Homooligomerization of ABCA3 and its functional significance

Frixel,Sabrina; Lotz-Havla,Amelie  S.; Kern,Sunčana; Kaltenborn,Eva; Wittmann,Thomas; Gersting,Søren  W.; Muntau,Ania  C.; Zarbock,Ralf; Griese,Matthias

doi:10.3892/ijmm.2016.2650

Homooligomerization of ABCA3 and its functional significance

Authors:
- Sabrina Frixel
- Amelie S. Lotz-Havla
- Sunčana Kern
- Eva Kaltenborn
- Thomas Wittmann
- Søren W. Gersting
- Ania C. Muntau
- Ralf Zarbock
- Matthias Griese
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Affiliations: German Centre for Lung Research, Dr von Hauner Children's Hospital, Ludwig-Maximilians University, D-80337 Munich, Germany, Department of Molecular Pediatrics, Dr von Hauner Children's Hospital, Ludwig-Maximilians University, D-80337 Munich, Germany, University Children's Hospital, University Medical Center Hamburg-Eppendorf, D-20246 Hamburg, Germany
Published online on: June 21, 2016 https://doi.org/10.3892/ijmm.2016.2650
Pages: 558-566

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Abstract

ABCA3 is a surfactant lipid transporter in the limiting membrane of lamellar bodies in alveolar type II cells. Mutations in the ATP-binding cassette, sub-family A (ABC1), member 3 (ABCA3) gene cause respiratory distress syndrome in newborns, and chronic interstitial lung disease in children and adults. ABCA3 belongs to the class of full ABC transporters, which are supposed to be functional in their monomeric forms. Although other family members e.g., ABCA1 and ABCC7 have been shown to function as oligomers, the oligomerization state of ABCA3 is unknown. In the present study, the oligomerization of ABCA3 was investigated in cell lysates and crude membrane preparations from transiently and stably transfected 293 cells using blue native PAGE (BN-PAGE), gel filtration and co-immunoprecipitation. Additionally, homooligomerization was examined in vivo in cells using bioluminescence resonance energy transfer (BRET). Using BN-PAGE and gel filtration, we demonstrate that non-denatured ABCA3 exists in different oligomeric forms, with monomers (45%) and tetramers (30%) being the most abundant forms. Furthermore, we also show the existence of 20% dimers and 5% trimers. BRET analyses verified intermolecular interactions in vivo. Our results also demonstrated that the arrest of ABCA3 in the endoplasmic reticulum (ER), either through drug treatment or induced by mutations in ABCA3, inhibited the propensity of the protein to form dimers. Based on our results, we suggest that transporter oligomerization is crucial for ABCA3 function and that a disruption of oligomerization due to mutations represents a novel pathomechanism in ABCA3-associated lung disease.

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