Analysis of phosphatidylinositol mediated molecular mechanisms that related to phagocytosis process of the Entamoeba histolytica.

磷脂酰肌醇介导的溶组织内阿米巴吞噬过程相关分子机制分析。

基本信息

批准号：
20790323
负责人：
TSUKUI Kumiko
金额：
$ 2.75万
依托单位：
National Institute of Infectious Diseases
依托单位国家：
日本
项目类别：
Grant-in-Aid for Young Scientists (B)
财政年份：
2008
资助国家：
日本
起止时间：
2008 至 2009
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-20790323/
关键词：
原虫感染症イノシトールリン脂質小胞輸送赤痢アメーバ貪食 RhoGEF Rac PtdIns4P

项目摘要

To understand the roles of phosphoinositides [PtdIns] in phagocytosis of parasitic eukaryotes, we examined the interaction of phosphatidylinositol-3-phosphate [PtdIns(3)P] and putative PtdIns-P-binding proteins during phagocytosis in the enteric protozoan parasite Entamoeba histolytica. It was previously shown that phagocytosis in E. histolytica is indispensable for virulence and is inhibited by PtdIns 3-kinase inhibitors. We demonstrated by time-lapse live imaging that during the initiation of phagocytosis, the PtdIns(3)P biomarker GFP-Hrs-FYVE, was translocated to the phagocytic cup, phagosome, and to tunnel-like structures connecting the plasma membrane and phagosomes. E. histolytica possesses 12 FYVE domain-containing proteins (EhFP1-12), 11 of which also contain the RhoGEF/DH domain. Among them EhFP4 was shown to be recruited to the tunnel-like structures and to the proximal region of the phagosome. We further demonstrated that EhFP4 physically interacted with 4 of 10 predominant Rho/Rac small GTPases. Phosphoinositide binding assay showed that EhFP4 unexpectedly bound to PtdIns(4)P via the carboxyl-terminal domain and that the FYVE domain modulates the binding specificity of EhFP4 to PtdIns-P. Expression of the FYVE domain from EhFP4 inhibited phagocytosis while enhancement was observed when mammalian Hrs-FYVE domain was expressed. Altogether, we demonstrated that PtdIns(3)P, PtdIns(4)P and EhFP4 coordinately regulate phagocytosis and phagosome maturation in this parasitic eukaryote.

为了了解磷酸肌醇[ptdins]在寄生虫吞噬的吞噬中的作用，我们检查了磷脂酰肌醇-3-磷酸[PTDINS（3）P]（3）P]和PTDINS-P结合蛋白的相互作用在吞噬proteric protozoan protaSite paraSata paraSASA paraSASA paraSANITA paraSapa中。先前的表明，溶血性大肠杆菌中的吞噬作用对于毒力是必不可少的，受PTDINS 3-激酶抑制剂的抑制。我们通过延时现场成像证明，在吞噬作用的启动过程中，PTDINS（3）P生物标志物GFP-HRS-FYVE被转移到吞噬杯，吞噬体和连接质膜膜和phagosomes的隧道样结构。 E. histolictica具有12个含FYVE结构域的蛋白质（EHFP1-12），其中11个还包含Rhogef/DH结构域。其中显示了EHFP4被招募到隧道样结构和吞噬体的近端区域。我们进一步证明了EHFP4与10个主要的Rho/RAC小型GTPase中的4个物理相互作用。磷酸肌醇结合测定法显示，EHFP4通过羧基末端结构域意外地与PTDINS（4）P结合，而FYVE结构域调节EHFP4与Ptdins-P的结合特异性。在表达哺乳动物HRS-Fyve域时，观察到FYVE结构域的表达在EHFP4中抑制了吞噬作用。总的来说，我们证明了PTDINS（3）P，PTDINS（4）P和EHFP4在此寄生真核生物中协调调节吞噬作用和吞噬体成熟。