乳酸脱氢酶LDHA琥珀酰化修饰促进胃癌细胞糖酵解的功能机制研究

Lactate dehydrogenase (LDH)A act as a key glycolytic enzyme that plays an important role in tumor cell metabolism.As a nearly found style about post-translational modification of protein， ,Lysine succinylation is widely involved in cell proliferation, differentiation and energy metabolism. In the previous study, we found that there was a significant difference about the protein succinylation level in gastric cancer tissues compared with normal gastric mucosa tissues, in which the level of lysine succinylation at the 222nd site of LDHA in gastric cancer tissues was significantly increased. Pre-test showed that the succinyltransferase KAT2A and the de-succinylase Sirt5 can change the succinylation level of LDHA, while LDHA can promote the proliferation and migration of gastric cancer cells. We hypothesize that LDHA regulates the succinylation at K222 site by KAT2A and Sirt5 in gastric cancer cells, which is involved in the regulation of ubiquitin-proteasome and autophagy-lysosomal degradation pathways。Increased LDHA in turn activates glycolysis and promotes the progression of gastric cancer. This project intends to study the succinylation site and regulatory mechanism of LDHA in level of molecular, cellular, tissue and animal. The topic could reveal the role of succinylation of LDHA in the glycolytic metabolism of gastric cancer and can clarify its clinical relevance in the progression of gastric cancer , provide a new strategy for the prevention and treatment of gastric cancer.

乳酸脱氢酶(LDH)A是糖酵解关键酶，在肿瘤细胞代谢中起关键作用。赖氨酸琥珀酰化是新近发现的蛋白质翻译后修饰方式，广泛参与细胞增殖、分化及能量代谢。前期研究中我们发现胃癌组织中蛋白质琥珀酰化修饰较癌旁正常胃粘膜组织有显著差异，其中胃癌组织LDHA第222位点赖氨酸琥珀酰化水平明显增高。敲除实验结果显示琥珀酰转移酶KAT2A及去琥珀酰化酶Sirt5可以改变LDHA的琥珀酰化水平，而LDHA能促进胃癌的增殖及迁移。我们提出假想：胃癌细胞中LDHA通过KAT2A和Sirt5调节 K222位点琥珀酰化修饰，参与调控其泛素-蛋白酶体及自噬-溶酶体降解途径，进而活跃糖酵解，促进胃癌的进展。本课题拟从分子、细胞、组织以及动物水平等多层次研究LDHA的琥珀酰化位点和调节机制，揭示LDHA琥珀酰化修饰在胃癌糖酵解代谢中的作用，明确其在胃癌进展中的临床相关性，为胃癌的防治提供新策略。

赖氨酸琥珀酰化是一种新兴的翻译后修饰，近年来得到了越来越多的关注，但其在胃癌中的作用仍有待进一步研究。我们在胃癌患者正常组织与肿瘤组织中共鉴定出303个蛋白的503个赖氨酸琥珀酰化位点。质谱数据显示沃伯格效应相关的关键代谢酶乳酸脱氢酶A (LDHA)的赖氨酸K222位点发生较高水平的琥珀酰化修饰。其中重要的是，这种修饰并不影响LDHA的泛素化作用，但降低了泛素化LDHA与SQSTM1的结合，从而抑制了其溶酶体降解途径。在研究中我们证明了CPT1A作为一种赖氨酸琥珀酰转移酶，与LDHA相互作用并使其琥珀酰化。此外，LDHA的K222位点琥珀酰化与GC患者预后与生存存在直接关系。最后，通过过表达与干扰琥珀酰化修饰LDHA可以调节细胞增殖、侵袭和迁移。揭示了一种新的LDHA降解的溶酶体途径是通过k63泛素化LDHA与自噬相关功能蛋白1（SQSTM1）介导的。肉碱棕榈酰转移酶（CPT1A）通过琥珀酰化修饰LDHAK222位点，抑制LDHA的降解，促进GC侵袭和增殖。