Chen Xian-Ming, Huang Bing Q, Splinter Patrick L, Cao Hong, Zhu Guan, McNiven Mark A, LaRusso Nicholas F
Center for Basic Research in Digestive Diseases, Division of Gastroenterology and Hepatology, Mayo Medical School, Clinic and Foundation, Rochester, Minnesota 55905, USA.
Gastroenterology. 2003 Jul;125(1):216-28. doi: 10.1016/s0016-5085(03)00662-0.
BACKGROUND & AIMS: Cryptosporidium parvum invasion of epithelia requires polymerization of host cell actin at the attachment site. We analyzed the role of host cell c-Src, a cytoskeleton-associated protein tyrosine kinase, in C. parvum invasion of biliary epithelia.
In vitro models of biliary cryptosporidiosis using a human biliary epithelial cell line were used to assay the role of c-Src signaling pathway in C. parvum invasion.
c-Src and cortactin, an actin-binding protein and a substrate for c-Src, were recruited to the parasite-host cell interface during C. parvum invasion. Tyrosine phosphorylation of cortactin in infected cells was also detected. Inhibition of host cell c-Src significantly blocked C. parvum -induced accumulation and tyrosine phosphorylation of cortactin and actin polymerization at the attachment sites, thereby inhibiting C. parvum invasion of biliary epithelial cells. A triple mutation of tyrosine of cortactin in the epithelia also diminished C. parvum invasion. In addition, proteins originating from the parasite were detected within infected cells at the parasite-host cell interface. Antiserum against C. parvum membrane proteins blocked accumulation of c-Src and cortactin and significantly decreased C. parvum invasion. No accumulation of the endocytosis-related proteins, dynamin 2 and clathrin, was found at the parasite-host cell interface; also, inhibition of dynamin 2 did not block C. parvum invasion.
C. parvum invasion of biliary epithelial cells requires host cell tyrosine phosphorylation of cortactin by a c-Src-mediated signaling pathway to induce actin polymerization at the attachment site, a process associated with microbial secretion but independent of host cell endocytosis.
微小隐孢子虫侵袭上皮细胞需要宿主细胞肌动蛋白在附着位点聚合。我们分析了宿主细胞c-Src（一种与细胞骨架相关的蛋白酪氨酸激酶）在微小隐孢子虫侵袭胆管上皮细胞中的作用。
使用人胆管上皮细胞系建立胆管隐孢子虫病的体外模型，以检测c-Src信号通路在微小隐孢子虫侵袭中的作用。
在微小隐孢子虫侵袭过程中，c-Src和纽蛋白（一种肌动蛋白结合蛋白且为c-Src的底物）被募集到寄生虫-宿主细胞界面。还检测到感染细胞中纽蛋白的酪氨酸磷酸化。抑制宿主细胞c-Src可显著阻断微小隐孢子虫诱导的纽蛋白积累和酪氨酸磷酸化以及附着位点的肌动蛋白聚合，从而抑制微小隐孢子虫对胆管上皮细胞的侵袭。上皮细胞中纽蛋白酪氨酸的三重突变也减少了微小隐孢子虫的侵袭。此外，在寄生虫-宿主细胞界面的感染细胞内检测到源自寄生虫的蛋白质。抗微小隐孢子虫膜蛋白的抗血清可阻断c-Src和纽蛋白的积累，并显著降低微小隐孢子虫的侵袭。在寄生虫-宿主细胞界面未发现内吞作用相关蛋白发动蛋白2和网格蛋白的积累；同样，抑制发动蛋白2也未阻断微小隐孢子虫的侵袭。
微小隐孢子虫侵袭胆管上皮细胞需要宿主细胞通过c-Src介导的信号通路使纽蛋白发生酪氨酸磷酸化，以诱导附着位点的肌动蛋白聚合，这一过程与微生物分泌有关，但独立于宿主细胞内吞作用。
