巨噬细胞转运SIRT2去除肿瘤微环境蛋白乙酰化促进肿瘤转移的机制研究

Protein post-translational modifications precisely regulate the metastasis of tumor cells in the tumor microenvironment. Proteomics studies revealed that there are lots of acetylation modifications in extracellular proteins, and the mechanism has not been elucidated. We found that Toll-like receptor activated macrophage secretes the deacetylase SIRT2 into the tumor microenvironment. This secretory process relies on the E3 ubiquitin ligase activity of TRAF6. Secreted SIRT2 promotes invasion and migration of lung cancer cells. Based on these, we hypothesized that TRAF6 catalyzes the ubiquitination of SIRT2 in activated macrophages, promoting its secretion into the tumor microenvironment, and secreted SIRT2 induces deacetylation of extracellular proteins and promotes metastasis of tumor cells. This project intends to use immuno-precipitation, immuno-fluorescence, immuno-electron microscopy, proteomics, mouse tumor model etc, 1) to elucidate the mechanism of TRAF6 promoting SIRT2 secretion in macrophages, and 2) to identify the substrate of SIRT2 in extracellular microenvironment, 3) to reveal the biological function that secreted SIRT2 promotes the tumor development by reducing extracellular substrates acetylation. The aim of this project is to reveal the relationship between the secretion of SIRT2 and the acetylation level in tumor microenvironment that promotes tumor metastasis, and provide a new theoretical basis and drug target for tumor therapy.

肿瘤微环境中的蛋白翻译后修饰精密调控肿瘤细胞的迁移。蛋白质组学研究发现胞外蛋白存在丰富的乙酰化修饰，其具体机制尚未阐明。我们发现，Toll样受体活化的巨噬细胞，能分泌去乙酰化酶SIRT2到肿瘤微环境，且分泌过程依赖TRAF6介导的泛素化修饰。分泌的SIRT2促进肺癌细胞的侵袭迁移。据此我们提出假说：Toll样受体活化的巨噬细胞中TRAF6催化SIRT2泛素化，促进其分泌至胞外微环境诱导了胞外蛋白的去乙酰化，促进肿瘤细胞的迁移。本项目拟用免疫沉淀、免疫荧光、免疫电镜、蛋白组学、小鼠肿瘤模型等方法，1）阐明巨噬细胞中TRAF6促进SIRT2分泌机制，2）鉴定SIRT2在胞外微环境中的底物蛋白，3）揭示分泌的SIRT2通过降低细胞外底物乙酰化水平促进肿瘤发展的生物功能。本项目旨在揭示肿瘤微环境中巨噬细胞分泌SIRT2去除底物乙酰化水平与肿瘤发展的关系，为肿瘤治疗提供新的理论依据和药物靶点。

肿瘤微环境中的蛋白翻译后修饰精密调控肿瘤细胞的迁移。细胞外蛋白在细胞内乙酰化后转运到微环境，然而它们在肿瘤微环境中的去乙酰化调控仍不清楚。我们发现了多种乙酰基vWA结构域携带蛋白，包括整合素3（ITGB3）和胶原6A（COL6A）在细胞外被Sirtuin家族成员SIRT2去乙酰化。Toll样受体（TLR）家族成员TLR4或TLR2活化的巨噬细胞分泌SIRT2。TLR激活的SIRT2通过自噬体转运。肿瘤坏死因子受体相关因子6（TRAF6）介导SIRT2的分泌。在细胞外空间，分泌的SIRT2去乙酰化细胞外蛋白，促进细胞附着和迁移，进而促进癌细胞转移。因此，细胞外空间是亚细胞细胞器样的场所，分泌的SIRT2通过催化细胞外蛋白脱乙酰化促进癌细胞转移。