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Saison C.,National Institute of Blood Transfusion INTS | Helias V.,National Institute of Blood Transfusion INTS | Ballif B.A.,University of Vermont | Peyrard T.,National Institute of Blood Transfusion INTS | And 10 more authors.
Nature Genetics | Year: 2012

The breast cancer resistance protein, also known as ABCG2, is one of the most highly studied ATP-binding cassette (ABC) transporters because of its ability to confer multidrug resistance. The lack of information on the physiological role of ABCG2 in humans severely limits cancer chemotherapeutic approaches targeting this transporter. We report here that ABCG2 comprises the molecular basis of a new blood group system (Junior, Jr) and that individuals of the Jr(a-) blood type have inherited two null alleles of ABCG2. We identified five frameshift and three nonsense mutations in ABCG2. We also show that the prevalence of the Jr(a-) blood type in the Japanese and European Gypsy populations is related to the p.Gln126* and p.Arg236 * protein alterations, respectively. The identification of ABCG2 -/- (Jr(a-)) individuals who appear phenotypically normal is an essential step toward targeting ABCG2 in cancer and also in understanding the physiological and pharmacological roles of this promiscuous transporter in humans. © 2012 Nature America, Inc. All rights reserved.


Helias V.,National Institute of Blood Transfusion INTS | Saison C.,National Institute of Blood Transfusion INTS | Peyrard T.,National Institute of Blood Transfusion INTS | Peyrard T.,National Reference Center for Blood Groups | And 5 more authors.
Human Mutation | Year: 2013

KLF1 encodes an erythroid transcription factor, whose essential function in erythropoiesis has been demonstrated by extensive studies in mouse models. The first reported mutations in human KLF1 were found in individuals with a rare and asymptomatic blood type called In(Lu). Here, we show that KLF1 haploinsufficiency is responsible for the In(Lu) blood type, after redefining this peculiar blood type using flow cytometry to quantify the levels of BCAM and CD44 on red blood cells. We found 10 (seven novel) heterozygous KLF1 mutations responsible for the In(Lu) blood type. Although most were obligate loss-of-function mutations due to the truncation of the DNA-binding domain of KLF1, three were missense mutations that were located in its DNA-binding domain and impaired the transactivation capacity of KLF1 in vitro. We further showed that the levels of the hemoglobin variants HbF and HbA2 were increased in the In(Lu) blood type, albeit differently. The levels of the membrane glycoproteins BCAM and CD44 were also differently reduced on In(Lu) red blood cells. This biochemical and genetic analysis of the In(Lu) blood type tackles the phenotypic outcome of haploinsufficiency for a transcription factor. © 2012 Wiley Periodicals, Inc.


Peyrard T.,National Institute of Blood Transfusion INTS
Immunohematology / American Red Cross | Year: 2013

LAN (Langereis) was officially recognized by the International Society of Blood Transfusion in 2012 as being the 33rd human blood group system. It consists of one single high-prevalence antigen,Lan (LANl). The ABCB6 protein is the carrier of the Lan blood group antigen. The ABCB6 gene (chromosome 2q36, 19 exons)encodes the ABCB6 polypeptide (ATP-binding cassette protein,subfamily B, member 6), known as a porphyrin transporter.The exceptional Lan- people do not express ABCB6 (Lan null phenotype), owing to several different molecular mechanisms affecting ABCB6: frameshift leading to a premature stop codon(deletion, insertion, or nonsense mutation of nucleotides);missense mutation; or intronic mutation responsible for RNA splicing defect. Despite the Lan antigen's being reported to play a key role in erythropoiesis and detoxification of cells, Lan people do not appear to demonstrate susceptibility to any disease or seemingly physiologic disorder. Anti-Lan has been described as having variable clinical significance, either for hemolytic transfusion reactions (none to severe) or hemolytic disease of the fetus and newborn (none to mild). Despite challenging conditions caused by the scarcity of Lan- donors worldwide, Lan- blood should ideally be given to patients with anti-Lan, especially those with a high-titer antibody.


Saison C.,National Institute of Blood Transfusion INTS | Helias V.,National Institute of Blood Transfusion INTS | Peyrard T.,National Institute of Blood Transfusion INTS | Peyrard T.,National Reference Center for Blood Groups | And 3 more authors.
Vox Sanguinis | Year: 2013

Background and Objective The membrane transporter ABCB6 has recently been shown to carry the high-frequency red-blood-cell (RBC) antigen Lan. All the Lan- individuals genotyped so far have inherited two recessive null mutations in ABCB6. The finding of a family with the Lan- blood type occurring in two successive generations prompted this study. Methods Mutations in ABCB6 were searched by Sanger sequencing of exons and flanking intronic regions. Expression analysis of the Lan antigen was carried out by serology and flow cytometry. PCR-RFLP genotyping and Western blot analysis were also applied. Results All the Lan- members of this family were homozygous for c.574C>T, p.Arg192Trp in ABCB6 while the Lan+ members were heterozygous for this missense mutation encoded by the SNP rs149202834. Homozygosity for p.Arg192Trp was associated not only with absence of the Lan antigen, but also of the ABCB6 transporter in RBC membrane. The complete absence of Lan expression resulting from p.Arg192Trp homozygosity was confirmed by the subsequent identification of five unrelated Lan- individuals who were homozygous for this mutation and who developed an anti-Lan. We also provide evidence that three other single amino acid mutations in ABCB6 (c.826C>T, p.Arg276Trp; c.85_87delTTC, p.Phe29del; c.1762G>A, p.Gly588Ser) may also define ABCB6 null alleles. Conclusion p.Arg192Trp is the first ABCB6 missense mutation causing the Lan- blood type and appears to be a relatively frequent cause of this rare blood type. Like the previously reported frameshift, nonsense and essential splice-site mutations in ABCB6, this missense mutation is recessive and defines an ABCB6 null allele. Other single amino acid mutations in ABCB6 may also cause the Lan- blood type. © 2012 International Society of Blood Transfusion.


Ballif B.A.,University of Vermont | Helias V.,National Institute of Blood Transfusion INTS | Peyrard T.,National Institute of Blood Transfusion INTS | Peyrard T.,National Reference Center for Blood Groups | And 9 more authors.
EMBO Molecular Medicine | Year: 2013

Here, we report the biochemical and genetic basis of the Vel blood group antigen, which has been a vexing mystery for decades, especially as anti-Vel regularly causes severe haemolytic transfusion reactions. The protein carrying the Vel blood group antigen was biochemically purified from red blood cell membranes. Mass spectrometry-based de novo peptide sequencing identified this protein to be small integral membrane protein 1 (SMIM1), a previously uncharacterized single-pass membrane protein. Expression of SMIM1 cDNA in Vel- cultured cells generated anti-Vel cell surface reactivity, confirming that SMIM1 encoded the Vel blood group antigen. A cohort of 70 Vel- individuals was found to be uniformly homozygous for a 17 nucleotide deletion in the coding sequence of SMIM1. The genetic homogeneity of the Vel- blood type, likely having a common origin, facilitated the development of two highly specific DNA-based tests for rapid Vel genotyping, which can be easily integrated into blood group genotyping platforms. These results answer a 60-year-old riddle and provide tools of immediate assistance to all clinicians involved in the care of Vel- patients. © 2013.


Helias V.,National Institute of Blood Transfusion INTS | Saison C.,National Institute of Blood Transfusion INTS | Ballif B.A.,University of Vermont | Peyrard T.,National Institute of Blood Transfusion INTS | And 15 more authors.
Nature Genetics | Year: 2012

The human ATP-binding cassette (ABC) transporter ABCB6 has been described as a mitochondrial porphyrin transporter essential for heme biosynthesis, but it is also suspected to contribute to anticancer drug resistance, as do other ABC transporters located at the plasma membrane. We identified ABCB6 as the genetic basis of the Lan blood group antigen expressed on red blood cells but also at the plasma membrane of hepatocellular carcinoma (HCC) cells, and we established that ABCB6 encodes a new blood group system (Langereis, Lan). Targeted sequencing of ABCB6 in 12 unrelated individuals of the Lan(-) blood type identified 10 different ABCB6 null mutations. This is the first report of deficient alleles of this human ABC transporter gene. Of note, Lan(-) (ABCB6 -/-) individuals do not suffer any clinical consequences, although their deficiency in ABCB6 may place them at risk when determining drug dosage. © 2012 Nature America, Inc. All rights reserved.


Le Toriellec E.,National Institute of Blood Transfusion INTS | Chenet C.,National Institute of Blood Transfusion INTS | Kaplan C.,National Institute of Blood Transfusion INTS
Transfusion | Year: 2013

Background Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is due to maternal alloimmunization against fetal platelet (PLT) antigens. Antenatal management strategies have been developed to avoid complications such as intracranial hemorrhage. The aim of this study was to set up two reliable, noninvasive fetal genotyping assays to determine the fetal risk in pregnancies in which the father is heterozygous for the offending antigen. This study focused on human PLT antigen (HPA)-1, the most frequently implicated antigen in FNAIT in Caucasians. Study Design and Methods Two assays based on cell-free fetal DNA extracted from maternal blood samples and on real-time polymerase chain reaction (QPCR) were developed: an allele-specific QPCR specifically targeting the polymorphic sequence in HPA-1 and the study of the variation in the high-resolution melting curve of amplicons containing the polymorphic region. Results All results from the 49 samples obtained from 29 pregnant women were consistent with expectations. Six women were compatible with their fetuses (three HPA-1aa women and three HPA-1bb women), 41 HPA-1bb women were incompatible with their fetuses, as were two HPA-1aa women. Conclusion Two fetal PLT genotyping assays on maternal blood samples proved to be reliable as of 15 weeks of gestation, thereby avoiding invasive techniques such as amniocentesis. © 2012 American Association of Blood Banks.


PubMed | National Institute of Blood Transfusion INTS, PathoQuest and Tenon Hospital
Type: Journal Article | Journal: Blood transfusion = Trasfusione del sangue | Year: 2016

Characterisation of human-associated viral communities is essential for epidemiological surveillance and to be able to anticipate new potential threats for blood transfusion safety. In high-resource countries, the risk of blood-borne agent transmission of well-known viruses (HBV, HCV, HIV and HTLV) is currently considered to be under control. However, other unknown or unsuspected viruses may be transmitted to recipients by blood-derived products. To investigate this, the virome of plasma from individuals at high risk for parenterally and sexually transmitted infections was analysed by high throughput sequencing (HTS).Purified nucleic acids from two pools of 50 samples from recipients of multiple transfusions, and three pools containing seven plasma samples from either HBV-, HCV- or HIV-infected blood donors, were submitted to HTS.Sequences from resident anelloviruses and HPgV were evidenced in all pools. HBV and HCV sequences were detected in pools containing 3.810(3) IU/mL of HBV-DNA and 1.710(5) IU/mL of HCV-RNA, respectively, whereas no HIV sequence was found in a pool of 150 copies/mL of HIV-RNA. This suggests a lack of sensitivity in HTS performance in detecting low levels of virus. In addition, this study identified other issues, including laboratory contaminants and the uncertainty of taxonomic assignment of short sequence. No sequence suggestive of a new viral species was identified.This study did not identify any new blood-borne virus in high-risk individuals. However, rare and/or viruses present at very low titre could have escaped our protocol. Our results demonstrate the positive contribution of HTS in the detection of viral sequences in blood donations.


PubMed | National Institute of Blood Transfusion INTS
Type: Journal Article | Journal: Nature genetics | Year: 2012

The breast cancer resistance protein, also known as ABCG2, is one of the most highly studied ATP-binding cassette (ABC) transporters because of its ability to confer multidrug resistance. The lack of information on the physiological role of ABCG2 in humans severely limits cancer chemotherapeutic approaches targeting this transporter. We report here that ABCG2 comprises the molecular basis of a new blood group system (Junior, Jr) and that individuals of the Jr(a-) blood type have inherited two null alleles of ABCG2. We identified five frameshift and three nonsense mutations in ABCG2. We also show that the prevalence of the Jr(a-) blood type in the Japanese and European Gypsy populations is related to the p.Gln126* and p.Arg236* protein alterations, respectively. The identification of ABCG2(-/-) (Jr(a-)) individuals who appear phenotypically normal is an essential step toward targeting ABCG2 in cancer and also in understanding the physiological and pharmacological roles of this promiscuous transporter in humans.


PubMed | National Institute of Blood Transfusion INTS
Type: Case Reports | Journal: Vox sanguinis | Year: 2013

The membrane transporter ABCB6 has recently been shown to carry the high-frequency red-blood-cell (RBC) antigen Lan. All the Lan- individuals genotyped so far have inherited two recessive null mutations in ABCB6. The finding of a family with the Lan- blood type occurring in two successive generations prompted this study.Mutations in ABCB6 were searched by Sanger sequencing of exons and flanking intronic regions. Expression analysis of the Lan antigen was carried out by serology and flow cytometry. PCR-RFLP genotyping and Western blot analysis were also applied.All the Lan- members of this family were homozygous for c.574C>T, p.Arg192Trp in ABCB6 while the Lan+ members were heterozygous for this missense mutation encoded by the SNP rs149202834. Homozygosity for p.Arg192Trp was associated not only with absence of the Lan antigen, but also of the ABCB6 transporter in RBC membrane. The complete absence of Lan expression resulting from p.Arg192Trp homozygosity was confirmed by the subsequent identification of five unrelated Lan- individuals who were homozygous for this mutation and who developed an anti-Lan. We also provide evidence that three other single amino acid mutations in ABCB6 (c.826C>T, p.Arg276Trp; c.85_87delTTC, p.Phe29del; c.1762G>A, p.Gly588Ser) may also define ABCB6 null alleles.p.Arg192Trp is the first ABCB6 missense mutation causing the Lan- blood type and appears to be a relatively frequent cause of this rare blood type. Like the previously reported frameshift, nonsense and essential splice-site mutations in ABCB6, this missense mutation is recessive and defines an ABCB6 null allele. Other single amino acid mutations in ABCB6 may also cause the Lan- blood type.

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