Gu Jing, Lv Yang-Fan, Xia Ji-Ying, Bai Fu-Hai, Gong Ji, Pan Guang-Qiang, Liu Bo, Huang Lu, Guo Qiao-Nan, Hao Xiang-Lin
Institute of Toxicology, College of Preventive Medicine, Third Military Medical University, Chongqing, 400038, PR China.
Department of Pathology, Xinqiao Hospital, Third Military Medical University, 183 Xinqiao Street, Shapingba District, Chongqing, 400037, PR China.
Cell Biosci. 2025 Jan 23;15(1):8. doi: 10.1186/s13578-025-01348-3.
Tandem C2 domains, nuclear (TC2N) is a protein that has been characterized to contain C2A domain, C2B domain, and a short C-terminus with a WHXL motif. In previous studies, we have uncovered the oncogenic role and mechanisms of TC2N in lung cancer: TC2N achieves this by inhibiting the p53 signaling pathway and activating the NF-kappaB signaling pathway. Beyond that, its precise function in tumorigenesis is not fully understood.
TC2N-engineered mice model was used to assess the effect of TC2N knockout on normal lung and urethane-induced carcinogenesis. Tumor tissues of 395 lung cancer patients were subjected to tissue microarray and further assessed the associations of TC2N expression with tumor differentiation degree. The protein levels of TC2N and stem cell markers in cell lines and tissue specimens were monitored by WB and immunohistochemistry. In vitro cell assays were performed to assess the effect of TC2N ectopic expression on the stem cell-like characteristics of lung cancer cells. The downstream signaling pathway or target molecule of TC2N was mined using a combination of transcriptomics and proteomics, and the underlying mechanism was explored by WB and co-IP assays.
Herein, TC2N appeared to have a strong effect in promoting lung tumorigenesis caused by urethane, whereas it seemed to lose its function in the normal lung. Meanwhile, we found that the functional differences of TC2N between lung tumor and normal lung were linked to its potential role in cancer cell stemness. Function-wise, TC2N overexpression maintained stem-like properties of lung cancer cell. Mechanism-wise, TC2N upregulated the phosphorylation of EGFR, ERK, STAT3 and FAK1 to activate these signaling pathways by the inhibition of DUSP3 phosphatase via a dual mechanism. Firstly, TC2N competes with EGFR, ERK, STAT3 and FAK1 for binding to DUSP3. This competition prevents these signaling molecules from being dephosphorylated by DUSP3, resulting in their sustained activation. Secondly, TC2N bind to DUSP3 and restrict the enzyme's ability to dephosphorylate the signaling molecules.
Overall, this study revealed a previously unknown role and mechanism of TC2N in the regulation of tumorigenesis and stemness in lung cancer cells.
串联C2结构域，细胞核型（TC2N）是一种蛋白质，其特征在于含有C2A结构域、C2B结构域以及带有WHXL基序的短C末端。在先前的研究中，我们已经揭示了TC2N在肺癌中的致癌作用和机制：TC2N通过抑制p53信号通路和激活NF-κB信号通路来实现这一点。除此之外，其在肿瘤发生中的精确功能尚未完全了解。
使用TC2N基因工程小鼠模型来评估TC2N敲除对正常肺和氨基甲酸乙酯诱导的致癌作用的影响。对395例肺癌患者的肿瘤组织进行组织芯片检测，并进一步评估TC2N表达与肿瘤分化程度的相关性。通过蛋白质免疫印迹法（WB）和免疫组织化学法监测细胞系和组织标本中TC2N和干细胞标志物的蛋白质水平。进行体外细胞实验以评估TC2N异位表达对肺癌细胞干细胞样特性的影响。使用转录组学和蛋白质组学相结合的方法挖掘TC2N的下游信号通路或靶分子，并通过WB和免疫共沉淀实验（co-IP）探索其潜在机制。
在本文中，TC2N似乎在促进氨基甲酸乙酯引起的肺肿瘤发生中具有强大作用，而在正常肺中它似乎失去了其功能。同时，我们发现TC2N在肺肿瘤和正常肺之间的功能差异与其在癌细胞干性中的潜在作用有关。从功能角度来看，TC2N过表达维持了肺癌细胞的干细胞样特性。从机制角度来看，TC2N通过双重机制抑制双特异性磷酸酶3（DUSP3）来上调表皮生长因子受体（EGFR）、细胞外信号调节激酶（ERK）、信号转导和转录激活因子3（STAT3）以及粘着斑激酶1（FAK1）的磷酸化，从而激活这些信号通路。首先，TC2N与EGFR、ERK、STAT3和FAK1竞争与DUSP3的结合。这种竞争阻止了这些信号分子被DUSP3去磷酸化，从而导致它们持续激活。其次，TC2N与DUSP3结合并限制该酶使信号分子去磷酸化的能力。
总体而言，本研究揭示了TC2N在调节肺癌细胞肿瘤发生和干性方面以前未知的作用和机制。
