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.

中文翻译：

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四种 ATP 结合盒转运蛋白 A3 基因 (ABCA3) 变体的功能表征。

ABCA3 在 II 型肺泡细胞中跨板层体膜转运磷脂，并且是表面活性剂组装所必需的。罕见的双等位基因致病性ABCA3变异会导致致命的新生儿呼吸窘迫综合征和儿童间质性肺病。ABCA3错义变体的定性功能表征表明有两种致病类别：细胞内运输中断（I 型突变体）或 ATP 酶介导的磷脂转运至层状体受损（II 型突变体）。我们定性比较了野生型 (WT-AB​​CA3) 与四种未表征的ABCA3变体 (c.418A>C;p.Asn140His、c.3609_3611delCTT;p.Phe1203del、c.3784A>G;p.Ser1262Gly 和 c.4195G> A;p.Val1399Met) 在 A549 细胞中使用蛋白质加工、与细胞内细胞器的共定位、层状体超微结构和 ATP 酶活性。我们使用基于荧光的共定位定量测量了层状体样囊泡直径和细胞内 ABCA3 运输。三个ABCA3变体（p.Asn140His、p.Ser1262Gly 和 p.Val1399Met）正常加工和运输，并表现出组织良好的层状体样囊泡，但与 II 型突变体一致，ATP 酶活性降低。P.Phe1203del 加工正常，ATP 酶活性降低，具有组织良好的层状体样囊泡，但与内质网和溶酶体标记物定量共定位，这是一种中间表型，表明细胞内运输和磷脂运输均受到破坏。所有 ABCA3 突变体的平均囊泡直径均小于 WT-AB​​CA3。ABCA3变体的定性和定量功能表征可揭示致病机制。