GTF2I mutation induces metabolic alterations and cell transformation in thymic epithelial cells [I]. GTF2I mutation induces metabolic alterations and cell transformation in thymic epithelial cells [I]

The molecular mechanisms underlying the pathogenesis of TETs are poorly understood. Recently we reported a common missense mutation on GTF2I gene in thymic tumors and hypothesized that GTF2I mutation might contribute to thymic tumorigenesis. Expression of mutant TFII-I altered the transcriptome of thymic epithelial cells and up-regulated several oncogenic genes. Using CRISPR/Cas9n, Gtf2i T1211A knock-in cells exhibited distinct features of cancerous cells including cell transformation, aneuploidy, tumor growth in xenograft, and survival after DNA damage or glucose deprivation. We also observed that Gtf2i mutation increased the expression of several glycolytic enzymes, cyclooxygenase-2, and altered lipid metabolism. Elevated COX-2 expression by Gtf2i mutation was required for survival under metabolic stress and cellular transformation of thymic epithelial cells. Our findings identify GTF2I mutation as a new oncogenic driver mutation that is responsible for transformation of thymic epithelial cells. Overall design: We profiled transcriptome alteration by mutant (beta or delta) TFII-I expression under doxycycline-inducilble system in immortalized mouse thymic epithelial cell line (TEC100.4). Total RNA was prepared using the RNeasy Plus Mini Kit (QIAGEN), and microarray anlysis was performed using the Agilent SurePrint G3 mouse gene expression V2 8×60K microarray kit (design ID G4852B, Single color) according to the manufacturer's instructions. Data analysis was performed using DAVID functional annotation analysis.

胸腺上皮肿瘤（Thymic Epithelial Tumors, TETs）的发病分子机制目前仍未得到充分阐释。本团队此前曾报道胸腺肿瘤中存在GTF2I基因的常见错义突变，并提出假说：GTF2I突变可能参与胸腺肿瘤发生过程。突变型TFII-I的表达可改变胸腺上皮细胞的转录组，并上调多个致癌基因的表达。
采用CRISPR/Cas9n技术构建的Gtf2i T1211A敲入细胞，展现出癌细胞的多种典型特征：包括细胞转化、非整倍体、异种移植瘤生成，以及在DNA损伤或葡萄糖剥夺条件下的存活能力。本团队还观察到，Gtf2i突变可提升多种糖酵解酶及环氧合酶-2（cyclooxygenase-2, COX-2）的表达水平，并改变脂质代谢过程。Gtf2i突变所介导的COX-2表达上调，是胸腺上皮细胞在代谢应激下存活以及发生细胞转化的必要条件。
本研究结果证实，GTF2I突变是一种全新的致癌驱动突变，可直接介导胸腺上皮细胞的转化过程。
实验设计：本研究以永生化小鼠胸腺上皮细胞系（TEC100.4）为模型，采用多西环素诱导表达系统，分析突变型（β或δ型）TFII-I表达所引发的转录组变化。总RNA提取采用RNeasy Plus Mini Kit（QIAGEN公司），基因芯片检测使用Agilent SurePrint G3小鼠基因表达V2 8×60K芯片试剂盒（设计编号G4852B，单通道），实验操作严格遵循厂商说明书进行。数据分析采用DAVID功能注释分析工具完成。