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Further characterization of ATP6V0A2-related autosomal recessive cutis laxa

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Abstract

Autosomal recessive cutis laxa (ARCL) syndromes are phenotypically overlapping, but genetically heterogeneous disorders. Mutations in the ATP6V0A2 gene were found to underlie both, autosomal recessive cutis laxa type 2 (ARCL2), Debré type, and wrinkly skin syndrome (WSS). The ATP6V0A2 gene encodes the a2 subunit of the V-type H+-ATPase, playing a role in proton translocation, and possibly also in membrane fusion. Here, we describe a highly variable phenotype in 13 patients with ARCL2, including the oldest affected individual described so far, who showed strikingly progressive dysmorphic features and heterotopic calcifications. In these individuals we identified 17 ATP6V0A2 mutations, 14 of which are novel. Furthermore, we demonstrate a localization of ATP6V0A2 at the Golgi-apparatus and a loss of the mutated ATP6V0A2 protein in patients’ dermal fibroblasts. Investigation of brefeldin A-induced Golgi collapse in dermal fibroblasts as well as in HeLa cells deficient for ATP6V0A2 revealed a delay, which was absent in cells deficient for the ARCL-associated proteins GORAB or PYCR1. Furthermore, fibroblasts from patients with ATP6V0A2 mutations displayed elevated TGF-β signalling and increased TGF-β1 levels in the supernatant. Our current findings expand the genetic and phenotypic spectrum and suggest that, besides the known glycosylation defect, alterations in trafficking and signalling processes are potential key events in the pathogenesis of ATP6V0A2-related ARCL.

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Acknowledgments

We are grateful to the patients and their family members whose cooperation made this study possible. We would like to thank the family of patient 2 especially for their great contribution and interest in our work. We thank Traute Burmester for her excellent help with fibroblast cultivation from skin biopsies. We additionally thank E. Ntrivalas for providing the ATP6V0A2 antibody. This study was funded by the Fritz Thyssen Stiftung to Uwe Kornak.

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The authors declare no conflict of interest.

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Authors and Affiliations

  1. Institut fuer Medizinische Genetik und Humangenetik, Charité-Universitaetsmedizin Berlin, Berlin, Germany

    Björn Fischer, Aikaterini Dimopoulou, Johannes Egerer, Dominik Jost, Stefan Mundlos & Uwe Kornak

  2. FG Development and Disease, Max-Planck-Institut fuer Molekulare Genetik, Berlin, Germany

    Johannes Egerer, Stefan Mundlos & Uwe Kornak

  3. Department of Pediatrics, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands

    Thatjana Gardeitchik & Eva Morava

  4. Central and Southern Regional Genetic Services, Wellington Hospital, Wellington South, New Zealand

    Alexa Kidd & Emma Felix

  5. Medical Genetics Department, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey

    Hülya Kayserili

  6. Department of Child Health and Diseases, Acibadem University, Istanbul, Turkey

    Yasemin Alanay

  7. Department of Dermatology, University of Cologne, Cologne, Germany

    Iliana Tantcheva-Poor

  8. Institute of Human Genetics, University of Bonn, Bonn, Germany

    Elisabeth Mangold

  9. Prenatal Medicine Munich, Munich, Germany

    Cornelia Daumer-Haas

  10. Department of Medical Genetics, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, India

    Shubha Phadke

  11. Beiersdorf AG, R&D, Skin Research Center, Unnastrasse 48, 20253, Hamburg, Germany

    Reto I. Peirano & Julia Heusel

  12. Department of Clinical Genetics, Guy’s Hospital, London, UK

    Charu Desphande

  13. Department of Pediatrics, All India Institute of Medical Sciences, New Delhi, India

    Neerja Gupta & Madhulika Kabra

  14. Pediatric Dermatology Unit, Asad Al-Hamad Dermatology Center, Al-Sabah Hospital, Kuwait City, Kuwait

    Arti Nanda

  15. Department of Pediatrics, Neurological Sciences and Biochemistry, Rush University Medical Center, Chicago, IL, USA

    Elisabeth Berry-Kravis

  16. Laboratory of Genetic, Endocrine and Metabolic Diseases, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

    Ron A. Wevers

  17. Cutis laxa, Debré type Study Group, Centre de Génétique Humaine, Centre Hospitalier Universitaire, Université de Franche-Comté, 25000, Besançon, France

    Lionel van Maldergem

  18. Berlin-Brandenburg Center for Regenerative Therapies, Charité-Universitaetsmedizin Berlin, Berlin, Germany

    Stefan Mundlos

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  1. Björn Fischer
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  2. Aikaterini Dimopoulou
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Corresponding author

Correspondence to Uwe Kornak.

Additional information

J. Egerer and T. Gardeitchik contributed equally. B. Fischer and A. Dimopoulou contributed equally.

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Supplementary Fig. 1 Control experiments: Target mRNA expression in patient fibroblasts and HeLa cells after RNAi. a ATP6V0A2 mRNA level was determined by quantitative PCR in patient and control dermal fibroblasts. In patient 5 and 2 a significant reduction of ATP6V0A2 mRNA was detectable whereas in patient 10 transcript levels were almost unchanged. b Verification of target gene knockdown efficiency upon 72 h RNAi by quantitative PCR.

Supplementary Fig. 2 Control experiments: Specificity of delayed retrograde trafficking defect after brefeldin A treatment in HeLa cells upon RNAi. The experiment was performed in HeLa cells after 72 h RNA interference. After 10 min of 5 μg/ml brefeldin A treatment, 20 % of control cells and 60 % of ATP6V0A2-depleted cells had incompletely collapsed Golgi structures, respectively. GORAB- as well as PYCR1-depleted cells behaved similar like control cells. All experiments were done at least three times and errors are given as SEM.

Supplementary Fig. 3 ATP6V0A2 and TGFB1 expression level in three day confluent fibroblasts. To investigate whether elevated TGF-β1 levels are due to transcriptional upregulation we determined TGFB1 mRNA levels. As control we measured ATP6V0A2 mRNA and found on average a ~50 % reduction whereas TGFB1 remained unchanged. P-values were determined by t test.

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Fischer, B., Dimopoulou, A., Egerer, J. et al. Further characterization of ATP6V0A2-related autosomal recessive cutis laxa. Hum Genet 131, 1761–1773 (2012). https://doi.org/10.1007/s00439-012-1197-8

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