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Molecular Dissection of the Organization and the Dynamics of Intracellular Lipid Domains

组织的分子解剖和细胞内脂质结构域的动力学

基本信息

批准号：
12672143
负责人：
KOBAYASHI Toshihide
金额：
$ 2.24万
依托单位：
RIKEN
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

项目摘要

I. Involvement of mitochondrial cardiolipin domains in apoptosis.Cytochrome c is a proapoptotic factor that binds preferentially to cardiolipin (CL), a mitochondrial lipid, but not to cardiolipin hydroperoxide (Cl-OOH). Our studies using overexpression of mitochondrial phospholipid hydroperoxide glutathione peroxidase (PHGPx) suggests that the generation of CL-OOH in mitochondria might be a primary event that triggers the release of cytochrome c.II. Characterization of lysobisphosphatidic acid domain in late endosomes.Late endosomes accumulates internal membranes within the lumen of the organelle. These intemal membranes are enriched in the specific lipid, Iysobisphosphatidic acid (LBPA). Using LBPA-specific monoclonal antibody and fluorescence correlation spectroscopy, we have determined that LBPA domain is exclusively localized in the lumen of late endosomes.III. Caveolae and glycosylphosphatidylinositol domain.Most mammalian cells have in their membrane at least two types of lipid m … More icrodomains, non-invaginated lipid rafts and caveolae. Glycosylphosphatidylinositol (GPI)-anchored proteins constitute a class of proteins that are enriched in rafts but not caveolae at steady state. By using GPI-deficient cells and the cells overexpressing caveolin-1, we showed there is an inverse correlation between the expression of GPI-anchored proteins and caveolin-1.IV. Sphingomyelin-specific proteinLysenin is a novel 4lkDa sphingomyelin-binding protein obtained from coelomic fluid of the earthworrn Eisenia foetida. We have developed recombinant lysenin and examined the distribution of sphingomyelin under electron microscope. Sphingomyelin was accumulated in caveolae in endothelial cells. Sphingomyelin was also distributed in endosomes and the trans-Golgi network but excluded from the Golgi cistemae. We also examined sphingomyelin in Niemann-Pick type A (NPA) fibroblpsts, which accumulate sphingomyelin intracellularly Sphingomyelin is accumuiated in late endosomes lysosomes in NPA. Interestingly, other lipid raft components such as ganglioside GM1, and cholesterol were aiso accumulated in the same organelle. Our results suggest that re-distribution of lipid raft occurs in NPA. Less

线粒体心磷脂结构域参与细胞凋亡。细胞色素c是一种促凋亡因子，优先结合心肌磷脂（CL），一种线粒体脂质，但不结合过氧化心肌磷脂（CL - ooh）。我们利用线粒体磷脂氢过氧化物谷胱甘肽过氧化物酶（PHGPx）过表达的研究表明，线粒体中CL-OOH的产生可能是触发细胞色素c.II释放的主要事件。内体晚期溶二磷脂酸结构域的表征。晚期核内体在细胞器腔内的内膜内积聚。这些细胞膜富含一种特殊的脂质，即双磷脂酸（LBPA）。利用LBPA特异性单克隆抗体和荧光相关光谱，我们已经确定LBPA结构域完全定位于晚期内核体的腔内。囊泡和糖基磷脂酰肌醇结构域。大多数哺乳动物细胞的膜中至少有两种类型的脂质结构域，更多的是微结构域，非内陷脂筏和小泡。糖基磷脂酰肌醇（GPI）锚定蛋白是一类在筏状结构中富集而在稳定状态下不富集的蛋白。通过gpi缺陷细胞和过表达caveolin-1的细胞，我们发现gpi锚定蛋白的表达与caveolin-1 - iv呈负相关。鞘磷脂特异性蛋白lysenin是从蚯蚓叶eisenia foetida体腔液中获得的一种新型鞘磷脂结合蛋白。我们研制了重组溶菌素，并在电镜下观察了鞘磷脂的分布。鞘磷脂在内皮细胞的小泡中积累。鞘磷脂也分布在核内体和反式高尔基网络中，但不存在于高尔基池中。我们还检测了neemann - pick型A （NPA）纤维母细胞中的鞘磷脂，它们在细胞内积累鞘磷脂。鞘磷脂在NPA的内核溶酶体中积累。有趣的是，其他脂质筏成分如神经节苷脂GM1和胆固醇也在同一细胞器中积累。我们的结果表明，脂筏的重新分布发生在NPA中。少