Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, USA.
N Engl J Med. 2013 Jul 4;369(1):54-65. doi: 10.1056/NEJMoa1301296. Epub 2013 Jun 5.
Neutrophils are the predominant phagocytes that provide protection against bacterial and fungal infections. Genetically determined neutrophil disorders confer a predisposition to severe infections and reveal novel mechanisms that control vesicular trafficking, hematopoiesis, and innate immunity.
We clinically evaluated seven children from five families who had neutropenia, neutrophil dysfunction, bone marrow fibrosis, and nephromegaly. To identify the causative gene, we performed homozygosity mapping using single-nucleotide polymorphism arrays, whole-exome sequencing, immunoblotting, immunofluorescence, electron microscopy, a real-time quantitative polymerase-chain-reaction assay, immunohistochemistry, flow cytometry, fibroblast motility assays, measurements of apoptosis, and zebrafish models. Correction experiments were performed by transfecting mutant fibroblasts with the nonmutated gene.
All seven affected children had homozygous mutations (Thr224Asn or Glu238Lys, depending on the child's ethnic origin) in VPS45, which encodes a protein that regulates membrane trafficking through the endosomal system. The level of VPS45 protein was reduced, as were the VPS45 binding partners rabenosyn-5 and syntaxin-16. The level of β1 integrin was reduced on the surface of VPS45-deficient neutrophils and fibroblasts. VPS45-deficient fibroblasts were characterized by impaired motility and increased apoptosis. A zebrafish model of vps45 deficiency showed a marked paucity of myeloperoxidase-positive cells (i.e., neutrophils). Transfection of patient cells with nonmutated VPS45 corrected the migration defect and decreased apoptosis.
Defective endosomal intracellular protein trafficking due to biallelic mutations in VPS45 underlies a new immunodeficiency syndrome involving impaired neutrophil function. (Funded by the National Human Genome Research Institute and others.).
中性粒细胞是主要的吞噬细胞，可提供针对细菌和真菌感染的保护。遗传决定的中性粒细胞疾病赋予了对严重感染的易感性，并揭示了控制囊泡运输、造血和先天免疫的新机制。
我们对来自五个家庭的七名患有中性粒细胞减少症、中性粒细胞功能障碍、骨髓纤维化和肾肿大的儿童进行了临床评估。为了确定致病基因，我们使用单核苷酸多态性阵列、全外显子组测序、免疫印迹、免疫荧光、电子显微镜、实时定量聚合酶链反应检测、免疫组织化学、流式细胞术、成纤维细胞迁移检测、凋亡测量和斑马鱼模型进行了纯合子作图。通过将突变基因转染到突变的成纤维细胞中进行纠正实验。
所有七名受影响的儿童均携带 VPS45 基因的纯合突变（根据儿童的种族起源，分别为 Thr224Asn 或 Glu238Lys），该基因编码一种调节通过内体系统的膜运输的蛋白质。VPS45 蛋白水平降低，与其结合的 rabenosyn-5 和 syntaxin-16 减少。VPS45 缺陷中性粒细胞和成纤维细胞表面的 β1 整合素水平降低。VPS45 缺陷成纤维细胞的特征是迁移能力受损和凋亡增加。vps45 缺陷斑马鱼模型显示髓过氧化物酶阳性细胞（即中性粒细胞）明显减少。用非突变 VPS45 转染患者细胞可纠正迁移缺陷并减少凋亡。
由于 VPS45 的双等位基因突变导致内体细胞内蛋白运输缺陷，导致一种新的免疫缺陷综合征，涉及中性粒细胞功能障碍。（由国家人类基因组研究所等资助）。
