Mutations in ABCD4 cause a new inborn error of vitamin B12 metabolism

Abstract

Inherited disorders of vitamin B12 (cobalamin) have provided important clues to how this vitamin, which is essential for hematological and neurological function, is transported and metabolized. We describe a new disease that results in failure to release vitamin B12 from lysosomes, which mimics the cblF defect caused by LMBRD1 mutations. Using microcell-mediated chromosome transfer and exome sequencing, we identified causal mutations in ABCD4, a gene that codes for an ABC transporter, which was previously thought to have peroxisomal localization and function. Our results show that ABCD4 colocalizes with the lysosomal proteins LAMP1 and LMBD1, the latter of which is deficient in the cblF defect. Furthermore, we show that mutations altering the putative ATPase domain of ABCD4 affect its function, suggesting that the ATPase activity of ABCD4 may be involved in intracellular processing of vitamin B12.

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Figure 1: Distribution of free and protein-bound cobalamin in cell lines from controls and affected individuals.
Figure 2: Somatic complementation analysis confirms a new cobalamin complementation group cblJ.
Figure 3: Expression of wild-type and mutant ABCD4 alleles.
Figure 4: ABCD4 is a membrane protein with ATPase function.
Figure 5: Subcellular localization of ABCD4 detected by fluorescence and confocal microscopy.

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Acknowledgements

We thank S. Lutz for technical assistance in carrying out enzyme assays and K. Locher for fruitful discussions. We thank J. Gärtner (University of Göttingen) for providing cell lines with peroxisomal defects and R. Newbold (Brunel University) for providing the mouse-human monochromosomal hybrid cell lines used for microcell-mediated chromosome transfer. This work was supported by the Swiss National Science Foundation (320000_122568 and 31003A_138521) and by the Deutsche Forschungsgemeinschaft (RU816/5-1). D.S.R. is supported by the Canadian Institutes for Health Research (15078).

Author information

M.R.B., F.R., B.F. and D.S.R. supervised the project. M.R.B., F.R., B.F., D.S.R., J.C.K., T.S. and D.C. designed the study, analyzed the data and wrote the manuscript. T.S., M.d.M. and D.C. performed microcell-mediated chromosome transfer. M.S., D.C. and M.F. designed primers and built constructs. M.d.M., I.B., J.C.K., I.R.M., D.W. and D.C. performed sequencing analysis. J.M., P.N. and H.T. performed whole-exome sequencing. S.F. and E.A.S. performed immortalization and stable transduction of fibroblasts. T.S., P.B., J.C.K. and D.W. performed somatic cell complementation, enzymatic assays, Cbl coenzyme synthesis and expression studies. H.R., I.B. and D.C. analyzed the subcellular localization of ABCD4. D.C. and P.B. performed protein blots. W.H. and D.C. analyzed the structure of ABCD4. N.L., M.P. and E.M. provided clinical information about the affected individuals. All authors discussed the results and reviewed the manuscript.

Correspondence to David S Rosenblatt or Brian Fowler.

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Supplementary Figures 1 and 2, Supplementary Tables 1–7 and Supplementary Note (PDF 126 kb)

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Coelho, D., Kim, J., Miousse, I. et al. Mutations in ABCD4 cause a new inborn error of vitamin B12 metabolism. Nat Genet 44, 1152–1155 (2012). https://doi.org/10.1038/ng.2386

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