ABCB6 is dispensable for erythropoiesis and specifies the new blood group system Langereis

Journal name:
Nature Genetics
Year published:
Published online

The human ATP-binding cassette (ABC) transporter ABCB6 has been described as a mitochondrial porphyrin transporter essential for heme biosynthesis1, but it is also suspected to contribute to anticancer drug resistance2, 3, 4, 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.

At a glance


  1. Characterization of the Lan blood group antigen expression on RBCs with the OSK43 monoclonal antibody.
    Figure 1: Characterization of the Lan blood group antigen expression on RBCs with the OSK43 monoclonal antibody.

    (a) Specificity of the OSK43 monoclonal antibody as shown by flow cytometry analysis of Lan(+) and Lan(−) RBCs. OSK43 shows a strict Lan specificity, such that the histogram profiles of Lan(−) RBCs labeled with and without OSK43 were superimposable. (b) Immunofluorescence analysis of Lan(+) and Lan(−) RBCs co-labeled with the OSK43 human monoclonal antibody (green) and a mouse monoclonal antibody to glycophorin A, the most abundant glycoprotein on RBCs (red). Scale bars, 5 μm. (c) Lan antigen expression on RBCs from 39 random blood donors from France (ABO type: O), as measured by flow cytometry with OSK43 as in a. The geometric means of fluorescence intensity (in arbitrary units of the flow cytometer) are shown as a box-and-whisker diagram, with the lower and upper boundaries of the boxes representing the 25th and 75th percentiles, respectively. Horizontal lines, medians; whiskers, the minimum and maximum non-atypical values; circles, the atypical values. (d) Lan antigen expression on RBCs from adult or cord blood, as measured by flow cytometry with OSK43 as in a. Bars, mean fluorescence intensity; error bars, s.d. (N = 4 individuals); **P < 0.01 by Mann-Whitney U test.

  2. Identification of ABCB6 as the gene encoding the Lan blood group antigen.
    Figure 2: Identification of ABCB6 as the gene encoding the Lan blood group antigen.

    (a) Purification of the Lan antigen from RBCs by immunoprecipitation with OSK43. Lysates were prepared from the membranes of Lan(+) RBCs labeled with or without OSK43, and the corresponding immune complexes were analyzed by PAGE and silver staining. The position of the heavy (Ig(γ1)) and light (Ig(κ)) chains of OSK43 are indicated. (b) ABCB6 peptides identified by mass spectrometry from the band of ~80 kDa that was immunoprecipitated with OSK43 from RBCs. In total, 27 tryptic peptides of ABCB6 were identified (green); the predicted trypsin cleavage sites are underlined. (c) ABCB6 is present in the plasma membrane of RBCs from Lan(+) but not Lan(−) individuals, as shown by protein blot analysis. RBC membranes were prepared from two Lan(+) and six Lan(−) individuals and analyzed by PAGE, under reducing conditions without heat denaturation, and protein blotting with a rabbit polyclonal antibody to ABCB6 (top). ABCB6 runs as monomeric and multimeric forms, even under reducing conditions (arrows; see Supplementary Fig. 6 for details). The protein blot membrane was reprobed with a mouse monoclonal antibody to ABCG2, another ABC transporter able to transport porphyrin (bottom; only the band corresponding to a monomeric form of ABCG2 is shown; see Supplementary Fig. 6). (d) Exogenous expression of ABCB6 in K-562 cells results in cell surface expression of the Lan antigen. Live cells of representative K-562 clones stably transfected with an expression construct of ABCB6 cDNA (green) or the corresponding empty vector (red) were analyzed by flow cytometry with OSK43. (e) Diagram showing the positions of ABCB6 null mutations identified in this study (see Table 1 for details). Green boxes, exons; broken lines, introns; red diamonds at the top, the number of unrelated Lan(−) subjects with each mutation.

  3. Expression of the Lan antigen on human cancer cell lines.
    Figure 3: Expression of the Lan antigen on human cancer cell lines.

    (af) Cell surface expression of the Lan antigen was analyzed on native HepG2 (a), A-498 (b), A-431 (c), MOLT-4 (d), HeLa (e) and HuH-7 (f) cells by flow cytometry with OSK43 (green) or T27S (a human monoclonal antibody to an unrelated antigen; gray).


  1. Krishnamurthy, P.C. et al. Identification of a mammalian mitochondrial porphyrin transporter. Nature 443, 586589 (2006).
  2. Szakács, G. et al. Predicting drug sensitivity and resistance: profiling ABC transporter genes in cancer cells. Cancer Cell 6, 129137 (2004).
  3. Kelter, G. et al. Role of transferrin receptor and the ABC transporters ABCB6 and ABCB7 for resistance and differentiation of tumor cells towards artesunate. PLoS ONE 2, e798 (2007).
  4. Yasui, K. et al. Alteration in copy numbers of genes as a mechanism for acquired drug resistance. Cancer Res. 64, 14031410 (2004).
  5. Cartron, J.-P. Blood group antigens structure and function: recent advances. in Hematology Education: the Education Program for the Annual Congress of the European Hematology Association Vol. 2, 158174 (2008).
  6. Daniels, G. The molecular genetics of blood group polymorphism. Hum. Genet. 126, 729742 (2009).
  7. van der Hart, M., Moes, M., van der Veer, M. & van Loghem, J.J. Ho and Lan—two new blood group antigens. in Proceedings of the 8th Congress of the European Society of Haematology 493 (Vienna, 1962).
  8. Smith, D.S. et al. Haemolytic disease of the newborn caused by anti-Lan antibody. BMJ 3, 9092 (1969).
  9. Page, P.L. Hemolytic disease of the newborn due to anti-Lan. Transfusion 23, 256257 (1983).
  10. Okubo, Y. et al. The rare red cell phenotype Lan negative in Japanese. Transfusion 24, 534535 (1984).
  11. Holland, I.B., Cole, S.P.C., Kuchler, K. & Higgins, C.F. ABC Proteins, From Bacteria to Man (Academic Press, New York, 2003).
  12. Paterson, J.K. et al. Human ABCB6 localizes to both the outer mitochondrial membrane and the plasma membrane. Biochemistry 46, 94439452 (2007).
  13. Borst, P. & Elferink, R.O. Mammalian ABC transporters in health and disease. Annu. Rev. Biochem. 71, 537592 (2002).
  14. Szakács, G., Paterson, J.K., Ludwig, J.A., Booth-Genthe, C. & Gottesman, M.M. Targeting multidrug resistance in cancer. Nat. Rev. Drug Discov. 5, 219234 (2006).
  15. Puy, H., Gouya, L. & Deybach, J.C. Porphyrias. Lancet 375, 924937 (2010).
  16. Zhou, S. et al. Increased expression of the Abcg2 transporter during erythroid maturation plays a role in decreasing cellular protoporphyrin IX levels. Blood 105, 25712576 (2005).
  17. Saison, C. et al. Null alleles of ABCG2 encoding the breast cancer resistance protein define the new blood group system Junior. Nat. Genet. published online (15 January 2012), doi:10.1038/ng.1070.
  18. Emadi-Konjin, H.P. et al. Isolation of a genomic clone containing the promoter region of the human ATP binding cassette (ABC) transporter, ABCB6. Biochim. Biophys. Acta 1574, 117130 (2002).
  19. Ballif, B.A., Carey, G.R., Sunyaev, S.R. & Gygi, S.P. Large-scale identification and evolution indexing of tyrosine phosphorylation sites from murine brain. J. Proteome Res. 7, 311318 (2008).
  20. Elias, J.E. & Gygi, S.P. Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry. Nat. Methods 4, 207214 (2007).
  21. Gouya, L. et al. Contribution of a common single-nucleotide polymorphism to the genetic predisposition for erythropoietic protoporphyria. Am. J. Hum. Genet. 78, 214 (2006).

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Author information


  1. National Institute of Blood Transfusion (INTS), Paris, France.

    • Virginie Helias,
    • Carole Saison,
    • Thierry Peyrard,
    • Camille Sureau,
    • Bach-Nga Pham,
    • Pierre-Yves Le Pennec,
    • Jean-Pierre Cartron &
    • Lionel Arnaud
  2. Department of Biology, University of Vermont, Burlington, Vermont, USA.

    • Bryan A Ballif
  3. National Reference Center for Blood Groups (CNRGS), Paris, France.

    • Thierry Peyrard,
    • Bach-Nga Pham &
    • Pierre-Yves Le Pennec
  4. Japanese Red Cross Osaka Blood Center, Osaka, Japan.

    • Junko Takahashi,
    • Hideo Takahashi,
    • Mitsunobu Tanaka &
    • Yoshihiko Tani
  5. Assistance Publique–Hôpitaux de Paris, Centre Français des Porphyries, Institut National de la Santé et de la Recherche Médicale (Inserm), Centre de Recherche Biomédicale Bichat-Beaujon, U 773, Université Paris Diderot–Paris 7, Paris, France.

    • Jean-Charles Deybach &
    • Hervé Puy
  6. Centre de Recherche des Cordeliers, Inserm, Université Pierre et Marie Curie–Paris 6, Unité Mixte de Recherche (UMR) S 872, Paris, France.

    • Maude Le Gall


V.H. carried out flow cytometry and protein blot analysis, made expression constructs and performed cell culture. C. Saison performed immunoprecipitation and genomic DNA sequencing. B.A.B. performed mass spectrometry analysis. T.P. provided immunohematological information and most Lan(−) blood samples. J.T., H.T. and M.T. isolated and produced the OSK43 monoclonal antibody. Y.T. conceived and supervised the study at the Japanese Red Cross Osaka Blood Center. H.P. performed porphyrin analysis. M.L.G. performed statistical analysis and provided human cell lines. C. Sureau provided the HuH-7 cell line. B.-N.P. provided two Lan(−) blood samples. P.-Y.L.P. was a former chief operating officer of the CNRGS involved in identifying and cryopreserving Lan(–) blood samples. J.-P.C. initiated the study conducted by L.A. by obtaining OSK43 from Y.T., contacted J.-C.D. for porphyrin analysis and continuously supported the study. L.A. conceived and supervised the study at the INTS, performed experiments, made the figures and wrote the manuscript, which was reviewed by J.-P.C., T.P., M.L.G., H.P., C. Sureau and B.A.B. All authors approved the submitted manuscript.

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    Supplementary Figures 1–6 and Supplementary Table 1

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