Phosphatidylinositol 3-phosphate is generated in phagosomal membranes
Phosphatidylinositol 3-phosphate is generated in phagosomal membranes
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DOI：
10.1016/s0960-9822(01)00447-x
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发表时间：
2001-10-16
期刊：
影响因子：
9.2
通讯作者：
Hawkins, PT
中科院分区：
文献类型：
--
作者：
Ellson, CD;Anderson, KE;Hawkins, PT
吞噬细胞，如中性粒细胞和巨噬细胞，吞噬并摧毁入侵的微生物。在内化后，吞噬体膜内捕获的物质被消化酶和活性氧物种协调输送的复杂过程破坏。最近，几种预期参与这些活动的内切酶、溶酶体和氧化酶成分被证明与PtdIns3P结合，这表明这种脂类可能在这一过程中发挥作用[1-5]。我们使用稳定表达PtdIns3P结合GFP-PX结构域或GFP-FYVE结构域的RAW 264.7细胞的实时数字荧光成像来观察吞噬Ig G调理酵母多聚糖颗粒过程中PtdIns3P水平和亚细胞定位的变化。两个PtdIns3P探针的结果非常相似，每个PtdIns3P探针的基本分布部分是胞浆的，部分定位于EEA-1阳性的内体结构。在酵母多糖附着并伴随吞噬体膜关闭的大约2-3分钟内，GFP阳性囊泡向吞噬小体的局部区域移动并附着。随之而来的是GFP探针在整个吞噬小体周围戏剧性的、瞬时的积累，伴随着胞浆GFP荧光的瞬时下降。PtdIns3P升高的幅度和时间清楚地表明，它是控制吞噬小体成熟的早期阶段的理想候选者。
Phagocytic cells such as neutrophils and macrophages engulf and destroy invading microorganisms. After internalization, material captured within the phagosomal membrane is destroyed by a complex process of coordinated delivery of digestive enzymes and reactive oxygen species. Several endosomal, lysosomal, and oxidase components expected to participate in these events have recently been shown to bind PtdIns3P, suggesting that this lipid may play a role in this process [1-5]. We used live, digital fluorescence imaging of RAW 264.7 cells stably expressing either a PtdIns3P binding GFP-PX domain or a GFP-FYVE domain to visualize changes in the levels and subcellular localization of PtdIns3P during phagocytic uptake of IgG-opsonized zymosan particles. Very similar results were obtained using both PtdIns3P probes, The basal distribution of each PtdIns3P probe was partially cytosolic and partially localized to EEA-1-positive endosomal structures. Within about 2-3 min of zymosan attachment and concomitant with the closure of the phagosomal membrane, GFP-positive vesicles moved toward and attached to a localized area of the phagosome. A dramatic, transient accumulation of GFP probe around the entire phagosome rapidly ensued, accompanied by a transient drop in cytosolic GFP fluorescence. The magnitude and timing of this rise in PtdIns3P clearly suggest that it is an ideal candidate for controlling the early stages of phagosomal maturation.