Apical organelle discharge by Cryptosporidium parvum is temperature, cytoskeleton, and intracellular calcium dependent and required for host cell invasion
Apical organelle discharge by Cryptosporidium parvum is temperature, cytoskeleton, and intracellular calcium dependent and required for host cell invasion
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DOI：
10.1128/iai.72.12.6806-6816.2004
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发表时间：
2004-12-01
影响因子：
3.1
通讯作者：
LaRusso, NF
中科院分区：
文献类型：
--
作者：
Chen, XM;O'Hara, SP;LaRusso, NF
顶复合体寄生虫的顶细胞器是含有复杂分子混合物的特征性分泌囊泡。虽然已证实一些顶复合体寄生虫（包括弓形虫和疟原虫）的顶细胞器放电参与了细胞侵袭，但细小隐孢子虫孢子子的顶细胞器放电机制及其在侵染宿主细胞中的作用尚不清楚。在这里，我们证明了小孢子虫顶端细胞器的放电是以温度依赖的方式发生的。细胞松弛素D和秋水仙碱分别抑制了寄生虫的肌动蛋白和微管蛋白聚合，抑制了寄生虫的顶端细胞器放电。寄生虫细胞内钙离子的螯合作用也抑制了顶端细胞器的放电，这一过程可以通过使用离子载体A23187提高细胞内钙离子浓度而部分逆转。细胞松弛素D和秋水仙碱对寄生虫细胞骨架聚合的抑制以及细胞内钙的耗竭也降低了小孢子虫孢子体的滑行能力。重要的是，细小梭菌孢子体对顶端细胞器放电的抑制作用阻断了寄生虫对宿主细胞（即培养的人胆管细胞）的侵袭，而不是附着。此外，在宿主-寄生虫界面区域检测到寄生虫蛋白CP2向宿主细胞膜的易位；CP2抗体可降低小弓形虫对胆管细胞的侵袭。这些数据表明，小孢子虫孢子体顶端细胞器内容物的排出是发生的，并且它依赖于温度、细胞内钙和细胞骨架，并且是入侵宿主细胞所必需的，证实了顶端细胞器在小孢子虫进入宿主细胞中起着核心作用。
The apical organelles in apicomplexan parasites are characteristic secretory vesicles containing complex mixtures of molecules. While apical organelle discharge has been demonstrated to be involved in the cellular invasion of some apicomplexan parasites, including Toxoplasma gondii and Plasmodium spp., the mechanisms of apical organelle discharge by Cryptosporidium parvum sporozoites and its role in host cell invasion are unclear. Here we show that the discharge of C. parvum apical organelles occurs in a temperature-dependent fashion. The inhibition of parasite actin and tubulin polymerization by cytochalasin D and colchicines, respectively, inhibited parasite apical organelle discharge. Chelation of the parasite's intracellular calcium also inhibited apical organelle discharge, and this process was partially reversed by raising the intracellular calcium concentration by use of the ionophore A23187. The inhibition of parasite cytoskeleton polymerization by cytochalasin D and colchicine and the depletion of intracellular calcium also decreased the gliding motility of C. parvum sporozoites. Importantly, the inhibition of apical organelle discharge by C. parvum sporozoites blocked parasite invasion of, but not attachment to, host cells (i.e., cultured human cholangiocytes). Moreover, the translocation of a parasite protein, CP2, to the host cell membrane at the region of the host cell-parasite interface was detected; an antibody to CP2 decreased the C. parvum invasion of cholangiocytes. These data demonstrate that the discharge of C. parvum sporozoite apical organelle contents occurs and that it is temperature, intracellular calcium, and cytoskeleton dependent and required for host cell invasion, confirming that apical organelles play a central role in C. parvum entry into host cells.