Microdomain abnormalities due to aberrant glycolipids

在本项目中，我们试图研究肿瘤细胞和神经细胞上表达的神经鞘糖脂在生物信号调节中的作用，并阐明其异常导致的发病机制。为了达到这些目的，我们分析了：1.基于糖基化模式重塑的黑色素瘤和小细胞肺癌中糖脂信号的调节机制；2.糖基转移酶基因敲除小鼠异常表型的建立和分析，以阐明糖脂在体内的作用。在黑色素瘤细胞中，GD3的特征性表达诱导了p130Cas和paxlin等接头分子酪氨酸磷酸化的增强，并增加了细胞的生长和侵袭活性。此外，粘着斑激酶(FAK)在GD3+细胞中的激活程度也高于GD3-细胞。另一方面，GD2的表达促进了小细胞肺癌细胞…的增殖和侵袭抗GD2抗体的增多和结合可诱导小细胞肺癌细胞发生凋亡。结果表明，抗GD2抗体可以诱导FAK的去磷酸化和p38的激活，从而导致失巢。因此，抗GD2抗体可触发失巢凋亡，破坏GD2的分子复合体是必需的。整合素和FAK是一种有效的肿瘤治疗策略。至于神经鞘糖脂在神经组织中的作用，人们一直怀疑酸性鞘糖脂在神经系统的发育和功能中发挥着重要作用，因为它们的高水平表达。为了阐明它们在神经组织中的作用，我们建立了基因敲除小鼠系，即GM2/GD2合成酶、GD3合成酶、GM3合成酶和乳糖基神经酰胺合成酶的双重敲除小鼠。作为对这些突变小鼠的表型分析的结果，我们已经证明它们根据神经节苷脂结构缺陷的范围而表现出异常的表型变化。一般来说，酸性糖脂似乎在维持神经组织的完整性和神经元损伤后的修复中是必不可少的。此外，对野生型和双基因敲除小鼠神经组织基因表达谱的比较表明，突变小鼠的神经变性不是单纯的萎缩性改变，而是伴随着炎症过程的活跃变化，表现为补充系统的激活和细胞因子的产生或分泌。较少

In this project, we have tried to investigate roles of glycosphingolipids expressed on cancer cells and neuronal cells in the regulation of biosignals, and to clarify mechanisms for the pathogenesis due to their abnormalities. In order to achieve these aims, we analysed ; 1. regulatory mechanisms of signaling with glycilipids in melanomas and small cell lung cancers based on the remodeling of glycosylation patterns, 2. establishment and analysis of abnormal phenotypes of gene knockout mice of glycosyltransferases to elucidate roles of glycolipids in vivo. In melanoma cells, characteristic expression of GD3 induced enhancement of tyrosine phosphorylation of adaptor molecules such as p130Cas and paxillin, and increased cell growth and invasion activity. Furthermore, focal adhesion kinase (FAK) was also activated more strongly in GD3+ cells than in GD3- cells. On the other hand, GD2 expression resulted in the enhancement of cell proliferation and invasion in small cell lung cancer cells, … More and binding of anti-GD2 antibodies could trigger apoptosis of small cell lung cancer cells. It was, then, demonstrated that anti-GD2 antibodies could induce dephosphorylation of FAK and activation of p38, leading to anoikis. Consequently, it was concluded that anti-GD2 antibodies trigger anoikis, and it was essential to destroy molecular complex sonsisting of GD2. Integrin and FAK as an efficient strategy toward cancer therapeutics.As for roles of glycosphingolipids in nervous tissues, it has been suspected that acidic glycosphingolipids paly important roles in the development and function of nervous systems based on their high levels of expression. In order to clarify their roles in the nervous tissues, we generated gene knockout mice lines, i. e. knockout mice of GM2/GD2 synthase, GD3 synthase, double knockout of those two, GM3 synthase, and lactosylceramide synthase. As results of phenotypic analyses of these mutant mice, we have demonstrated that they showed abnormal phenotypic changes according to the range of defects in ganglioside structures. Generally, acidic glycolipids appeared to be essential in the maintenance of the integrity of the nervous tissues and repair after neuronal damages. Furthermore, comparison of gene expression profiles in the nerve tissues between wild type and double knockout mice revealed that neurodegeneration detected in the mutant mice are not mere atrophic changes, but active changes with inflammatory process as indicated by the activation of complementary system and cytokine production or secretion. Less