MAPK15在三氧化二砷诱导的细胞凋亡中的作用及其分子机制研究

Arsenic trioxide can induce apoptosis, and is a promising drug commonly used for the treatment of acute promyelocytic leukemia and a variety of solid tumors. Studies have reported that arsenic trioxide treated cells showed reactive oxygen species (ROS) accumulation and NF-kB activation. Our study discovered that high MAPK15 expression in cells can promote apoptosis induced by arsenic trioxide. Due to the limited knowledge on the function of MAPK15 and its signal transduction, the underlying mechanism of this phenomenon requires further study. Preliminary results show that IkBa is a potential substrate phosphorylated by activated MAPK15 upon arsenic trioxide stimulation where this phosphorylation is likely linked to the ubiquitylation and degradation of IkBa, and modulates the subsequent NF-kB activity. From the basis of these findings, in the present project, from molecular, cellular and animal levels, we want to study the mechanisms and interaction of MAPK15 and the NF-kB pathway, the role of NF-kB signaling in MAPK15 promoted arsenic trioxide-induced apoptosis, and the impact of ROS on the MAPK15 activity and NF-kB signal transduction. The completion of this project will provide an in-depth understanding of the molecular mechanisms of NF-kB in the participation of apoptosis and the biological functions of MAPK15, and open up application for the scientific use of oxidant as anticancer drugs.

三氧化二砷能够诱导细胞凋亡，是治疗急性早幼粒细胞白血病的常用药物，有望用于多种实体瘤的治疗。有研究报道，三氧化二砷处理的细胞检测到活性氧ROS堆积和NF-kB的活化。我们研究中发现，MAPK15高表达能促进三氧化二砷诱导的细胞凋亡。由于对MAPK15的功能与信号转导认识有限，背后的机制有待进一步研究。预实验结果显示，IkBa是三氧化二砷刺激后活化的MAPK15潜在的磷酸化底物，该磷酸化很可能与IkBa的泛素化及随后的降解相关，影响NF-kB转录因子活性。本项目拟在此基础上，从分子、细胞和动物水平，研究NF-kB通路与 MAPK15相互作用的分子机制，NF-kB信号在MAPK15促进三氧化二砷诱导的细胞凋亡中的作用，并研究ROS对MAPK15活性及NF-kB信号转导的影响。本课题的完成将有助深入认识NF-kB参与细胞凋亡的分子机制和MAPK15的生物学功能，为科学使用氧化剂类抗癌药提供参考。

胞外信号调节激酶8（ERK8），也被称为有丝分裂原激活的蛋白激酶15（MAPK15），是最近发现的蛋白激酶ERK家族成员，目前对其研究较少，认识有限。我们在研究中发现，不同的肺癌细胞对抗癌药物三氧化二砷（As2O3）表现出不同的敏感性，通过对肺癌细胞中MAPK15表达水平的筛查，发现MAPK15在几种人肺癌细胞株中的的表达水平和其三氧化二砷的敏感性呈正相关。改变细胞内MAPK15的表达水平，能够改变细胞对三氧化二砷的反应性。分子机制的研究表明，三氧化二砷能够诱导MAPK15的TEY激酶结构域上Thr175和Tyr177位点的磷酸化，使其激活。活化的MAPK15又能进一步与IkappaBalpha (IkBa)相互作用，直接磷酸化IkBa的 Ser32/36位点，导致其降解，从而引发核促进因子-kB（NFkB p65）的核转位，促进其与染色质结合，诱导和激活参与细胞凋亡的下游蛋白质分子，导致细胞凋亡的发生。通过稳定knockdown肺癌H1299细胞中MAPK15的表达和裸鼠移植瘤成瘤和治疗实验，证实内源性MAPK15-IkBa-NFkB信号通路对细胞的生物学作用， MAPK15特异敲除减低了细胞对三氧化二砷治疗的敏感性，表明MAPK15在三氧化二砷治疗中潜在的中介效应。综上所述，我们的研究结果第一次阐明了MAPK15调节NFkB的激活模式，提示MAPK15可以作为在肺癌治疗中选择三氧化二砷方案的潜在的生物标志分子，或作为协同用药的靶点。