MEMBRANE TRAFFIC IN SPHINGOLIPID STORAGE DISEASES

鞘脂储存疾病中的膜运输

• 批准号: 6625111
• 负责人: RICHARD E PAGANO
• 金额: $ 23.64万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-12-22 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6625111
• 关键词: CHO cells; Golgi apparatus; N acylation; Niemann Pick disease; cell component structure /function; cell line; cholesterol; cholesterol ester storage disease; endocytosis; fibroblasts; fluorescent dye /probe; gene mutation; guanosinetriphosphatases; immunofluorescence technique; intracellular transport; lipid metabolism; lipid transport; lysosomes; membrane activity; membrane lipids; membrane structure; membrane transport proteins; molecular pathology; protein localization; sphingolipidosis

DESCRIPTION (Verbatim from the applicant's abstract): The investigator will study membrane lipid transport and sphingolipid/cholesterol interactions in sphingolipid storage disease (SLSD) fibroblasts. Recently he found that several fluorescent SL analogs were internalized from the plasma membrane (PM) predominantly to the Golgi complex of normal cells, while in 10 different SLSD cell types, these lipids accumulated in endosomes and lysosomes. This accumulation was in general unrelated to the extent of SL analog degradation in the different cell types. He also showed that cholesterol bomeostasis is perturbed in multiple SLSDs secondary to SL accumulation and that mistargeting of SL analogs was regulated by cholesterol. Based on these results, they hypothesize that endogenous lipids which accumulate in SLSD cells due to primary defects in lipid catabolism result in an altered intracellular distribution of cholesterol, and that this alteration in membrane composition then results in defective sorting and transport of SLs. Four broad projects pertaining to this hypothesis will.be pursued. They will (i) examine the itinerary of fluorescent internalized recycled from the PM to various intracellular compartments over time: Dominant negative constructs of several different Rab proteins will be used to help define the compartment(s) where defective SL sorting occurs; (ii) study the intracellular transport of endogenous (e.g., using fluorescent SL binding toxins, anti-SL antibodies, or resialylation of SLs) to confirm their preliminary observations that the perturbation in PM to Golgi traffic seen in SLSD fibroblasts with fluorescent SLs is mirrored by endogenous SLs; (iii) evaluate potential mechanisms responsible for the perturbation of cholesterol homeostasis in SLSD cells. In particular, they will determine whether the ability of exogenous SLs to perturb cholesterol homeostasis in cells is related to the strength of specific SL/cholesterol interactions in vitro. They will also evaluate the potential roles of several sterol-sensing proteins in modulating cholesterol homeostasis in SLSD cells; and (iv) evaluate mechanistic hypotheses modulating Sl transport targeting in SLSD cells. The role of "membrane fluidity" will be studied by modifying endogenous membrane lipids and by systematically varying the chain length and hydrophobicity of the analogs used to observe SL sorting in SLSD cells. They will also examine the function of the sterol transport protein, NPC 1, suggested by their preliminary data to play a critical role in SL targeting. These studies will test a novel concept concerning the involvement of SL/cholesterol interactions in the regulation of lipid trafficking in normal and SLSD cells.

描述（摘自申请人摘要）：研究者将