title: Adrenoleukodystrophy: subcellular localization and degradation of adrenoleukodystrophy protein (ALDP/ABCD1) with naturally occurring missense mutations.
abstract: Mutation in the X-chromosomal adrenoleukodystrophy gene (ALD; ABCD1) leads to X-linked adrenoleukodystrophy (X-ALD), a severe neurodegenerative disorder. The encoded adrenoleukodystrophy protein (ALDP/ABCD1) is a half-size peroxisomal ATP-binding cassette protein of 745 amino acids in humans. In this study, we chose nine arbitrary mutant human ALDP forms (R104C, G116R, Y174C, S342P, Q544R, S606P, S606L, R617H, and H667D) with naturally occurring missense mutations and examined the intracellular behavior. When expressed in X-ALD fibroblasts lacking ALDP, the expression level of mutant His-ALDPs (S606L, R617H, and H667D) was lower than that of wild type and other mutant ALDPs. Furthermore, mutant ALDP-green fluorescence proteins (S606L and H667D) stably expressed in CHO cells were not detected due to rapid degradation. Interestingly, the wild type ALDP co-expressed in these cells also disappeared. In the case of X-ALD fibroblasts from an ALD patient (R617H), the mutant ALDP was not detected in the cells, but appeared upon incubation with a proteasome inhibitor. When CHO cells expressing mutant ALDP-green fluorescence protein (H667D) were cultured in the presence of a proteasome inhibitor, both the mutant and wild type ALDP reappeared. In addition, mutant His-ALDP (Y174C), which has a mutation between transmembrane domain 2 and 3, did not exhibit peroxisomal localization by immunofluorescense study. These results suggest that mutant ALDPs, which have a mutation in the COOH-terminal half of ALDP, including S606L, R617H, and H667D, were degraded by proteasomes after dimerization. Further, the region between transmembrane domain 2 and 3 is important for the targeting of ALDP to the peroxisome.
authors.affiliation: Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.
authors.forename: Norimasa
authors.initials: N
authors.lastname: Takahashi
authors.forename: Masashi
authors.initials: M
authors.lastname: Morita
authors.forename: Takanori
authors.initials: T
authors.lastname: Maeda
authors.forename: Yuta
authors.initials: Y
authors.lastname: Harayama
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Shimozawa
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Suzuki
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Furuya
authors.forename: Ryuichiro
authors.initials: R
authors.lastname: Sato
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Kashiwayama
authors.forename: Tsuneo
authors.initials: T
authors.lastname: Imanaka
mesh_terms: D000074063:ATP Binding Cassette Transporter, Subfamily D, Member 1; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D000326:Adrenoleukodystrophy / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D020125:Mutation, Missense; D061988:Proteasome Inhibitors; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: C115572:ABCD1 protein, human; D000074063:ATP Binding Cassette Transporter, Subfamily D, Member 1; D018528:ATP-Binding Cassette Transporters; D007976:Leupeptins; D061988:Proteasome Inhibitors; C067713:lactacystin; C032854:leupeptin; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D000111:Acetylcysteine