Ciribilli Yari, Singh Prashant, Spanel Reinhard, Inga Alberto, Borlak Jürgen
Centre for Integrative Biology (CIBIO), University of Trento, 38123 Mattarello, Italy.
Centre for Pharmacology and Toxicology, Hannover Medical School, 30625 Hannover, Germany.
Oncotarget. 2015 Oct 13;6(31):31569-92. doi: 10.18632/oncotarget.5035.
The c-Myc gene codes for a basic-helix-loop-helix-leucine zipper transcription factor protein and is reported to be frequently over-expressed in human cancers. Given that c-Myc plays an essential role in neoplastic transformation we wished to define its activity in lung cancer and therefore studied its targeted expression to respiratory epithelium in a transgenic mouse disease model. Using histological well-defined tumors, transcriptome analysis identified novel c-Myc responsive cell cycle and apoptosis genes that were validated as direct c-Myc targets using EMSA, Western blotting, gene reporter and ChIP assays.Through computational analyses c-Myc cooperating transcription factors emerged for repressed and up-regulated genes in cancer samples, namely Klf7, Gata3, Sox18, p53 and Elf5 and Cebpα, respectively. Conversely, at promoters of genes regulated in transgenic but non-carcinomatous lung tissue enriched binding sites for c-Myc, Hbp1, Hif1 were observed. Bioinformatic analysis of tumor transcriptomic data revealed regulatory gene networks and highlighted mortalin and moesin as master regulators while gene reporter and ChIP assays in the H1299 lung cancer cell line as well as cross-examination of published ChIP-sequence data of 7 human and 2 mouse cell lines provided strong evidence for the identified genes to be c-Myc targets. The clinical significance of findings was established by evaluating expression of orthologous proteins in human lung cancer. Taken collectively, a molecular circuit for c-Myc-dependent cellular transformation was identified and the network analysis broadened the perspective for molecularly targeted therapies.
c-Myc基因编码一种碱性螺旋-环-螺旋-亮氨酸拉链转录因子蛋白，据报道在人类癌症中经常过度表达。鉴于c-Myc在肿瘤转化中起关键作用，我们希望确定其在肺癌中的活性，因此在转基因小鼠疾病模型中研究了其在呼吸道上皮中的靶向表达。利用组织学上明确的肿瘤，转录组分析鉴定出了新的c-Myc反应性细胞周期和凋亡基因，这些基因通过电泳迁移率变动分析（EMSA）、蛋白质免疫印迹法、基因报告和染色质免疫沉淀（ChIP）分析被验证为直接的c-Myc靶标。通过计算分析，在癌症样本中分别出现了与c-Myc协同作用的转录因子，即Klf7、Gata3、Sox18、p53和Elf5以及Cebpα，它们分别与被抑制和上调的基因相关。相反，在转基因但非癌性肺组织中受调控基因的启动子上，观察到了c-Myc、Hbp1、Hif1的富集结合位点。对肿瘤转录组数据的生物信息学分析揭示了调控基因网络，并突出了mortalin和埃兹蛋白作为主要调控因子，而在H1299肺癌细胞系中的基因报告和ChIP分析以及对7个人类和2个小鼠细胞系已发表的ChIP序列数据的交叉检验为所鉴定的基因是c-Myc靶标提供了有力证据。通过评估人类肺癌中直系同源蛋白的表达，确定了研究结果的临床意义。总体而言，确定了一个c-Myc依赖性细胞转化的分子回路，网络分析拓宽了分子靶向治疗的视野。
