Sphingolipid Metabolism in Drosophila Development

描述（由申请人提供）：鞘脂是建立在鞘氨醇碱骨架上的保守脂质家族。鞘脂具有结构膜功能，而它们的代谢产生参与调节哺乳动物发育、免疫功能、炎症和细胞应激反应的信号分子。该基金支持的研究探索了鞘脂在模式生物果蝇中的作用。这些研究导致果蝇鞘氨醇碱的化学表征和鉴定新的内源性果蝇鞘脂具有强效的生长抑制活性。此外，我们已经表明，缺乏果蝇鞘氨醇-1-磷酸（S1 P）裂解酶（SPL），这是负责鞘脂降解的最后一步表达的Sply突变体，积累鞘脂中间体，并在动力飞行所需的胸肌中发展为进行性肌病。Sply肌病通过减少鞘脂产生而得到纠正，表明鞘脂中间体起致病作用。我们已经观察到关键膜蛋白如发动蛋白和肌营养不良蛋白聚糖的突变体与Sply肌病表型相似。我们还进行了基于细胞的研究，表明Sply/SPL是正常AKT信号传导、蛋白质翻译、成肌细胞融合、成肌细胞基因表达和自噬控制所必需的。这些相互作用中的许多已经在鼠C2 C12细胞中得到证实，表明SPL在成肌细胞存活、融合和分化中起保守作用。我们还发现鞘氨醇激酶和S1 P裂解酶在小鼠肌肉再生过程中动态上调，导致再生肌肉中S1 P水平的瞬时峰值。这些集体观察使我们提出了我们的中心假设，即SPL在肌肉生物学，发育和稳态中起着关键作用。因此，我们建议的具体目标是：1）定义SPL在肌肉细胞生物学中的作用; 2）剖析SPL在肌肉发育中的作用; 3）建立SPL在肌肉萎缩和再生中的作用。我们的长期目标是利用果蝇阐明鞘脂在生物学、膜功能和组织稳态中的作用，并确定这些发现与人类疾病的相关性。由于鞘脂的物种特异性结构差异和低等真核生物中缺乏S1 P受体，我们不期望这两个系统是等同的。因此，我们的目标是比较果蝇和小鼠细胞和动物模型中SPL损失相关的生化和分子事件，目的是全面了解SPL在肌肉组织中的功能以及其功能在整个进化过程中如何被修改。我们的团队在鞘脂生物化学、果蝇遗传学以及MD的遗传学和病理学方面具有丰富的经验，这些研究应该很容易完成。

DESCRIPTION (provided by applicant): Sphingolipids are a conserved family of lipids built upon a sphingoid base backbone. Sphingolipids serve structural membrane functions, whereas their metabolism produces signaling molecules involved in regulating mammalian development, immune function, inflammation and cellular stress responses. Studies supported by this grant have explored the role of sphingolipids in the model organism Drosophila melanogaster. These studies have resulted in the chemical characterization of Drosophila sphingoid bases and the identification of novel endogenous Drosophila sphingolipids with potent growth-inhibitory activity. Further, we have shown that Sply mutants lacking expression of Drosophila sphingosine-1-phosphate (S1P) lyase (SPL), which is responsible for the final step of sphingolipid degradation, accumulate sphingolipid intermediates and develop a progressive myopathy in the thoracic muscles needed to power flight. The Sply myopathy is corrected by reducing sphingolipid production, indicating sphingolipid intermediates play a causative role. We have observed that mutants in key membrane proteins such as dynamin and dystroglycan phenocopy the Sply myopathy. We have also conducted cell-based investigations that suggest that Sply/SPL is required for normal AKT signaling, protein translation, myoblast fusion, myoblast gene expression and control of autophagy. Many of these interactions have been corroborated in murine C2C12 cells, indicating that SPL plays a conserved role in myoblast survival, fusion and differentiation. We have also found that sphingosine kinase and S1P lyase are dynamically upregulated during murine muscle regeneration, leading to a transient peak in S1P levels in regenerating muscle. These collective observations have led us to propose our central hypothesis, which states that SPL plays a critical role in muscle biology, development and homeostasis. The specific aims of our proposal are, thus: 1) To define the role of SPL in muscle cell biology; 2) To dissect the role of SPL in muscle development; and 3) To establish the role of SPL in muscle atrophy and regeneration. Our long-term goals are to exploit Drosophila to elucidate the role of sphingolipids in biology, membrane function, and tissue homeostasis and to define the relevance of these findings to human disease. Due to species-specific structural differences in sphingolipids and the lack of S1P receptors in lower eukaryotes, we do not expect the two systems to be equivalent. Thus, our goals are to compare the biochemical and molecular events associated with SPL loss in Drosophila and murine cell and animal models, with the intent of developing a comprehensive understanding of how SPL functions in the context of muscle tissue and how its function may have been modified throughout evolution. These studies should be readily achieved by our team, which has extensive experience in sphingolipid biochemistry, Drosophila genetics, and the genetics and pathology of MD.