SOX10蛋白乙酰化修饰作用调控肿瘤恶性进展的机制及临床意义

SOX10 is reported to be upregulated in variety of malignant tumors including melanoma and its upregulation is positively correlated with the cancer progression. However, the mechanisms of SOX10 being upregulated in these cancers remain unknown. Recently, we reported that the E3 ligase Fbxw7 negatively regulated the stability of SOX10. In this project, thepreliminary results indicated thatCBPacetylated SOX10 in K256 sites and this acetylation resulted in the enhanced stability of SOX10.Based on these observations, we made the hypothesis of this project. The acetylation of SOX10 by CBP inhibits the ubiquitination mediated degradation of SOX10 by Fbxw7. The dysregulation of Fbxw7 and CBP leads the upregulation of SOX10 and thus the progression of malignant cancers. We will validate our hypothesis by variety of experiments. Firstly, we will explore the mechanisms by which CBP and Fbxw7 regulate the stability using mutation, ubiquitination and CHX assay etc. Secondly, we will develop the acetylation site mutated (K/R, K/Q) colorectal cells using CRISP-CAS9 system and examined their functional changes including proliferation, apoptosis and migratory ability in vitro and in vivo. To reveal the clinical significance of SOX10 acetylation by CBP, we will collect 200 melanomaand glioma samples and explore the correlation of acetylated SOX10 with CBP expression. Besides, we will also analyze the correlation of SOX10 acetylation level with the clinical characteristic and overall survival of cancer patients. This study would strive to elucidate the mechanisms and clinical significance of SOX10 acetylation on the cancer progression.

SOX10在黑色素瘤等多种肿瘤中表达上调，其在肿瘤中表达上调的机制尚未阐明。课题组前期报道了SOX10稳定性受E3连接酶Fbxw7负调控。在本项目中，我们发现乙酰化修饰正调控SOX10的稳定性。预实验结果显示CBP介导SOX10蛋白K256位乙酰化修饰，增强SOX10的稳定性。据此我们提出科学假设：CBP介导的乙酰化修饰能抑制Fbxw7对SOX10的泛素化降解作用，CBP与Fbxw7的表达失衡，导致SOX10在肿瘤中的表达上调，促进肿瘤的恶性进展。项目接下来将深入研究CBP与Fbxw7共同调控SOX10稳定性的机制；使用CRISP-CAS9系统建立SOX10 K256突变的黑色素瘤细胞株，在细胞与动物水平验证乙酰化修饰对肿瘤恶性进展的影响；在临床标本中分析SOX10乙酰化水平与CBP表达的相关性，以及与肿瘤病人总的生存率、预后的相关性，揭示乙酰化修饰调控SOX10稳定性的机制及临床意义。

黑色素瘤是一类高度恶性的肿瘤，约50%的黑色素瘤发生BRAF V600E突变，BRAF突变是黑色素瘤发生的驱动因素，此外，SOX10表达上调也导致黑色素瘤的发生。.探讨SOX10表达上调的机制有助于了黑色素瘤的发生发展机制，发现黑色素瘤临床诊断标志物以及治疗靶点。本项目发现SOX10受乙酰化修饰作用，K253位是SOX10的乙酰化位点。乙酰化修饰促进SOX10蛋白的稳定性，抑制SOX10蛋白的泛素化修饰，从而促进SOX10的表达。CBP是介导SOX10乙酰化修饰的乙酰转移酶，敲低CBP降低SOX10的乙酰化修饰，以及SOX10的蛋白表达水平。重要的是，敲低CBP降低黑色素细胞的迁移侵袭。.BRAF抑制剂维罗菲尼等对BRAF V600E突变的黑色素瘤具有良好的疗效，然而耐药常常发生，限制了黑色素瘤病人的受益，探讨BRAF抑制剂耐药机制并找出克服耐药的新策略有助于提高黑色素瘤病人的生存时间及生活质量。本项目研究发现NONO蛋白通过上调CRAF/ARAF，使BRAF下游信号通路pERK1/2不依赖于BRAF而激活，导致黑色素瘤BRAF抑制剂耐药。NONO蛋白受p300介导的乙酰化修饰，促进NONO蛋白表达。在耐药黑色素瘤中，乙酰化修饰导致NONO蛋白表达上调，进而激活CRAF/ARAF-pERK1/2信号通路。此外pERK1/2上调又反过来促进p300的表达，导致NONO表上调，形成一个p300-NONO-CRAF/ARAF-pERK1/2正反馈回路，促进黑色素瘤更加耐药。P300抑制剂处理下调NONO蛋白表达，进而下调CRAF/ARAF-pERK1/2信号通路，部分恢复耐药细胞对BRAF抑制剂维罗菲尼等的敏感性，p300抑制剂有望成分克服黑色素瘤BRAF抑制剂耐药的新选择。