Molecular dissection of organization and dynamics of lipid domains in biomembranes

生物膜中脂质结构域的组织和动力学的分子解剖

基本信息

批准号：
17390025
负责人：
KOBAYASHI Toshihide
金额：
$ 9.6万
依托单位：
RIKEN
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

(1) Characterization of lipid probesLysenin is a pore-forming toxin that specifically binds sphingomyelin (SM). The binding of the toxin to the membrane is accompanied by the oligomerization of the protein, leading to pore formation. Although a previous study showed that SM/cholesterol liposomes were 10,000 times more effective than SM liposomes in inhibiting lysenin-induced hemolysis, the role of cholesterol is not precisely clarified. Our results indicate that both cholesterol and the SM/lysenin ratio control the amount of lysenin monomer via altering the state of protein oligomerization, thus affecting hemolysis.Duramycin is a 19 amino acid tetracyclic lantibiotic closely related to cinnamycin, which is known to bind phosphatidyl-ethanolamine (PE). The lipid specificity of duramycin was not established. Our study indicates that both duramycin and cinnamycin exclusively bind to ethanolamine phospholipids in curvature-dependent manner.(2) Regulation of endocytosis by cholesterolCellular cholesterol increases when cells reach confluency in Chinese hamster ovary (CHO) cells. We showed that the endocytic pathways of several fluorescent lipids were altered in cell confluency-dependent manner. The crucial role of cellular cholesterol in cell-confluency dependent endocytosis was suggested. Our results also suggest that cholesterol controls endocytic routes of a subset of membrane lipids through rab11.(3) Regulation of cellular cholesterol homeostasis by endosome lipid domainsUsing D-PDMP and ganglioside GM1, it is shown that the structure alteration of bis (monoacylglycero) phosphate (BMP)-rich late endosome lipid domain plays a crucial role in cellular cholesterol accumulation. We also employed anti-BMP monoclonal antibody to examine the role of BMP domain in cholesterol homeostasis in macrophages.

(1)脂质探针的表征溶菌素是一种特异性结合鞘磷脂(SM)的成孔毒素。毒素与膜的结合伴随着蛋白质的寡聚化，导致孔的形成。尽管之前的研究表明，SM/胆固醇脂质体在抑制赖氨酸引起的溶血方面比SM脂质体有效10,000倍，但胆固醇的作用尚未明确阐明。我们的结果表明，胆固醇和 SM/lysenin 比例通过改变蛋白质寡聚状态来控制 lysenin 单体的量，从而影响溶血。耐久霉素是一种与肉桂霉素密切相关的 19 个氨基酸的四环羊毛硫抗生素，已知肉桂霉素可结合磷脂酰-乙醇胺（PE）。耐久霉素的脂质特异性尚未确定。我们的研究表明，耐久霉素和肉桂霉素都以曲率依赖性方式专门与乙醇胺磷脂结合。(2)胆固醇对内吞作用的调节当细胞在中国仓鼠卵巢(CHO)细胞中达到汇合时，细胞胆固醇增加。我们发现几种荧光脂质的内吞途径以细胞汇合依赖性方式发生改变。提出了细胞胆固醇在细胞汇合依赖性内吞作用中的关键作用。我们的结果还表明，胆固醇通过 rab11 控制膜脂子集的内吞途径。 (3) 内体脂质结构域调节细胞胆固醇稳态使用 D-PDMP 和神经节苷脂 GM1，表明双（单酰基甘油）磷酸酯的结构改变富含 BMP 的晚期内体脂质结构域在细胞胆固醇积累中起着至关重要的作用。我们还使用抗 BMP 单克隆抗体来检查 BMP 结构域在巨噬细胞胆固醇稳态中的作用。