Sphingolipid Metabolism in Drosophila Development

描述(申请人提供)：鞘磷脂是建立在狮身人面像基础脊椎上的一种保守的脂类家族。鞘磷脂具有结构膜功能，而它们的新陈代谢产生信号分子，参与调节哺乳动物的发育、免疫功能、炎症和细胞应激反应。由这笔赠款支持的研究探索了鞘磷脂在模式生物黑腹果蝇中的作用。这些研究结果导致了果蝇狮身人面像碱基的化学表征和具有强大生长抑制活性的新型内源性果蝇鞘脂的鉴定。此外，我们已经证明，缺乏果蝇鞘氨醇-1-磷酸(S1P)裂解酶(SPL)表达的Sply突变体，负责鞘磷脂降解的最后一步，积累鞘磷脂中间体，并在飞行所需的胸肌中发展进行性肌病。鞘磷脂产生减少可纠正鞘脂肌病，表明鞘脂中间体起致病作用。我们已经观察到，关键膜蛋白的突变，如动力蛋白和营养不良糖链，表现为Sply肌病。我们还进行了基于细胞的研究，表明Sply/SPL是正常的AKT信号、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。其中许多相互作用已经在小鼠C2C12细胞中得到证实，表明SPL在成肌细胞的存活、融合和分化中发挥着保守的作用。我们还发现，在小鼠肌肉再生过程中，鞘氨醇激酶和S1P裂解酶动态上调，导致再生肌肉中S1P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、发育和动态平衡中发挥着关键作用。因此，我们建议的具体目标是：1)确定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)确定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来阐明鞘磷脂在生物学、膜功能和组织动态平衡中的作用，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以及低等真核生物中S1P受体的缺乏，我们预计这两个系统不会对等。因此，我们的目标是比较果蝇、小鼠细胞和动物模型中与SPL丢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可能被修改的方式。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拥有丰富的经验，这些研究应该很容易实现。

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.