CD82棕榈酰化修饰对抑制肿瘤淋巴结转移能力的影响及分子机制的研究

KAI1/CD82 is a tumor metastasis suppressor factor, the mechanism has not been clarified. Our previous work found that CD82 had synergistic effects on the activation of some growth factor receptors (EGFR, cMet), downregulation of PI3K/AKT signal transduction pathway activity and inhibition of cell migration with the cell membrane of ganglioside GM2/GM3. Research shows that the CD82 has function in formation of CD82 membrane protein complexes (Tetraspanin Web) , and the palmitoylation of CD82 plays an important role in the integrity of the inhibition of tumor invasion. We speculate that① the palmitoylation is an important molecular structure of mutual recognition and interaction between CD82 and other related molecules; ② the protein structure formed by CD82 palmitoylation is the important fundamental structure involved in the correct formation of CD82 complex in the membrane. To confirm this hypothesis, we intend to construct 7 different palmitoylation site mutation of CD82 mutants, using molecular biology, mass spectrum analysis technology, to investigate the effect of the palmitoylation modification of CD82 in the formation of the CD82 complex, signal transduction, cell migration ability and lymph node metastasis ,then explore its molecular mechanism. This topic will clarify that the molecular mechanism of CD82 inhibiting tumor metastasis from a new point of view, and provide new theory and new drug targets of new therapeutic drugs for inhibiting tumor metastasis.

KAI1/CD82是广谱的肿瘤转移抑制因子,作用机制尚未阐明。我们前期研究发现CD82与细胞膜上神经节苷脂GM2/GM3具有协同作用，下调生长因子受体（EGFR和cMet）介导的PI3K/AKT信号通路活性，从而抑制细胞迁移能力。研究表明CD82发挥功能依赖于CD82膜蛋白复合体（Tetraspanin Web）的形成，且CD82的棕榈酰化对于其功能完整性具有重要作用。我们推测①棕榈酰化是CD82与其它相关分子之间相互识别、相互作用的重要分子结构；②棕榈酰化修饰形成的蛋白结构是参与CD82复合体形成的重要基础结构。我们拟通过构建7种不同棕榈酰化位点突变的CD82突变体，采用分子生物学、质谱分析等技术，探讨棕榈酰化对CD82 复合体的形成、信号转导通路及淋巴结转移能力的影响及其分子机制。本课题将从新的视角阐明CD82抑制肿瘤转移的分子机制，并为抑制肿瘤转移新的治疗药物提供新理论和新靶点。

广谱的转移抑制基因CD82 /KAI1，是四跨膜蛋白家族的成员之一，研究表明通过四跨膜蛋白微域（Tetraspanin Web）发挥其活性。然而，CD82棕榈酰化在该微域的作用尚未完全阐明。本研究的目的是通过对CD82的棕榈酰化的突变，来研究细胞的迁移和凋亡，并观察这种突变与表皮生长因子受体(EGFR)和肝细胞生长因子受体(HGFR / cMet)的活化相关性。我们发现乳腺癌细胞转染CD82棕榈酰化的突变的基因，显示细胞凋亡增加。此外，CD82的棕榈酰化突变会选择性激活EGFR和cMet不同的酪氨酸位点。这些结果表明, CD82在半胱氨酸5位点或半胱氨酸83位点单独突变或半胱氨酸5位点和半胱氨酸74位点同时突变或半胱氨酸74位点和半胱氨酸83位点同时突变，细胞凋亡增加, 然而, CD82其他形式的棕榈酰化突变对细胞凋亡没有影响。总之,我们的研究结果有力地表明, 仅有CD82半胱氨酸位点74位点或/和半胱氨酸83位点的棕榈酰化促使CD82在质膜上的表达具有重要的作用; CD82的半胱氨酸位点5位点和半胱氨酸83位点同时棕榈酰化与维持CD82 的生物学功能密切相关。