Folding defects in P-type ATP 8B1 associated with hereditary cholestasis are ameliorated by 4-phenylbutyrate

Authors

  • Lieke M. van der Velden,

    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
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    • These authors contributed equally to this work.

  • Janneke M. Stapelbroek,

    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
    2. Department of Pediatric Gastroenterology, UMC Utrecht, Utrecht, The Netherlands
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    • These authors contributed equally to this work.

  • Elmar Krieger,

    1. Centre for Molecular and Biomolecular Informatics, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands
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  • Peter V. E. van den Berghe,

    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
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  • Ruud Berger,

    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
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  • Patricia M. Verhulst,

    1. Department of Membrane Enzymology, Bijvoet Center and Institute of Biomembranes, Utrecht University, The Netherlands
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  • Joost C. M. Holthuis,

    1. Department of Membrane Enzymology, Bijvoet Center and Institute of Biomembranes, Utrecht University, The Netherlands
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  • Roderick H. J. Houwen,

    1. Department of Pediatric Gastroenterology, UMC Utrecht, Utrecht, The Netherlands
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  • Leo W. J. Klomp,

    Corresponding author
    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
    • University Medical Center Utrecht, Department of Metabolic and Endocrine Diseases, Room KC-02.069.1, P.O. Box 85090, 3508 AB Utrecht, The Netherlands
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    • These authors have equal senior authorship

    • fax: +31 30 2504295.

  • Stan F. J. van de Graaf

    1. Department of Metabolic and Endocrine Diseases, University Medical Center (UMC) Utrecht, and Netherlands Metabolomics Centre, Utrecht, The Netherlands
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    • These authors have equal senior authorship


  • Potential conflict of interest: Nothing to report.

Abstract

Deficiency in P-type ATP8B1 is a severe and clinically highly variable hereditary disorder that is primarily characterized by intrahepatic cholestasis. It presents either as a progressive (progressive familial intrahepatic cholestasis type 1 [PFIC1]) or intermittent (benign recurrent intrahepatic cholestasis type 1 [BRIC1]) disease. ATP8B1 deficiency is caused by autosomal recessive mutations in the gene encoding ATP8B1, a putative aminophospholipid-translocating P-type adenosine triphosphatase. The exact pathogenesis of the disease is elusive, and no effective pharmacological therapy is currently available. Here, the molecular consequences of six distinct ATP8B1 missense mutations (p.L127P, p.G308V, p.D454G, p.D554N, p.I661T, and p.G1040R) and one nonsense mutation (p.R1164X) associated with PFIC1 and/or BRIC1 were systematically characterized. Except for the p.L127P mutation, all mutations resulted in markedly reduced ATP8B1 protein expression, whereas messenger RNA expression was unaffected. Five of seven mutations resulted in (partial) retention of ATP8B1 in the endoplasmic reticulum. Reduced protein expression was partially restored by culturing the cells at 30°C and by treatment with proteasomal inhibitors, indicating protein misfolding and subsequent proteosomal degradation. Protein misfolding was corroborated by predicting the consequences of most mutations onto a homology model of ATP8B1. Treatment with 4-phenylbutyrate, a clinically approved pharmacological chaperone, partially restored defects in expression and localization of ATP8B1 substitutions G308V, D454G, D554N, and in particular I661T, which is the most frequently identified mutation in BRIC1. Conclusion: A surprisingly large proportion of ATP8B1 mutations resulted in aberrant folding and decreased expression at the plasma membrane. These effects were partially restored by treatment with 4-phenylbutyrate. We propose that treatment with pharmacological chaperones may represent an effective therapeutic strategy to ameliorate the recurrent attacks of cholestasis in patients with intermittent (BRIC1) disease. (HEPATOLOGY 2009.)

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