摘要
背景与目的:组蛋白赖氨酸去甲基化酶1(lysine specific demethylase 1,LSD1)是重要的染色质修饰蛋白之一,可以通过调节染色质的结构调节基因的转录调控,进而影响肿瘤的发生、发展、侵袭、转移以及代谢异常等恶性潜能,是判断肿瘤预后的生物标志物。Sirtuins家族去乙酰化酶3(sirtuin3,SIRT3)基因是位于线粒体内的抑癌基因,通过调控肿瘤代谢异常以及氧化损伤行使抑癌基因的功能。本研究通过基因转录调控的手段,研究胰腺癌细胞PANC-1中LSD1与SIRT3的关系。方法:通过RNA干扰(RAN interference,RNAi)、免疫共沉淀(co-immunoprecipitation,CoIP)、染色质免疫共沉淀(chromatin immunoprecipitation assay,ChIP)及启动子活性分析等分子生物学实验手段,探讨LSD1与SIRT3在PANC-1细胞中的关系。结果:通过RNAi的手段干扰LSD1的表达,发现SIRT3基因转录水平和蛋白水平明显上升;通过蛋白相互作用的手段,发现LSD1可以与SIRT3转录调控的重要转录因子过氧化物酶增殖体激活受体辅激动子-1α(peroxisome proliferator-activated receptor gamma,coactivator 1 alpha,PGC-1α)相互作用;通过ChIP方法,发现LSD1与PGC-1α共同募集到SIRT3基因的启动子区域染色质上;通过启动子活性分析,发现LSD1基因可以显著抑制PGC-1α对SIRT3基因的转录调控。结论:LSD1可以表观遗传调控抑癌基因SIRT3的转录,为深入研究LSD1与肿瘤代谢异常以及氧化应激提供了理论依据。
Background and purpose： Lysine specific demethylase I（LSD1） is an important chromatin modifier. It epigenetically regulates gene expression pattern through chromatin modification and participates in maintenance of tumor malignant properties, such as oncogenesis, development, invasion, migration and metabolic transformation. SIRT3 （sirtuin 3） transformation and oxidative stress is a mitochondria localized tumor The correlation between LSD1 and suppressor and regulates tumor metabolic SIRT3 has never been reported before. This study aimed to elucidate the correlation between LSD1 and SIRT3 with gene transcriptional regulation methods. Methods： RNA interference technique, co-immunoprecipitation assay（ColP）, chromatin immune-precipitation assay（ChiP） and firefly luciferase activity assay were employed to elucidate the correlation between LSD1 and SIRT3 in pancreatic cancer. Results： mRNA and protein levels of SIRT3 were significantly elevated in LSD1 knock-down PANC-1 cells. LSD1 interacts with PGC-lct, an important regulator of SIRT3 gene expression. LSD1 and PGC-1α occupied the same region in SIRT3 promoter region through ChIP analysis. Luciferase activity assay validated LSD1as a negative regulator of PGC-1α in SIRT3 gene transcriptional regulation. Conclusion： LSD1, as an important tumor promoter, negatively regulates the expression of tumor suppressor gene SIRT3, these results provide important clues for the role that LSD1 plays in aberrant metabolism and oxidative stress.
出处
《中国癌症杂志》
CAS
CSCD
北大核心
2014年第2期87-92,共6页
China Oncology
