood group polymorphism as determined by Southern analysis. Blood 78: 2747-2752, 1991. [PubMed: 1824267] [Full Text: https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)76726-1]
du Toit, E. D., Martell, R. W., Botha, I., Kriel, C. J. Anti-D antibodies in the Rh-positive mothers. (Letter) S. Afr. Med. J. 75: 452 only, 1989. [PubMed: 2497525]
Goossens, D., Trinh-Trang-Tan, M.-M., Debbia, M., Ripoche, P., Vilela-Lamego, C., Louache, F., Vainchenker, W., Colin, Y., Cartron, J.-P. Generation and characterization of Rhd and Rhag null mice. Brit. J. Haemat. 148: 161-172, 2009. [PubMed: 19807729] [Full Text: https://doi.org/10.1111/j.1365-2141.2009.07928.x]
Hyland, C. A., Wolter, L. C., Liew, Y. W., Saul, A. A Southern analysis of Rh blood group genes: association between restriction fragment length polymorphism patterns and Rh serotypes. Blood 83: 566-572, 1994. [PubMed: 7904488] [Full Text: https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)77686-X]
Innan, H. A two-locus gene conversion model with selection and its application to the human RHCE and RHD genes. Proc. Nat. Acad. Sci. 100: 8793-8798, 2003. [PubMed: 12857961] [Full Text: https://doi.org/10.1073/pnas.1031592100]
Issitt, P. D. The Rh blood group system, 1988: eight new antigens in nine years and some observations on the biochemistry and genetics of the system. Transfus. Med. Rev. 3: 1-12, 1989. [PubMed: 2520535] [Full Text: https://doi.org/10.1016/s0887-7963(89)70064-x]
Kemp, T. J., Poulter, M., Carritt, B. A recombination hot spot in the Rh genes revealed by analysis of unrelated donors with the rare D-- phenotype. Am. J. Hum. Genet. 59: 1066-1073, 1996. Note: Erratum: Am. J. Hum. Genet. 60: 749 only, 1997. [PubMed: 8900235]
Le Van Kim, C., Cherif-Zahar, B., Raynal, V., Mouro, I., Lopez, M., Cartron, J. P., Colin, Y. Multiple Rh messenger RNA isoforms are produced by alternative splicing. Blood 80: 1074-1078, 1992. [PubMed: 1379850]
Le Van Kim, C., Mouro, I., Cherif-Zahar, B., Raynal, V., Cherrier, C., Cartron, J.-P., Colin, Y. Molecular cloning and primary structure of the human blood group RhD polypeptide. Proc. Nat. Acad. Sci. 89: 10925-10929, 1992. [PubMed: 1438298] [Full Text: https://doi.org/10.1073/pnas.89.22.10925]
Matassi, G., Cherif-Zahar, B., Pesole, G., Raynal, V., Cartron, J. P. The members of the RH gene family (RH50 and RH30) followed different evolutionary pathways. J. Mol. Evol. 48: 151-159, 1999. [PubMed: 9929383] [Full Text: https://doi.org/10.1007/pl00006453]
Miyoshi, O., Yabe, R., Wakui, K., Fukushima, Y., Koizumi, S., Uchikawa, M., Kajii, T., Numakura, C., Takahashi, S., Hayasaka, K., Niikawa, N. Two cases of mosaic RhD blood-group phenotypes and paternal isodisomy for chromosome 1. Am. J. Med. Genet. 104: 250-256, 2001. [PubMed: 11754053]
Okuda, H., Kawano, M., Iwamoto, S., Tanaka, M., Seno, T., Okubo, Y., Kajii, E. The RHD gene is highly detectable in RhD-negative Japanese donors. J. Clin. Invest. 100: 373-379, 1997. [PubMed: 9218514] [Full Text: https://doi.org/10.1172/JCI119543]
Race, R. R. An 'incomplete' antibody in human serum. (Letter) Nature 153: 771-772, 1944.
Rouillac, C., Le Van Kim, C., Beolet, M., Cartron, J.-P., Colin, Y. Leu110-to-pro substitution in the RhD polypeptide is responsible for the D(VII) category blood group phenotype. Am. J. Hemat. 49: 87-88, 1995. [PubMed: 7741145] [Full Text: https://doi.org/10.1002/ajh.2830490115]
Smythe, J. S., Avent, N. D., Judson, P. A., Parsons, S. F., Martin, P. G., Anstee, D. J. Expression of RHD and RHCE gene products using retroviral transduction of K562 cells establishes the molecular basis of Rh blood group antigens. Blood 87: 2968-2973, 1996. [PubMed: 8639918] [Full Text: https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)64811-X]
Wagner, F. F., Flegel, W. A. RHD gene deletion occurred in the Rhesus box. Blood 95: 3662-3668, 2000. [PubMed: 10845894] [Full Text: https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)63931-3]
Wagner, F. F., Flegel, W. A. RHCE represents the ancestral RH position, while RHD is the duplicated gene. (Letter) Blood 99: 2272-2274, 2002. [PubMed: 11902138] [Full Text: https://doi.org/10.1182/blood-2001-12-0153]
Wagner, F. F., Gassner, C., Muller, T. H., Schonitzer, D., Schunter, F., Flegel, W. A. Molecular basis of weak D phenotypes. Blood 93: 385-393, 1999. [PubMed: 9864185] [Full Text: https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)57619-2]
Wagner, F. F., Ladewig, B., Angert, K. S., Heymann, G. A., Eicher, N. I., Flegel, W. A. The DAU allele cluster of the RHD gene. Blood 100: 306-311, 2002. [PubMed: 12070041] [Full Text: https://doi.org/10.1182/blood-2002-01-0320]
Wiener, A. S. The Rh series of allelic genes. Science 100: 595-597, 1944. [PubMed: 17776134] [Full Text: https://doi.org/10.1126/science.100.2609.595]