Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knöll Institute (Leibniz-HKI), Jena, Germany.
Department of Microbiology and Molecular Biology, Institute of Microbiology, Friedrich Schiller University Jena, Jena, Germany.
mSphere. 2023 Feb 21;8(1):e0052322. doi: 10.1128/msphere.00523-22. Epub 2023 Jan 31.
Alveolar macrophages belong to the first line of defense against inhaled conidia of the human-pathogenic fungus Aspergillus fumigatus. In lung alveoli, they contribute to phagocytosis and elimination of conidia. As a counterdefense, conidia have a gray-green pigment that enables them to survive in phagosomes of macrophages for some time. Previously, we showed that this conidial pigment interferes with the formation of flotillin-dependent lipid raft microdomains in the phagosomal membrane, thereby preventing the formation of functional phagolysosomes. Besides flotillins, stomatin is a major component of lipid rafts and can be targeted to the membrane. However, only limited information on stomatin is available, in particular on its role in defense against pathogens. To determine the function of this integral membrane protein, a stomatin-deficient macrophage line was generated by CRISPR/Cas9 gene editing. Immunofluorescence microscopy and flow cytometry revealed that stomatin contributes to the phagocytosis of conidia and is important for recruitment of the β-glucan receptor dectin-1 to both the cytoplasmic membrane and phagosomal membrane. In stomatin knockout cells, fusion of phagosomes and lysosomes, recruitment of the vATPase to phagosomes, and tumor necrosis factor alpha (TNF-α) levels were reduced when cells were infected with pigmentless conidia. Thus, our data suggest that stomatin is involved in maturation of phagosomes via fostering fusion of phagosomes with lysosomes. Stomatin is an integral membrane protein that contributes to the uptake of microbes, e.g., spores of the human-pathogenic fungus Aspergillus fumigatus. By generation of a stomatin-deficient macrophage line by advanced genetic engineering, we found that stomatin is involved in the recruitment of the β-glucan receptor dectin-1 to the phagosomal membrane of macrophages. Furthermore, stomatin is involved in maturation of phagosomes via fostering fusion of phagosomes with lysosomes. The data provide new insights on the important role of stomatin in the immune response against human-pathogenic fungi.
肺泡巨噬细胞属于抵御人类致病真菌烟曲霉吸入分生孢子的第一道防线。在肺肺泡中，它们有助于吞噬和消除分生孢子。作为一种反击，分生孢子有一种灰绿色的色素，使它们能够在巨噬细胞的吞噬体中存活一段时间。以前，我们表明这种分生孢子色素会干扰吞噬体膜中依赖于 flotillin 的脂筏微域的形成，从而阻止功能性吞噬溶酶体的形成。除了 flotillin 外，stomatin 也是脂筏的主要组成部分，并且可以靶向到膜上。然而，关于 stomatin 的信息有限，特别是关于其在防御病原体方面的作用。为了确定这种整合膜蛋白的功能，通过 CRISPR/Cas9 基因编辑生成了 stomatin 缺陷型巨噬细胞系。免疫荧光显微镜和流式细胞术显示，stomatin 有助于分生孢子的吞噬作用，并且对于 β-葡聚糖受体 dectin-1 向细胞质膜和吞噬体膜的募集都很重要。在 stomatin 敲除细胞中，当细胞被无色分生孢子感染时，吞噬体和溶酶体的融合、vATPase 向吞噬体的募集以及肿瘤坏死因子-α（TNF-α）水平降低。因此，我们的数据表明 stomatin 通过促进吞噬体与溶酶体的融合参与吞噬体的成熟。stomatin 是一种整合膜蛋白，有助于微生物的摄取，例如人类致病真菌烟曲霉的孢子。通过先进的基因工程生成 stomatin 缺陷型巨噬细胞系，我们发现 stomatin 参与了 β-葡聚糖受体 dectin-1 向巨噬细胞吞噬体膜的募集。此外，stomatin 通过促进吞噬体与溶酶体的融合参与吞噬体的成熟。这些数据为 stomatin 在针对人类致病真菌的免疫反应中的重要作用提供了新的见解。
