Abstract
Mutations in XPR1, a gene encoding an inorganic phosphate exporter, have recently been identified in patients with primary familial brain calcification (PFBC). Using Sanger sequencing, we screened XPR1 in 18 unrelated patients with PFBC and no SLC20A2, PDGFB, or PDGFRB mutation. XPR1 variants were tested in an in vitro physiological complementation assay and patient blood cells were assessed ex vivo for phosphate export. We identified a novel c.260T > C, p.(Leu87Pro) XPR1 variant in a 41-year-old man complaining of micrographia and dysarthria and demonstrating mild parkinsonism, cerebellar ataxia and executive dysfunction. Brain 123I-Ioflupane scintigraphy showed marked dopaminergic neuron loss. Peripheral blood cells from the patient exhibited decreased phosphate export. XPR1 in which we introduced the mutation was not detectable at the cell surface and did not lead to phosphate export. These results confirm that loss of XPR1-mediated phosphate export function causes PFBC, occurring in less than 8 % of cases negative for the other genes, and may be responsible for parkinsonism.
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Acknowledgments
We are grateful to all collaborators who sent blood samples and medical charts, and all members of our labs for constant support and helpful discussions. This work was supported by grants from the Fondation pour la Recherche Médicale (FRM) and ANR Blanc (to M.S.); Ligue Nationale contre le Cancer (Comité de l’Hérault) and Agence Nationale de la Recherche JCJC (to J.-L.B.); U.L.-S. was supported by Labex EpiGenMed (ANR-10-LABX-12-01) and Secretaría de Ciencia, Tecnología e Innovación de la Ciudad de México (CM-102/15 SECITI 030/2016) fellowships; D.G. was supported by FRM, Institut National du Cancer (INCa), and Labex GR-Ex (ANR-11-LABX-0051) fellowships; Labex is funded by the program ‘Investissements d’Avenir’ of the French National Research Agency. J.-L.B. and M.S. are supported by INSERM; G.N., D.C., T.F., and D.H. are supported by Inserm, the University Hospital of Rouen and the French CNR-MAJ.
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J-L.B. and M.S. are inventors on provisional a patent describing the use of ligands, including XRBD, for the analysis of human cells (PCT/EP2010/050139); M.S. is a co-founder of METAFORA-biosystems, a start-up company that focuses on metabolite transporters under physiological and pathological conditions.
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M. Anheim, U. López-Sánchez contributed equally to the work.
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415_2016_8166_MOESM1_ESM.jpg
Supplementary material 1 Figure. a) Uptake of inorganic 33P (Pi) assay performed in HEK293T cells transfected with siRNA directed against either luciferase (siLUC) or XPR1 (siXPR1), or a combination of siXPR1 with an expression vector encoding HA-tagged WT XPR1 or the L87P mutant human XPR1. Results are shown as mean ± s.e.m. in a representative experiment (n = 3); b) Cell surface detection of PiT1 and PiT2 in HEK293T transfected cells as in (a). Specific binding with KoRBD, the PiT1 ligand derived from the receptor-binding domain of KoRV-MLV Env, or with ASU, the PiT2 ligand derived from the Amphotropic-MLV Env (grey histograms, upper and lower panels, respectively). Grey histograms represent the non-specific staining with the secondary IgG antibody. Numbers indicate the specific mean fluorescence intensity of a representative experiment (n = 3). (JPEG 295 kb)
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Anheim, M., López-Sánchez, U., Giovannini, D. et al. XPR1 mutations are a rare cause of primary familial brain calcification. J Neurol 263, 1559–1564 (2016). https://doi.org/10.1007/s00415-016-8166-4
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DOI: https://doi.org/10.1007/s00415-016-8166-4