Functional characterization of four ATP-binding cassette transporter A3 gene (ABCA3) variants

June Y. Hu, Ping Yang, Daniel J. Wegner, Hillary B. Heins, Cliff J. Luke, Fuhai Li, Frances V. White, Gary A. Silverman, F. Sessions Cole, Jennifer A. Wambach

Research output: Contribution to journalArticle

Abstract

ABCA3 transports phospholipids across lamellar body membranes in pulmonary alveolar type II cells and is required for surfactant assembly. Rare, biallelic, pathogenic ABCA3 variants result in lethal neonatal respiratory distress syndrome and childhood interstitial lung disease. Qualitative functional characterization of ABCA3 missense variants suggests two pathogenic classes: disrupted intracellular trafficking (type I mutant) or impaired ATPase-mediated phospholipid transport into the lamellar bodies (type II mutant). We qualitatively compared wild-type (WT-ABCA3) with four uncharacterized ABCA3 variants (c.418A>C;p.Asn140His, c.3609_3611delCTT;p.Phe1203del, c.3784A>G;p.Ser1262Gly, and c.4195G>A;p.Val1399Met) in A549 cells using protein processing, colocalization with intracellular organelles, lamellar body ultrastructure, and ATPase activity. We quantitatively measured lamellar body-like vesicle diameter and intracellular ABCA3 trafficking using fluorescence-based colocalization. Three ABCA3 variants (p.Asn140His, p.Ser1262Gly, and p.Val1399Met) were processed and trafficked normally and demonstrated well-organized lamellar body-like vesicles, but had reduced ATPase activity consistent with type II mutants. P.Phe1203del was processed normally, had reduced ATPase activity, and well-organized lamellar body-like vesicles, but quantitatively colocalized with both endoplasmic reticulum and lysosomal markers, an intermediate phenotype suggesting disruption of both intracellular trafficking and phospholipid transport. All ABCA3 mutants demonstrated mean vesicle diameters smaller than WT-ABCA3. Qualitative and quantitative functional characterization of ABCA3 variants informs mechanisms of pathogenicity.

Original languageEnglish (US)
Pages (from-to)1298-1307
Number of pages10
JournalHuman mutation
Volume41
Issue number7
DOIs
StatePublished - Jul 1 2020

Keywords

  • ABCA3
  • childhood interstitial lung disease (chILD)
  • lamellar bodies
  • respiratory distress syndrome (RDS)
  • surfactant

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

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    Hu, J. Y., Yang, P., Wegner, D. J., Heins, H. B., Luke, C. J., Li, F., White, F. V., Silverman, G. A., Sessions Cole, F., & Wambach, J. A. (2020). Functional characterization of four ATP-binding cassette transporter A3 gene (ABCA3) variants. Human mutation, 41(7), 1298-1307. https://doi.org/10.1002/humu.24014