Department of Cell Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA.
PLoS One. 2011;6(12):e29092. doi: 10.1371/journal.pone.0029092. Epub 2011 Dec 21.
CD36 is a type 2 scavenger receptor with multiple functions. CD36 binding to oxidized LDL triggers signaling cascades that are required for macrophage foam cell formation, but the mechanisms by which CD36 signals remain incompletely understood. Mass spectrometry analysis of anti-CD36 immuno-precipitates from macrophages identified the tetraspanin CD9 as a CD36 interacting protein. Western blot showed that CD9 was precipitated from mouse macrophages by anti-CD36 monoclonal antibody and CD36 was likewise precipitated by anti-CD9, confirming the mass spectrometry results. Macrophages from cd36 null mice were used to demonstrate specificity. Membrane associations of the two proteins on intact cells was analyzed by confocal immunofluorescence microscopy and by a novel cross linking assay that detects proteins in close proximity (<40 nm). Functional significance was determined by assessing lipid accumulation, foam cell formation and JNK activation in wt, cd9 null and cd36 null macrophages exposed to oxLDL. OxLDL uptake, lipid accumulation, foam cell formation, and JNK phosphorylation were partially impaired in cd9 null macrophages. The present study demonstrates that CD9 associates with CD36 on the macrophage surface and may participate in macrophage signaling in response to oxidized LDL.
CD36 是一种具有多种功能的 II 型清道夫受体。CD36 与氧化型 LDL 的结合触发了信号级联反应，这是巨噬细胞泡沫细胞形成所必需的，但 CD36 信号的机制仍不完全清楚。从巨噬细胞中抗 CD36 免疫沉淀物的质谱分析鉴定出四跨膜蛋白 CD9 是 CD36 的相互作用蛋白。Western blot 显示 CD9 可被抗 CD36 单克隆抗体从小鼠巨噬细胞中沉淀出来，而 CD36 也可被抗 CD9 沉淀出来，证实了质谱分析的结果。用 cd36 缺失型小鼠的巨噬细胞来证明特异性。通过共聚焦免疫荧光显微镜和一种新的交联测定法（检测近距离（<40nm）的蛋白质）来分析这两种蛋白质在完整细胞上的膜结合。通过评估 wt、cd9 缺失型和 cd36 缺失型巨噬细胞在暴露于 oxLDL 后脂质积累、泡沫细胞形成和 JNK 激活来确定功能意义。oxLDL 摄取、脂质积累、泡沫细胞形成和 JNK 磷酸化在 cd9 缺失型巨噬细胞中部分受损。本研究表明 CD9 与巨噬细胞表面的 CD36 相关联，并可能参与氧化型 LDL 刺激的巨噬细胞信号转导。
