Balce Dale R, Rybicka Joanna M, Greene Catherine J, Ewanchuk Benjamin W, Yates Robin M
Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, Calgary, Canada.
Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Calgary, Calgary, Canada.
Traffic. 2016 Jul;17(7):786-802. doi: 10.1111/tra.12396. Epub 2016 May 13.
Proteolysis and the reduction of disulfides, both major components of protein degradation, are profoundly influenced by phagosomal redox conditions in macrophages. We evaluated the activation of phagocytic receptors that are known to influence activation of the phagocyte NADPH oxidase (NOX2), and its effect on phagosomal protein degradation. Population-based and single phagosome analyses of phagosomal chemistries in murine macrophages revealed that activation of NOX2 via the Fcγ receptor (FcγR) during phagocytosis decreased rates of proteolysis and disulfide reduction. Immunoglobulin G (IgG)-stimulated reactive oxygen species (ROS) production and the inhibition of phagosomal proteolysis and disulfide reduction were dependent on NOX2, FcγR and protein kinase C (PKC)/spleen tyrosine kinase (Syk) signaling. In contrast, low levels of ROS production were observed following the phagocytosis of unopsonized beads, which resulted in higher rates of phagosomal proteolysis and disulfide reduction. Phagosomes displayed autonomy with respect to FcγR-mediated differences in NOX2 activation and proteolysis, as phagosomes containing unopsonized cargo retained low NOX2 activation and high proteolysis even in the presence of phagosomes containing IgG-opsonized cargo in the same macrophage. These results show that opsonization of phagocytic cargo results in vastly different phagosomal processing of proteins through the FcγR-triggered, PKC/Syk-dependent local assembly and activation of NOX2.
蛋白水解作用以及二硫键的还原，这两个蛋白质降解的主要组成部分，都受到巨噬细胞吞噬体氧化还原状态的深刻影响。我们评估了已知会影响吞噬细胞NADPH氧化酶（NOX2）激活的吞噬受体的激活情况，及其对吞噬体蛋白降解的影响。对小鼠巨噬细胞吞噬体化学性质进行的群体分析和单个吞噬体分析表明，吞噬过程中通过Fcγ受体（FcγR）激活NOX2会降低蛋白水解速率和二硫键还原速率。免疫球蛋白G（IgG）刺激产生的活性氧（ROS）以及对吞噬体蛋白水解作用和二硫键还原的抑制依赖于NOX2、FcγR和蛋白激酶C（PKC）/脾酪氨酸激酶（Syk）信号传导。相比之下，吞噬未调理珠后观察到低水平的ROS产生，这导致吞噬体蛋白水解和二硫键还原的速率更高。吞噬体在FcγR介导的NOX2激活和蛋白水解差异方面表现出自主性，因为即使在同一巨噬细胞中存在含有IgG调理货物的吞噬体，含有未调理货物的吞噬体仍保持低水平的NOX2激活和高蛋白水解作用。这些结果表明，吞噬货物的调理作用通过FcγR触发的、PKC/Syk依赖的NOX2局部组装和激活，导致蛋白质在吞噬体中的加工过程大不相同。
