LCMR1调控DEK蛋白在肺癌细胞凋亡和增殖中的作用及机制研究

Lung cancer is one of the most common human cancers and the leading cause of cancer death worldwide. The identification of lung cancer associated genes is essential for the diagnosis and treatment of this disease. Lung cancer metastasis related protein 1 (LCMR1) is a novel gene cloned by differential display polymerase chain reaction in our laboratory. Our previous studies have confirmed that the interacting protein of LCMR1 is DEK. Our previous experiments found that inhibiting LCMR1 in cells leads to down-regulation of DEK protein expression, promoting apoptosis and inhibiting proliferation, but the specific mechanism is not clear. We speculate that "LCMR1 regulates cell apoptosis and proliferation through DEK". In order to confirm the above scientific hypothesis, we intends to use the tumor model of nude mice with LCMR1 gene knockout and lung cancer cells with LCMR1 and DEK gene knockdown respectively. Firstly, we will conform the roles and region of LCMR1 in regulating DEK protein in apoptosis and proliferation. Then, we will explore the molecular mechanism of LCMR1 in regulating the expression of DEK protein, PI3K/Akt pathway proteins, p53 pathway proteins, as well as the involved regulatory signal network. At last, we will elucidate the mechanism by which LCMR1 regulates DEK protein to participate in apoptosis and proliferation of lung cancer cells. Our research will find new molecular targets for the prevention and treatment of lung cancer from the gene level, and provide clues for the development of anticancer drugs and new strategies for the treatment of lung cancer.

肺癌发病率居于恶性肿瘤之首，严重危害人类健康和生命。肺癌转移相关蛋白1（LCMR1）是本实验室首先发现的功能基因，LCMR1可以抑制肺癌细胞凋亡促进细胞增殖，但具体作用机制尚不清楚。我们寻找到LCMR1的相互作用蛋白DEK，DEK原癌基因在细胞凋亡及增殖中发挥重要作用。课题组预实验显示抑制LCMR1表达可以引起DEK表达下降，检测肺癌细胞凋亡增加及增殖减少，我们推测“LCMR1通过DEK调控肺癌细胞凋亡和增殖”。为证实上述假设，本课题组拟开展体外及体内实验，首先明确LCMR1调节其相互作用蛋白DEK的作用机制；随后研究LCMR1通过DEK调控肺癌细胞凋亡和增殖的作用及分子机制；最后探讨LCMR1对DEK参与的PI3K/Akt及p53信号通路的调控作用，以及涉及的信号调控网络。相关研究结果为在基因水平预防和治疗肺癌寻找到新的分子靶点，为下一步将LCMR1运用于肺癌临床诊断及治疗提供分子基础。

肺癌转移相关蛋白1（LCMR1）是本实验室首先发现的功能基因，LCMR1可以抑制肺癌细胞凋亡促进细胞增殖，但具体作用机制尚不清楚。我们寻找到LCMR1的相互作用蛋白DEK，DEK原癌基因在细胞凋亡及增殖中发挥重要作用。我们推测“LCMR1通过DEK调控肺癌细胞凋亡和增殖”。本课题组开展实验，首先明确LCMR1对其相互作用蛋白DEK的调节作用；随后研究LCMR1通过DEK调控肺癌细胞凋亡和增殖的作用及分子机制；最后探讨LCMR1对DEK参与的PI3K/Akt及p53信号通路的调控作用，以及涉及的信号调控网络。.重要研究结果如下：.(1)明确LCMR1调节DEK蛋白作用机制。研究显示LCMR1与DEK（N端）功能区蛋白有直接相互作用；LCMR1能够正向调节DEK蛋白表达；LCMR1通过泛素-蛋白酶体途径改变DEK蛋白水平。.(2)明确LCMR1通过DEK调控肺癌细胞凋亡和增殖的作用。采用MTT、CCK8、流式细胞和Western Blot方法，发现LCMR1表达下调后，DEK表达下降，肺癌细胞凋亡增加及增殖减少；凋亡相关蛋白caspase-3表达增加及活性增强，增殖相关蛋白Cyclin D表达减少。.(3)明确LCMR1调控DEK在肺癌细胞凋亡和增殖的分子机制。采用Western Blot、Caspases活性免疫荧光等方法，发现LCMR1下调可增加凋亡启动蛋白Caspase-8及Caspase-9表达及活性，LCMR1通过外源性及内源性通路调控细胞凋亡。LCMR1可通过Bax、Mcl-1、c-Myc、Ras等机制调控凋亡及增殖。.(4)明确LCMR1对PI3K/Akt、p53通路的调控作用。检测LCMR1过表达或敲减后PI3K/Akt、p53通路关键蛋白表达及磷酸化活性表达，明确LCMR1通过PI3K/Akt及p53信号通路调控凋亡与增殖。.(5)确定LCMR1与DEK在肺癌凋亡和增殖中的信号调控网络。采用cDNA芯片技术检测并分析差异表达基因，通过生物信息学方法综合分析LCMR1与DEK在肺癌凋亡和增殖中共同相关的基因及信号通路。.本课题为原创性研究，为在基因水平预防和治疗肺癌寻找新的分子靶点，为抗癌药物的研发和肺癌治疗新策略提供线索，为下一步将LCMR1运用于肺癌临床诊断、治疗及预后预测提供分子基础。