ABCMdb

ABCD1 p.Ala616Thr

Predicted by SNAP2:	C: D (80%), D: D (95%), E: D (95%), F: D (95%), G: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (95%), M: D (95%), N: D (95%), P: D (95%), Q: D (95%), R: D (95%), S: D (91%), T: D (80%), V: D (95%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: N, T: D, V: D, W: D, Y: D,

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[hide] Conservation of targeting but divergence in functi... Biochem J. 2011 Jun 15;436(3):547-57. Zhang X, De Marcos Lousa C, Schutte-Lensink N, Ofman R, Wanders RJ, Baldwin SA, Baker A, Kemp S, Theodoulou FL
Conservation of targeting but divergence in function and quality control of peroxisomal ABC transporters: an analysis using cross-kingdom expression.
Biochem J. 2011 Jun 15;436(3):547-57., [PMID:21476988]

Abstract [show]
ABC (ATP-binding cassette) subfamily D transporters are found in all eukaryotic kingdoms and are known to play essential roles in mammals and plants; however, their number, organization and physiological contexts differ. Via cross-kingdom expression experiments, we have explored the conservation of targeting, protein stability and function between mammalian and plant ABCD transporters. When expressed in tobacco epidermal cells, the mammalian ABCD proteins ALDP (adrenoleukodystrophy protein), ALDR (adrenoleukodystrophy-related protein) and PMP70 (70 kDa peroxisomal membrane protein) targeted faithfully to peroxisomes and P70R (PMP70-related protein) targeted to the ER (endoplasmic reticulum), as in the native host. The Arabidopsis thaliana peroxin AtPex19_1 interacted with human peroxisomal ABC transporters both in vivo and in vitro, providing an explanation for the fidelity of targeting. The fate of X-linked adrenoleukodystrophy disease-related mutants differed between fibroblasts and plant cells. In fibroblasts, levels of ALDP in some 'protein-absent' mutants were increased by low-temperature culture, in some cases restoring function. In contrast, all mutant ALDP proteins examined were stable and correctly targeted in plant cells, regardless of their fate in fibroblasts. ALDR complemented the seed germination defect of the Arabidopsis cts-1 mutant which lacks the peroxisomal ABCD transporter CTS (Comatose), but neither ALDR nor ALDP was able to rescue the defect in fatty acid beta-oxidation in establishing seedlings. Taken together, our results indicate that the mechanism for trafficking of peroxisomal membrane proteins is shared between plants and mammals, but suggest differences in the sensing and turnover of mutant ABC transporter proteins and differences in substrate specificity and/or function.

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153 Approximately 60% of X-ALD ABCD1 mutations are missense mutations, 65% of which result in no detectable ALDP, based on IF (immunofluorescence), indicating that they affect protein Table 1 Quantification of ALDP levels in X-ALD fibroblasts ALDP Mutation IF Immunoblot (% of control) p.Arg74Trp Absent 7.5 + - 0.6 p.Arg104Cys Reduced 35 + - 3.0 p.Ser149Asn Present 77 + - 3.0 p.Asp194His Present 60 + - 13.6 p.Leu220Pro Reduced 21.8 + - 5.4 p.Arg389His Present 40.6 + - 3.6 p.Arg554His Absent 1.0 + - 0.5 p.Ser606Leu Present 25 + - 1.5 p.Glu609Gly Absent 2.1 + - 1.3 p.Glu609Lys Absent 1.8 + - 0.9 p.Ala616Thr Absent 4.3 + - 1.7 p.Leu654Pro Absent 1.5 + - 1.3 p.Arg660Trp Absent 1.6 + - 0.8 p.His667Asp Absent 2.9 + - 1.0 p.Arg113fs Absent - Figure 3 Interaction of mammalian ABCD proteins with Arabidopsis Pex19 in vivo Tobacco plants stably expressing CFP-SKL were co-transfected with 35S::ABCD-YFP fusions andNLS-Pex19constructs.Leafepidermalcellswereimagedusingconfocalmicroscopy:(A-D) ALDP-YFP plus NLS-HsPex19; (E-H) ALDP-YFP plus NLS-AtPex19_1; (I-L) ALDR-YFP plus NLS-AtPex19_1. Login to comment
169 Results are means + - S.D.; n = 3. significantly in response to low temperature in ten wild-type control lines tested (see Supplementary Figure S3A at http://www.BiochemJ.org/bj/436/bj4360547add.htm); however, increased expression levels of ALDP were found in several of the X-ALD fibroblasts investigated: p.Arg74Cys, p.Arg104Cys, p.Arg554His, p.Glu609Gly, p.Ala616Thr, p.Leu654Pro and p.Arg660Trp (Figures 4A and 4B). Login to comment
172 ALDP was increased from 2-4% to ~20% of wild-type levels in cell lines bearing the mutations p.Glu609Gly, p.Ala616Thr and p.Arg660Trp, from 1 to 10% in p.Glu609Lys and p.Arg554His cells and from 45 to 75% in the p.Asp194His cell line (Figure 4A). Login to comment
173 VLCFA β-oxidation was measured in cells that were cultured at 30◦ C for 72 h, but in only one case (p.Ala616Thr) was the capacity to degrade VLCFA restored to near-control levels (Figure 4C). Login to comment
174 However, after 4 weeks of culture at 30◦ C, VLCFA levels were partially corrected in p.Arg660Trp, p.Arg554His and p.Ala616Thr fibroblasts. Login to comment
154 Approximately 60% of X-ALD ABCD1 mutations are missense mutations, 65% of which result in no detectable ALDP, based on IF (immunofluorescence), indicating that they affect protein Table 1 Quantification of ALDP levels in X-ALD fibroblasts ALDP Mutation IF Immunoblot (% of control) p.Arg74Trp Absent 7.5 + - 0.6 p.Arg104Cys Reduced 35 + - 3.0 p.Ser149Asn Present 77 + - 3.0 p.Asp194His Present 60 + - 13.6 p.Leu220Pro Reduced 21.8 + - 5.4 p.Arg389His Present 40.6 + - 3.6 p.Arg554His Absent 1.0 + - 0.5 p.Ser606Leu Present 25 + - 1.5 p.Glu609Gly Absent 2.1 + - 1.3 p.Glu609Lys Absent 1.8 + - 0.9 p.Ala616Thr Absent 4.3 + - 1.7 p.Leu654Pro Absent 1.5 + - 1.3 p.Arg660Trp Absent 1.6 + - 0.8 p.His667Asp Absent 2.9 + - 1.0 p.Arg113fs Absent - Figure 3 Interaction of mammalian ABCD proteins with Arabidopsis Pex19 in vivo Tobacco plants stably expressing CFP-SKL were co-transfected with 35S::ABCD-YFP fusions andNLS-Pex19constructs.Leafepidermalcellswereimagedusingconfocalmicroscopy:(A-D) ALDP-YFP plus NLS-HsPex19; (E-H) ALDP-YFP plus NLS-AtPex19_1; (I-L) ALDR-YFP plus NLS-AtPex19_1. Login to comment
170 Results are means + - S.D.; n = 3. significantly in response to low temperature in ten wild-type control lines tested (see Supplementary Figure S3A at http://www.BiochemJ.org/bj/436/bj4360547add.htm); however, increased expression levels of ALDP were found in several of the X-ALD fibroblasts investigated: p.Arg74Cys, p.Arg104Cys, p.Arg554His, p.Glu609Gly, p.Ala616Thr, p.Leu654Pro and p.Arg660Trp (Figures 4A and 4B). Login to comment
175 However, after 4 weeks of culture at 30e6; C, VLCFA levels were partially corrected in p.Arg660Trp, p.Arg554His and p.Ala616Thr fibroblasts. Login to comment

[hide] X-linked adrenoleukodystrophy in women: a cross-se... Brain. 2014 Mar;137(Pt 3):693-706. doi: 10.1093/brain/awt361. Epub 2014 Jan 29. Engelen M, Barbier M, Dijkstra IM, Schur R, de Bie RM, Verhamme C, Dijkgraaf MG, Aubourg PA, Wanders RJ, van Geel BM, de Visser M, Poll-The BT, Kemp S
X-linked adrenoleukodystrophy in women: a cross-sectional cohort study.
Brain. 2014 Mar;137(Pt 3):693-706. doi: 10.1093/brain/awt361. Epub 2014 Jan 29., [PMID:24480483]

Abstract [show]
X-linked adrenoleukodystrophy is the most common peroxisomal disorder. The disease is caused by mutations in the ABCD1 gene that encodes the peroxisomal transporter of very long-chain fatty acids. A defect in the ABCD1 protein results in elevated levels of very long-chain fatty acids in plasma and tissues. The clinical spectrum in males with X-linked adrenoleukodystrophy has been well described and ranges from isolated adrenocortical insufficiency and slowly progressive myelopathy to devastating cerebral demyelination. As in many X-linked diseases, it was assumed that female carriers remain asymptomatic and only a few studies addressed the phenotype of X-linked adrenoleukodystrophy carriers. These studies, however, provided no information on the prevalence of neurological symptoms in the entire population of X-linked adrenoleukodystrophy carriers, since data were acquired in small groups and may be biased towards women with symptoms. Our primary goal was to investigate the symptoms and their frequency in X-linked adrenoleukodystrophy carriers. The secondary goal was to determine if the X-inactivation pattern of the ABCD1 gene was associated with symptomatic status. We included 46 X-linked adrenoleukodystrophy carriers in a prospective cross-sectional cohort study. Our data show that X-linked adrenoleukodystrophy carriers develop signs and symptoms of myelopathy (29/46, 63%) and/or peripheral neuropathy (26/46, 57%). Especially striking was the occurrence of faecal incontinence (13/46, 28%). The frequency of symptomatic women increased sharply with age (from 18% in women <40 years to 88% in women >60 years of age). Virtually all (44/45, 98%) X-linked adrenoleukodystrophy carriers had increased very long-chain fatty acids in plasma and/or fibroblasts, and/or decreased very long-chain fatty acids beta-oxidation in fibroblasts. We did not find an association between the X-inactivation pattern and symptomatic status. We conclude that X-linked adrenoleukodystrophy carriers develop an adrenomyeloneuropathy-like phenotype and there is a strong association between symptomatic status and age. X-linked adrenoleukodystrophy should be considered in the differential diagnosis in women with chronic myelopathy and/or peripheral neuropathy (especially with early faecal incontinence). ABCD1 mutation analysis deserves a place in diagnostic protocols for chronic non-compressive myelopathy.

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163 Table 1 Continued Family Age (years) Urinary incontinence Faecal incontinence Gait disorder Sensory complaints Sensory disturbance Spasticity Weakness Pathological reflexes EDSS Mutation ABCD1 protein S 76 Yes No Yes No Yes No No Yes 2.0 p.His283Tyr Reduced T 51 Yes Yes Yes No Yes No Yes Yes 4.0 p.Gln177* Absent U 47 Yes Yes No No Yes No No No 2.0 p.Arg464* Absent V 56 Yes No Yes No Yes No Yes Yes 2.5 p.Asp442Glyfs Absent W 45 Yes Yes Yes Yes Yes No No Yes 3.5 p.Ala616Thr Absent W 65 No Yes No No No No No No 2.0 p.Ala616Thr Absent X 47 Yes No Yes No No No No Yes 2.5 p.Arg113Alafs Absent Y 24 No No No No No No No No 0 p.Glu609Gly a Absent a Z 50 No No No No Yes No No Yes 2.0 p.Ser633Argfs Absent Age = age at examination; urinary incontinence = urge incontinence; faecal incontinence = soiling and urge incontinence; gait = gait disorder, for instance unable to run; sensory complaints = numbness or tingling; sensory disturbance = sensory symptoms on neurological examinations; spasticity = spasticity of the lower extremities; weakness = paresis of the lower extremities, Pathological reflexes: pathological reflexes in the lower extremities, EDSS = expanded disability status scale; mutation = mutation in ABCD1; ABCD1 protein = effect of mutation in ABCD1 on ABCD1 protein as determined by immunoblot. Login to comment