去泛素化酶USP16调节细胞衰老及恶性转化的研究

Cellular senescence, a state of irreversible growth arrest that limits excessive or aberrant cell proliferation, may protect against tumor development. Emerging evidence indicates that dysfunction in ubiquitin-dependent processes of proteins is involved in cell malignant transformation. Our preliminary experiments revealed that knockdown of deubiquitinase USP16 inhibited replicative senescence and oncogene H-Ras-induced senescence in primary fibroblast cell line IMR-90. Overexpression of USP16 in human liver cancer cells triggered cellular senescence. Moreover, we found that the expression of USP16 was downregulated in clinic samples of liver cancer compared with non-tumor tissues, and the low expression of USP16 was also found in clinic samples of lung cancer with lymph node metastasis compared with those without lymph node metastasis. Base on these results, using USP16 conditional knockout mice combined with animal models of primary tumor as well as human tumor cell lines, we will pursue the following specific aims: (1) to illustrate the role of USP16 in the process of cellular malignancy and tumor formation, (2) to identify the potential deubiquinated target and the mechanism of deubiquination in USP16-induced senescence, and (3) to investigate the relationship between USP16-mediated deubiquitination and senescence-related signaling pathway, such as p53-p21, p16-Rb. Thus, the study will be valuable for indentifying the deubiquitinases that function critically in cellular senescence and malignant transformation.

细胞衰老是维护组织稳态、抑制细胞恶性转化和预防肿瘤发生的重要细胞学机制。近来研究表明，蛋白泛素化的调控异常在细胞恶性转化过程中起重要作用。我们前期的工作显示，敲降去泛素化酶USP16可抑制细胞的复制衰老及H-Ras诱导的细胞衰老；在肝癌细胞中，过表达USP16可促进细胞衰老。通过对临床样本的分析表明，在肝癌组织中USP16呈低表达，同时在发生淋巴结转移的肺癌组织中也低表达USP16。提示USP16可能在细胞逃逸癌基因诱导的细胞衰老及恶性转化过程中起作用。本项目将在此基础上，应用USP16敲除鼠、小鼠原发瘤模型以及人肿瘤系统，进一步明确USP16在抑制细胞恶性转化、促进细胞衰老中的作用，阐明USP16对细胞衰老相关信号通路p53-p21、 p16-Rb的影响，确定USP16去泛素化底物及其调控细胞衰老的分子机制。本研究将为阐明控制细胞衰老和肿瘤发生的重要去泛素化事件，提供新的认识和实验依据。

近来的研究表明，蛋白泛素化的调控异常与肿瘤的发生发展密切相关。其中，去泛素化酶USP16可调节多种生物学过程，具有影响胚胎干细胞分化、造血干细胞生成等作用。但是，USP16在肿瘤发生、发展中的作用仍不十分清楚。.在本项目支持下，我们对USP16在肿瘤发生、发展中的作用及分子机制进行了研究。结果显示，敲降USP16虽然可以延迟细胞衰老的产生，但最终对肿瘤细胞的生长起抑制作用。体内实验也证明，敲除USP16降低SV40T和K-RasG12D共转化的MEF细胞的成瘤能力，而且USP16条件性敲除可抑制K-RasG12D诱导的鼠肺肿瘤的生长，延长小鼠生存期。机制研究显示，USP16对肿瘤细胞生长的影响不依赖于细胞衰老相关信号通路——p53信号通路，而敲降USP16可以提高p38的磷酸化水平。进一步的分子机制研究证明，USP16对肿瘤细胞生长的影响依赖于其去泛素化酶活性，而不依赖于其磷酸化。.总之，敲降USP16可抑制肿瘤细胞的生长，从而抑制K-Ras突变驱动的肺肿瘤的发生发展，延长生存期，提示USP16可作为潜在的肺肿瘤治疗靶点。因此，本项目的研究成果具有一定的临床应用价值。