MOLECULAR GENETIC ANALYSIS OF YEAST SPHINGOLIPIDS

鞘磷脂被认为在细胞间的相互作用中发挥作用， 动物生长、分化和肿瘤发生的调控。 最近的研究表明，鞘脂参与信号转导。 转导。尽管取得了很大的进展，但仍有许多事情需要了解 鞘磷脂的特殊作用及其作用机制。一 鞘磷脂研究的局限性在于缺乏实验 能够进行分子遗传分析的生物体。在.期间 在过去的资助期，我们的研究揭示了酵母菌 Cerevisiae提供将分子遗传分析与 生物化学技术在鞘磷脂合成与功能研究中的应用。 这项工作为我们提出的具体目标奠定了基础 我们将(1)检验神经鞘脂脂浓度为 部分通过调节丝氨酸的活性来维持 棕榈酰基转移酶，(2)分离肌醇-β-神经酰胺基因(S) (IPC)和神经酰胺合成酶，过表达，并纯化和 对这些酶进行了表征。IPC合成酶将受到重视，因为它 催化了一种真菌特有的反应，这应该是一个有效的目标 对于抗真菌药物，(3)确定遗传和生化基础 7R4菌株绕过制造鞘脂的能力。这个 在这种菌株中积累的独特的脂质将被表征为 确定它们是否在形成锚定的蛋白质过程中起中介作用 对于神经酰胺和磷脂酰肌醇-葡聚糖，(4)使用基因 研究鞘脂在信号中的潜在作用的策略 酿酒酵母中的转导。 我们的研究结果应该会让我们更好地理解 鞘磷脂的合成、功能(S)和真菌及其他中的调节 有机体。此外，它们还应提供一个独特的目标 设计治疗免疫功能低下所需的抗真菌药物 人类。作为我们努力的直接结果，应该可以使用 克隆酵母鞘脂生物合成基因以分离人的同源物。

Sphingolipids are thought to play roles i cell-to-cell interaction, regulation of growth, differentiation, and oncogenesis in animals. Recent developments have shown that sphingolipids participate in signal transduction. Despite great progress, much remains to be learned about the specific roles of sphingolipids and their mechanism of action. One limitation in sphingolipid research has been the lack of experimental organisms that are amenable to molecular genetic analysis. During the past grant period our research revealed the potential that Saccharomyces cerevisiae offers for combining molecular genetic analysis with biochemical techniques to studies of sphingolipid synthesis and function. This work has laid the foundation for our proposed Specific Aims in which we will (1) examine the hypothesis that sphingolipid concentrations is maintained in part by regulating the activity f serine palmitoyltransferase, (2) isolate the gene(s) for inositol-P-ceramide (IPC) and ceramide synthetases, overexpress them, and purify and characterized the enzymes. IPC synthetase will be emphasized since it catalyzed a reaction unique to fungi that should be an efficacious target for antifungal drugs, (3) determine the genetic and biochemical basis for the ability of strain 7R4 to bypass the need to make sphingolipids. The unique lipids that accumulate in this strain will be characterized to determine if they are intermediated in the formation of proteins anchored to ceramide and to phosphatidylinositol-glycans, (4) use a genetic strategy to examine the potential role of sphingolipids in signal transduction in S. cerevisiae. The results of our studies should provide a better understanding of sphingolipid synthesis, function(s), and regulation in fungi and other organisms. In addition, they should provide a unique target for designing antifungal drugs needed for treatment of immune compromised humans. As a direct results of our efforts it should be possible to used clone yeast sphingolipid biosynthetic genes to isolate human homologs.