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SPHINGOLIPID BIOSYNTHESIS IN YEAST

酵母中的鞘脂生物合成

基本信息

批准号：
2634757
负责人：
TERESA M DUNN
金额：
$ 17.64万
依托单位：
HENRY M. JACKSON FDN FOR THE ADV MIL/MED
依托单位国家：
美国
项目类别：
财政年份：
1995
资助国家：
美国
起止时间：
1995-01-01 至 2000-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2634757
关键词：
Saccharomyces cerevisiae alleles genetic regulation lipid biosynthesis molecular cloning mutant nucleic acid sequence protein sequence sphingolipids western blottings yeasts

项目摘要

Sphingolipids are essential in all eukaryotic cells and alterations in sphingolipid metabolism occur in many disease processes. Although a variety of cellular functions for sphingolipids and their metabolites have been proposed, little is known about their biosynthesis and physiological roles. Sphingolipid synthesis in the model eukaryote, S. cerevisiae will be investigated using a combined genetic/biochemical approach. A collection of mutants (csg2, scs1-scs7) with defects in sphingolipid metabolism has been isolated, and will be used to isolate the wild type genes required for sphingolipid synthesis. The scs mutants will be transformed with a genomic library and the wild type genes cloned by complementation of the scs phenotype. The function of each SCS gene will be probed by determining the biochemical consequence of disrupting the gene, by comparing the amino acid sequence of the SCS gene products with the protein data base, by investigating the epistatic relationships between scs, csg, sec and pmr mutants, and by identifying the cellular location of the gene products. Determination of the epistatic relationships between the csg2, scs, sec, and pmr mutants will provide insight into the biosynthetic pathway (both the order of action of the biosynthetic enzymes and their cellular location). Suppressor analysis of scs genes will be used to identify other genes and proteins in sphingolipid synthesis not found in the CSG and SCS collections. Since our scs mutant collection consists of several alleles of each scs gene, mutations that alter the activity in interacting proteins as well as those that bypass the scs gene function altogether will be identified. The results of these experiments should provide valuable information about the genes and proteins that catalyze and regulate sphingolipid synthesis as well as the roles of sphingolipids in all eukaryotic cells.

鞘脂在所有真核细胞中都是必需的，鞘脂代谢发生在许多疾病过程中。虽然鞘脂及其代谢物的各种细胞功能虽然已经提出，但对其生物合成和生理特性知之甚少。角色在模式真核生物S.酿酒厂将使用遗传/生物化学相结合的方法进行研究。一具有鞘脂缺陷的突变体（csg 2，scs 1-scs 7）的集合代谢已被分离，并将用于分离野生型鞘脂合成所需的基因。 scs的变种人用基因组文库转化，并通过 SCS表型的互补。每个SCS基因的功能将通过确定破坏细胞的生物化学后果来探测，基因，通过比较SCS基因产物的氨基酸序列与蛋白质数据库，通过研究上位关系在scs，csg，sec和pmr突变体之间，并通过鉴定细胞基因产物的位置。上位性的确定 csg 2、scs、sec和pmr突变体之间的关系将提供深入了解生物合成途径（无论是行动的顺序，生物合成酶及其细胞定位）。抑制因子分析 scs基因将被用来鉴定其他基因和蛋白质，在CSG和SCS集合中未发现鞘脂合成。由于我们 SCS突变体集合由每个SCS基因的几个等位基因组成，改变相互作用蛋白质活性的突变，完全绕过SCS基因功能的细胞将被鉴定出来。的这些实验的结果应该提供关于催化和调节鞘脂合成的基因和蛋白质，以及鞘脂在所有真核细胞中的作用。