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 Previous Article | Table of Contents | Next Article 
Blood, Vol. 92 No. 2 (July 15), 1998:
pp. 664-671
Rhnull Disease: The Amorph Type Results From a Novel
Double Mutation in RhCe Gene on D-Negative Background
By
Cheng-Han Huang,
Ying Chen,
Marion E. Reid, and
Christine Seidl
From the Laboratories of Biochemistry and Molecular Genetics and
Immunochemistry, Lindsley F. Kimball Research Institute, New York Blood
Center, New York, NY; and the Laboratory of Immunogenetics and Tissue
Typing, Frankfurt, Germany.
 |
ABSTRACT |
Rhnull disease, which includes the amorph and regulator
types, is a rare genetic disorder characterized by stomatocytosis and
chronic hemolytic anemia. We studied here a German family transmitting
a putative amorph Rhnull disease gene and identified a rare
mutation causing the loss-of-function phenotype. We analyzed the
genomic and transcript structure of RH30, RH50, and
CD47, the three loci thought to be most critical for expression
of the Rh complex in the red blood cell membrane. We showed that in
this family the Rh50 and CD47 transcripts were normal in primary
sequence. However, the RH30 locus contained an unusual double
mutation in exon 7 of the RhCe gene, in addition to a deletion of the
RhD gene. The mutation targeted two adjacent codons in multiple
arrangements probably via the mechanism of microgene conversion. One
scheme entails a noncontiguous deletion of two nucleotides,
[ATT(Ile322) AT] and [CAC(His323)CC],
whereas the other involves a TC transition [ATT(Ile322)ATC] and a dinucleotide deletion
[CAC(His323)C]. They caused the same shift in open
reading frame predicted to encode a shortened protein with 398 amino
acids. The loss of two transmembrane domains and gain of a new
C-terminal sequence are likely to alter the protein conformation and
impair the Rh complex assembly. Our findings establish the molecular
identity of an amorph Rhnull disease gene, showing that
Rh30 and Rh50 are both essential for the functioning of the Rh
structures as a multisubunit complex in the plasma membrane.
| |
INTRODUCTION |
Rh DEFICIENCY SYNDROME is a rare genetic
disorder of the red blood cell (RBC) transmitted via an autosomal
recessive mode generally through consanguineous pedigrees.1
Its occurrence is characterized by two phenotypic conditions: the
absence of all Rh antigens defines Rhnull disease, whereas
an extremely suppressed expression defines the Rhmod
phenotype. Both Rhnull and Rhmod patients
manifest a mild to moderate hemolytic anemia, and their RBCs show
changes in morphology (stomatocytosis) and abnormalities in plasma
membranes.2-5 Classic genetic studies suggest that Rhnull disease results from distinct mechanisms, depending
on whether the mutations target the Rh antigen locus itself (amorph type) or some unlinked loci that modulate the Rh protein expression (regulator type).1 Biochemical analyses show that
Rhnull cells not only lack the carrier antigens but are
deficient in several membrane glycoproteins, including Rh50, CD47, LW,
and GPB.2-5 This intricate relationship highlights the
organization of Rh and its related proteins as a multisubunit complex
in the RBC membrane and pinpoints the impaired protein-protein
interaction as the hallmark of Rh deficiency syndrome.
Although multiple components are likely involved, the nonglycosylated
Rh30 polypeptides (ie, RhD and RhCE) and Rh50 glycoprotein emerge as
the most important ones in formation of the Rh complex.3-5 Despite the localization of their loci to separate chromosomes, Rh30
and Rh50 belong to the same family and share a notable sequence homology, particularly in their putative  -helices that characterize a 12-transmembrane (TM) topology.6-11 Moreover, evidence is
accumulating that Rh30 and Rh50 may interact directly with each other
to form a structural core in the RBC membrane.12-14 The
other potentially crucial component of the Rh complex may be CD47, an
integrin-associated protein,15,16 whose expression is
profoundly reduced in Rhnull cells,17,18
regardless of the genetic mechanisms involved. The still others, namely
LW and GPB, are likely to be dispensable,19,20 because
their absence neither disrupts the Rh complex nor alters cell
morphology and physiology.
The recent identification of mutations associated with Rh50 gene in
regulator Rhnull patients defines the Rh50 glycoprotein as
a critical coexpressor of Rh30 polypeptides.21,22
Nevertheless, no mutations have been reported to date that target
either the RH30 or CD47 locus in Rhnull
patients.
We report here the identification of the first molecular defect that
occurs as an amorph Rhnull disease gene in a German family described 25 years ago.23 We show that in this family the
genomic structure and transcript expression of RH50 and
CD47 are apparently normal. However, the RH30 locus
contains a novel noncontiguous deletion of two nucleotides in exon 7 of
RhCe gene, in addition to a complete deletion of RhD gene. This novel
double mutation has caused frameshift and premature chain termination,
thereby leading to a loss-of-function phenotype.
| |
MATERIALS AND METHODS |
Blood samples, phenotyping, and immunoblotting.
Control blood samples were from D-positive (genotype, DCe/DCe)
and D-negative (genotype, dce/dce) human blood donors. Blood samples under study were from 4 members of the Rhnull
family, including 2 homozygotes (II-2, B.K., and II-3, D.R.) and 2 heterozygotes (III-1 and III-2; Fig 1).
Members I-1 and I-2 are related23 but not available for
molecular analysis. Retesting of the patients' RBCs confirmed the
absence of all Rh antigens. Immunoblot of RBC membrane proteins was
performed as described.24 Two antibodies, LOR-15C9 for
RhD25 and 2D10 for Rh50,26 were used.
Peroxidase-conjugated antihuman and antimouse Igs were used as the
respective secondary antibodies for band visualization.
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| Fig 1.
Gross structure of RH30 locus in the amorph
Rhnull family as determined by Sph I RFLPs. (Left)
Southern blot analysis of 4 members from the Rhnull family.
Lane 1, D-positive; lane 2, D-negative; and lanes 3 through 6, family
members II-2, II-3, III-1, and III-2, respectively. Genomic DNAs were
digested with Sph I and the polymorphic regions spanning exons
4-7 are shown. Note that RhD gene bands (8.9 and 1.2 kb) are missing in
II-2, II-3, and III-2, whereas the RH50 locus is grossly normal
in all members (gels not shown). Size marker (in kilobases) of  phage DNA cleaved by HindIII is indicated. (Right) Family
pedigree transmitting a putative amorph Rhnull disease
gene. I-1, I-2 (deceased), II-1, and II-4 were not available for
molecular analyses. II-2 (propositus) and II-3 are Rhnull
homozygotes, whereas their children, III-1 and III-2, are
heterozygotes. The genotype of each member was deduced from RFLP
analysis and phenotyping. The amorph copy of RhCe gene on RhD deletion
background is denoted by a crossed box with a vertical line.
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Nucleic acid isolation and Southern blot analysis.
RNA was isolated from hemolysates and genomic DNA from leukocytes, as
described.27,28 The Rh cDNA probes were as
described29; they span the 5 (exon 1-3), middle
(exon 4-7), and 3 (exon 8-10 plus 3-untranslated or UT)
regions, respectively. The Rh50 and CD47 cDNA probes were as detailed
previously.21 Southern blot was performed as
described.28
Reverse transcriptase-polymerase chain reaction (RT-PCR).
The expression of transcripts was analyzed by RT-PCR, as
described.21,29 Rh30-specific primers are shown
(Table 1), and those for Rh50 and CD47 were
detailed in our previous report.21 The mRNA was converted
into cDNA using either a gene-specific 3-UTa21 or
anchored (dT)16 oligomer.30 Total RNA (2 µg)
and 50 ng of primer were incubated at 65°C for 5 minutes and on ice for 5 minutes. Reverse transcription was incubated at 42°C for 75 minutes and inactivated at 72°C for 10 minutes. The cDNA was then
amplified by different pairs of upstream primers spanning the entire
coding region using AmpliTaq DNA polymerase (Perkin-Elmer, Norwalk,
CT). PCR was repeated 35 cycles as follows: 94°C for 1 minute,
55°C for 45 seconds, and 72°C for 1 minute. The last step for
chain extension at 72°C was for 7 to 10 minutes.
Amplification and analysis of RhCe amorph gene.
Sequences of Rh30 genes were amplified by PCR using total genomic DNA
as a template. Exon sequences were analyzed by diagnostic restriction
enzymes to distinguish their D or CE
origin.29 To retrieve unknown intron sequences for primer
design (Table 1), genomic libraries generated by 6 restriction enzymes
were amplified with Rh30 exon-specific primers, as
described.21 Amplification was repeated 30 cycles as
follows: 94°C for 1 minute, 60°C for 30 seconds, and 72°C
for 30 seconds.
Nucleotide determination and sequence analysis.
Amplified cDNA and genomic products were purified by native 5%
polyacrylamide gel electrophoresis (PAGE) and sequenced in both strands
on a Model 373A automated sequencer (Applied Biosystems, Foster City,
CA). The nucleotide and deduced amino acid sequences were analyzed
using the DNASIS program (Hitachi, South San Francisco, CA).
| |
RESULTS |
Gross structure of Rh30 polypeptide genes.
Figure 1 shows a representative Sph1 blot of the
Rhnull family hybridized with the cDNA probe spanning the
polymorphic region of RH30.31 Notably, the
homozygotes and one heterozygote each had a single 7.3-kb band
retaining exons 4-7 whose intensity was comparable to that of
D-negatives. This indicated that the Rhnull patients lacked
RhD gene but had two copies of RhCE gene (Fig 1, left). The other
heterozygote inherited from her father a DCe haplotype; thus,
her D-specific (8.9 and 1.2 kb) and Ce-specific (5.4 and 1.9 kb) bands showed reduced intensity. Analysis of RH50 and CD47 showed no difference between controls and
Rhnull family members (gels not shown), indicating that all
family members had two intact copies of Rh50 and CD47 genes. These
results indicate that the amorph gene occurs on background of
RHD deletion and that both copies of RHCE are defective
in the Rhnull patients (Fig 1, right).
To determine whether the RhD gene deletion is total or partial, we
analyzed 8 of the 10 exons of Rh30 genes by PCR and
diagnostic restriction cleavage (Fig 2).
Whereas heterozygote III-1 had a pattern apparently identical with that
of D-positives, other members lacked D-specific exons 1, 4, 5, 6, 7, 9, and 10. With regard to the cleavage of exons 1 and 2, the two
homozygotes contained Ce- or CE-specific exons, whereas
the heterozygotes exhibited a composite pattern. Taken together, these
data demonstrate a total absence of RhD gene in Rhnull
patients and indicate that some subtle molecular defect(s) have
silenced the phenotypic expression of their RhCE gene.
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| Fig 2.
Assay of Rh30 exons in the Rhnull family by
PCR and restriction analysis. Individual exons were amplified by PCR
and their gene origin was determined by unique restriction site or
primer specificity.29 The digested PCR products were
separated by native 6% PAGE and stained with ethidium bromide. Enzymes
used and exon numbers are indicated above gel panels. Lanes 1 through 6 are as in Fig 1. Below each panel, the size and gene origin of the respective fragments are shown ("+" indicates a cleavage). In the
exon 10 panel, the expected RhD (233 bp) and RhCe (163 bp) bands were
from coamplification of both gene fragments using a common 5
primer, Ex-10s, coupled with two 3-UTa primers (Table 1). Exons
3 and 8 were not analyzed, because the former lacks the unique site and
the latter is identical between RhCE and RhD.6-10
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Identification of the novel mutation in RhCe gene by transcript
analysis.
Although being a consanguineous pedigree,23 the inheritance
pattern of this Rhnull family (Fig 1) did not rule out the
occurrence of possible mutations in related suppressor loci. Therefore,
we analyzed and sequenced the Rh50 and CD47 cDNAs from the
Rhnull patients. No abnormality was found in the Rh50
(Fig 3A) or CD47 transcript (not shown),
conforming to the transmission of an amorph gene.
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| Fig 3.
Identification of the novel mutation in RhCe gene by RT-PCR
and sequencing. (A) Strategy for synthesis and amplification of Rh30
cDNA. Rh30 mRNA was reverse-transcribed into cDNA with either the
gene-specific 3-UTa primer or (dT)16 oligomer and
then amplified with two pairs of upstream primers (Table 1). The cDNA
products for Rh30 (left panel) and Rh50 (right panel) were separated on 1.8% agarose gels. Designations 1 through 6 are as in Fig 1. Lane a,
5-UT/Ex-5a segment; lane b, Ex-4s/3-UTa segment. Bands at bottom are primer dimers. (B) Nucleotide sequencing profiles for the
novel double mutation identified in the amorph form of RhCe cDNAs from
the 2 Rhnull patients (B.K. and D.R.). The mutation affects
2 codons (322 and 323) in exon 7, involving 2 single nucleotide deletions, ATTAT (nt 965 or 966) and CACCC (nt 968) (indicated by
arrow).
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To define the nature of the amorph mutation, the Rh30 cDNA was
amplified with different pairs of gene-specific primers. In both
Rhnull patients, only RhCE but not RhD cDNA was detected, whose size and amount were comparable to that of controls (Fig 3A).
Sequencing showed that the Rh30 cDNA was of Ce type, containing C- and e-specific polymorphisms.6,7,10 However, this cDNA harbored a very rare double mutation in exon 7, which targeted two
adjacent codons (322 and 323) and caused a noncontiguous deletion of
two nucleotides, 966T and 968A (Fig 3B). Significantly, the mutation
resulted in both frameshift and premature termination. Thus, the cDNA
was predicted to encode a shortened RhCe-like polypeptide of 398 amino
acids (Fig 4A). The premature termination
ablated the last 2 TM domains of the protein, whereas the frameshift
gave rise to a new C-terminal sequence of 76 amino acids likely facing the cytoplasmic side (Fig 4B). We did not try to establish the status
of the RhCe-like protein in the membrane, because no antibody can
detect RhCE proteins by immunoblot. However, we showed an absence of
RhD but a significant expression of Rh50 in the Rhnull patients (Fig 4C). In light of a normal structure of Rh50 and CD47, we
conclude that the double deletion is the primary defect underlying the
amorph Rhnull disease gene in this family.
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| Fig 4.
The deduced amino acid sequence and topology of amorph
RhCe from the 2 Rhnull patients. (A) Comparison of the
sequence between the normal (residues 251-417) and putative amorph RhCe
protein (residues 251-398). The frameshift starts from
Pro323 (marked by a star) and the new stretch of 76 amino
acids terminates at Gly398 (bold). (B) Topology and
organization of the putative amorph protein in the membrane. The amorph
protein lacks the last 2 TM domains and does not express any Rh
antigens, although it contains C and e antigen-specific polymorphisms,
Ser103 and Ala226, on the second and fourth
exofacial loops. (C) Immunoblot analysis of Rh30(D) and Rh50 proteins
in the Rhnull membrane. The upper panel was probed with
antibody LOR-15C9 and the lower one with 2D10. Lane designations are as
in Fig 1. Note that no RhD protein was detected in D-negative and
Rhnull patients, as expected. In contrast, a significant
amount of Rh50 protein was present in the Rhnull cells.
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Inheritance of amorph gene and confirmation of genomic
mutation.
The detection of a unique RhCe-like transcript (Fig 3) indicated that
the patients are homozygous, whereas their children are heterozygous,
for the mutation. As a new BamHI site was created, we performed
a diagnostic assay in the 4 members. Whereas no cleavage was seen in
controls, a total digestion in the 2 patients and a partial digestion
in their children were observed
(Fig 5A). This pattern is
entirely consistent with the transmission of the amorph gene in an
autosomal recessive fashion. Direct sequencing of the exon 7 containing
fragment showed the same noncontiguous deletion (Fig 3B), confirming
homozygosity of the mutation in the 2 patients.
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| Fig 5.
Family inheritance of the amorph Rhnull gene.
(A) The genomic region encompassing exon 7 of RhCE gene was amplified
by 2 pairs of primers In-6s/Ex-7a and In-6s/In-7a (Table 1). The
In-6s/Ex-7a fragments were cut by BamHI (G GATCC), which is
diagnostic of the mutation, and separated by native 6% PAGE. Although
no cleavage is seen in control lanes, the respective fragments of 264 bp (denoted by a star) were cut completely in the Rhnull
patient and partially in the heterozygotes. (B) Sequencing of the
genomic fragments In-6s/In-7a. The 2 patients showed the same mutation,
2 deleted nucleotides (966T and 968A, boxed), in this fragment, which
was sequenced on both strands. Intronic nucleotides are denoted by lowercase letters and exonic ones by uppercase letters. Dashes indicate
identical nucleotides. The 2 primers for PCR amplification are
overlined.
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DISCUSSION |
In this study, we examined a German family in which a putative amorph
Rhnull disease gene is transmitted on a consanguineous background.23 Molecular analysis of three genetic loci
relating to the expression of the Rh complex, RH30, RH50, and
CD47, has led to the detection of a rare double mutation in the
Rh30 gene and exclusion of possible candidate mutations in Rh50 and
CD47 genes. The mutation is defined as a noncontiguous deletion of 2 nucleotides targeting 2 adjacent codons in exon 7 of the RhCe gene and
thus becomes the first known amorph defect at RH30. With a
deleted RhD gene positioned in cis and trans, the
mutation has silenced the antigen expression of the remaining RhCe
gene, causing a loss-of-function phenotype in the two
Rhnull patients.
Of the double deletions identified, one hit the second or third
position of codon ATT(Ile322) and the other the second
position of codon CAC(His323). Based on the alignment of
deletion forms, two models may be postulated to account for their
origin. In spontaneous model, two hypothetical schemes could be
proposed, each evoking two molecular events. In scheme I, the deletion
of 966T (or 965T) and 968A is assumed to occur separately
(Fig 6A). In scheme II, one event causes a
transition [ATTATC] and the other a dinucleotide deletion
[CACC] (Fig 6A). In the microgene conversion model,32 the same end product may arise by a single event in which a
heteroduplex of RhD and RhCe could be formed via homologous pairing;
failure in repair synthesis involving codons 322 and 323 would result in patched transfer of the RhD-specific C residue and a contiguous deletion of TC dinucleotide (Fig 6B). Although microgene conversion might be infrequent in the Rh system,33 it is implicated as an important mechanism for the diversity of Ig and MHC gene families and provides a plausible explanation to the origin of some rare spontaneous mutations.34,35
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| Fig 6.
Two models accounting for origin of the double mutation
in the amorph Rhnull disease gene. (A) Spontaneous mutation
model. Sequences of codons 318-327 in RhD and RhCe genes are shown on top. Three mismatches at center right are marked by stars. Possible arrangements of the mutated region (boxed) in the amorph gene are
depicted in two hypothetical schemes (see Discussion for details). a
and b denote alternatives of the same scheme. Scheme I shows a
noncontiguous deletion of two nucleotides, whereas scheme II shows a
contiguous deletion of 2 nucleotides in association with a TC
transition. The BamHI site is shown. (B) Microgene conversion model. A heteroduplex is formed between RhD and RhCe genes via homologous pairing and strand synapsis. A failure in repair synthesis involving codons 322 and 323 would result in AC transition and contiguous deletion of 2 nucleotides (boxed). This model is compatible with scheme II shown above but accommodates the latter in a single event.
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The amorph mutation has targeted a nonconserved region, possibly the
fifth endofacial loop, in the Rh30 polypeptides.6-10 Thus,
its result as a loss-of-function phenotype is most likely due to the
inevitable shift in open reading frame. The status of the putative
RhCe-like protein in the Rhnull cells remains to be
established. Nevertheless, several lines of evidence suggest that the
mutant protein may be inserted into the membrane but not accessible to
the antibodies that are strictly conformation-dependent. (1) The
RhCe-like transcript was abundantly expressed, suggesting that the
steady-state level of mRNA is not affected by the mutation. (2) Albeit
at a reduced level, Rh50 was readily detectable by immunoblot. As a
critical interaction partner of Rh30,12-14 this expression
implies a possible association of Rh50 with the RhCe-like protein. (3)
Prior surface-labeling studies detected a structurally similar Rh30
polypeptide in the Rhnull membrane,36
indicating that the absence of Rh antigens does not necessarily mean
the absence of Rh proteins. As predicted, the primary sequence of the
RhCe-like protein indicates a loss of 2 TM domains and a gain of 76 new
amino acids in the C-terminal region. Such structural changes would
disturb the conformation of the RhCe-like protein and affect its
interaction with Rh50 and other related glycoproteins in the RBC
membrane.
Identification of the first amorph Rh30 gene establishes that Rh30,
akin to its interacting partner Rh50, is an essential member of the Rh
membrane structures. However, the molecular details regarding this
protein-protein interaction remain to be elucidated. Limited
proteolysis and antibody probing suggested that Rh30 and Rh50 are each
composed of two 6-TM domains, forming a tetramer through N-terminal
contacts.13 However, additional contact sites are likely
present to further dictate the assembly of Rh membrane structures. The
amorph protein has a change in the C-terminal portion after
Pro323, but its N-terminal sequence is identical with the
wild-type one. Its association with the dysfunction of the Rh complex
suggests a possible interaction of Rh30 with Rh50 through their
C-terminal regions. Notably, the C-termini of Rh50 mutants in regulator
Rhnull disease resulting from a splicing donor
mutation21 or a point deletion22 are also
abnormal because of the inherent frameshift and premature termination.
In the former, the only difference from the Rh50 protein lies
C-terminal to the 10th TM domain, whereas the latter is altered after
the 11th TM domain. Missense mutations targeting the conserved
C-terminal TM domains of Rh50 are also associated with regulator
Rhnull (our unpublished data). These
coincidental observations suggest that the C-terminal regions
of Rh30 and Rh50 play a crucial role in forming the multisubunit Rh
structures in the RBC membrane.
| |
FOOTNOTES |
Submitted December 17, 1997;
accepted March 20, 1998.
Supported by National Institutes of Health Grant No. HL 54459.
The accession numbers of the nucleotide and amino acid sequences are
AF056965, AF056966, and AF056967.
Address reprint requests to Cheng-Han Huang, MD, PhD, Laboratory of
Biochemistry and Molecular Genetics, Lindsley F. Kimball Research
Institute, New York Blood Center, 310 E 67th St, New York, NY 10021;
e-mail: chuang{at}nybc.org.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. section
1734 solely to indicate this fact.
| |
ACKNOWLEDGMENT |
The authors are grateful to Dr A. Blancher for his monoclonal antibody
LOR-15C9 against RhD and to Dr A.E.G.Kr. von dem Borne for his
monoclonal antibody 2D10 against Rh50. We thank G. Halverson for
immunoblot analysis and T. Huima-Byron, Y. Oskov, and R. Ratner for
photoprints and computer graphics.
| |
REFERENCES |
1. Race RR, Sanger R: Blood Groups in Man (ed 6). London, UK,
Blackwell Scientific, 1975, p 220
2.
Agre P,
Catron J-P:
Molecular biology of the Rh antigens.
Blood
78:551,
1991[Abstract/Free Full Text]
3.
Anstee DJ,
Tanner MJA:
Biochemical aspects of the blood group Rh (rhesus) antigens.
Clin Haematol
6:401,
1993
4.
Cartron J-P,
Agre P:
Rh blood groups and Rh deficiency syndrome.
Blood Cell Biochem
6:189,
1995
5.
Huang C-H:
Molecular insights into the Rh protein family and associated antigens.
Curr Opin Hematol
4:94,
1997[Medline]
[Order article via Infotrieve]
6.
Avent ND,
Ridgwell K,
Tanner MJA,
Anstee DJ:
cDNA cloning of a 30 kDa erythrocyte membrane protein associated with Rh (Rhesus)-blood group-antigen expression.
Biochem J
271:821,
1990[Medline]
[Order article via Infotrieve]
7.
Cherif-Zahar B,
Bloy B,
Le Van Kim C,
Blanchard D,
Bailly P,
Hermand P,
Salmon C,
Cartron J-P,
Colin Y:
Molecular cloning and protein structure of a human blood group Rh polypeptide.
Proc Natl Acad Sci USA
87:6243,
1990[Abstract/Free Full Text]
8.
Le Van Kim C,
Mouro I,
Cherif-Zahar B,
Raynal V,
Cherrie C,
Cartron J-P,
Colin Y:
Molecular cloning and primary structure of the human blood group RhD polypeptide.
Proc Natl Acad Sci USA
59:19925,
1992
9.
Arce MA,
Thompson ES,
Wagner S,
Coyne KE,
Ferdman BA,
Lubin DM:
Molecular cloning of RhD cDNA derived from a gene present in RhD-positive, but not RhD-negative individuals.
Blood
82:651,
1993[Abstract/Free Full Text]
10.
Mouro I,
Collin Y,
Cherif-Zahar B,
Cartron J-P,
Le Van Kim C:
Molecular genetic basis of the human Rhesus blood group system.
Nat Genet
5:62,
1993[Medline]
[Order article via Infotrieve]
11.
Ridgwell K,
Spurr NK,
Laguda B,
MacGeoch C,
Avent ND,
Tanner MJA:
Isolation of cDNA clones for a 50 kDa glycoprotein of the human erythrocyte membrane associated with Rh (Rhesus) blood group antigen expression.
Biochem J
287:223,
1992
12.
Ridgwell K,
Eyers SAC,
Mawby WJ,
Anstee DJ,
Tanner MJA:
Studies on the glycoprotein associated with Rh (Rhesus) blood group antigen expression in the human red cell membrane.
J Biol Chem
269:6410,
1994[Abstract/Free Full Text]
13.
Eyers SAC,
Ridgwell K,
Mawby WJ,
Tanner MJA:
Topology and organization of the human Rh (Rhesus) blood group-related polypeptides.
J Biol Chem
269:6417,
1994[Abstract/Free Full Text]
14.
Avent ND,
Liu W,
Warner KM,
Marby WJ,
Jones JW,
Ridgwell K,
Tanner MJA:
Immunochemical analysis of the human erythrocyte Rh polypeptides.
J Biol Chem
271:14233,
1996[Abstract/Free Full Text]
15.
Campbell IG,
Freemont PS,
Foulkes W,
Trowsdale J:
An ovarian tumor marker with homology to vaccinia virus contains an IgV-like region and multiple transmembrane domains.
Cancer Res
52:5416,
1992[Abstract/Free Full Text]
16.
Lindberg FP,
Gresham HD,
Schwartz E,
Brown EJ:
Molecular cloning of integrin-associated protein: An immunoglobulin family member with multiple membrane spanning domains implicated in v 3-dependent ligand binding.
J Cell Biol
123:485,
1993[Abstract/Free Full Text]
17.
Miller YE,
Daniels GL,
Jones C,
Palmer DK:
Identification of a cell-surface antigen produced by a gene on human chromosome 3 (cen-q22) and not expressed by Rh (null) cells.
Am J Hum Genet
41:1061,
1987[Medline]
[Order article via Infotrieve]
18.
Lindberg FP,
Lublin DM,
Telen MJ,
Veile RA,
Miller YE,
Donis-Keller H,
Brown EJ:
Rh-related antigen CD47 is the signal-transducer integrin-associated protein.
J Biol Chem
269:1567,
1994[Abstract/Free Full Text]
19.
Hermand P,
Le Pennec PY,
Rouger P,
Cartron J-P,
Bailly P:
Characterization of the gene encoding the human LW blood group protein in LW+ and LW- phenotypes.
Blood
87:2962,
1996[Abstract/Free Full Text]
20.
Huang C-H,
Blumenfeld OO:
MNSs blood groups and major glycophorins: Molecular basis for allelic variation.
Blood Cell Biochem
6:153,
1995
21.
Huang C-H:
The human Rh50 glycoprotein gene: Structural organization and associated splicing defect resulting in Rhnull disease.
J Biol Chem
273:2207,
1998[Abstract/Free Full Text]
22.
Cherif-Zahar B,
Raynal V,
Gane P,
Mattei M-G,
Bailly P,
Cartron J-P,
Colin Y:
Candidate gene acting as a suppressor of the RH locus in most cases of Rh deficiency.
Nat Genet
12:168,
1996[Medline]
[Order article via Infotrieve]
23.
Seidl S,
Spielmann W,
Martin H:
Two siblings with Rhnull disease.
Vox Sang
23:182,
1972[Medline]
[Order article via Infotrieve]
24.
Towbin H,
Staelin T,
Gordon J:
Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some implications.
Proc Natl Acad Sci USA
76:4350,
1979[Abstract/Free Full Text]
25.
Apoil PA,
Reid ME,
Halverson G,
Mouro I,
Cilin Y,
Roubinet F,
Cartron J-P,
Blancher A:
A human monoclonal anti-D antibody which detects a nonconformation-dependent epitope on the RhD protein by immunoblot.
Br J Haematol
98:365,
1997[Medline]
[Order article via Infotrieve]
26.
Mallinson G,
Anstee DJ,
Avent ND,
Ridgwell K,
Tanner MJA,
Daniels GL,
Tippett P,
von dem Borne AEGKr:
Murine monoclonal antibody MB-2D10 recognizes Rh-related glycoproteins in human red cell membrane.
Transfusion
30:222,
1990[Medline]
[Order article via Infotrieve]
27.
Goossens M,
Kan YW:
DNA analysis in the diagnosis of hemoglobin disorders.
Methods Enzymol
76:805,
1981[Medline]
[Order article via Infotrieve]
28.
Huang C-H,
Guizzo M-L,
Leigh N,
Blumenfeld OO:
Typing of MNSs blood group specific sequences in the human genome and characterization of a restriction fragment tightly linked to S-s-alleles.
Blood
83:222,
1991
29.
Huang C-H:
Alteration or RH gene structure and expression in dCCee and DCw-red blood cells: Phenotypic heterozygosity versus genotypic heterozygosity.
Blood
88:2326,
1996[Abstract/Free Full Text]
30.
Froham MA,
Dush MK,
Martin GR:
Rapid production of full-length cDNAs from rare transcripts: Amplification using a single gene-specific oligonucleotide primer.
Proc Natl Acad Sci USA
85:8998,
1988[Abstract/Free Full Text]
31.
Huang C-H,
Reid ME,
Chen Y,
Couglan G,
Okubo Y:
Molecular definition of red cell Rh haplotypes by tightly linked Sph I RFLPs.
Am J Hum Genet
58:133,
1996[Medline]
[Order article via Infotrieve]
32.
Szostak JW,
Orr-Weaver TL,
Rothstein RJ:
The double-strand-break model for recombination model.
Cell
33:25,
1983[Medline]
[Order article via Infotrieve]
33.
Huang C-H,
Chen Y,
Reid ME:
Human D IIIa erythrocytes: RhD protein is associated with multiple dispersed amino acid variations.
Am J Hematol
55:139,
1997[Medline]
[Order article via Infotrieve]
34.
Maizels N:
Might gene conversion be the mechanism of somatic hypermutation of mammalian immunoglobulin genes?
Trends Genet
5:4,
1989[Medline]
[Order article via Infotrieve]
35.
Kappes D,
Strominger JL:
Human class II major histocompatibility complex genes and proteins.
Annu Rev Biochem
57:991,
1988[Medline]
[Order article via Infotrieve]
36.
Connor J,
Bar-Eli M,
Gillum KD,
Schroit AJ:
Evidence for a structurally homologous Rh-like polypeptide in Rhnull erythrocytes.
J Biol Chem
267:26050,
1992[Abstract/Free Full Text]
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Abstract
Introduction
Abstract