1p36.22MIIP基因缺失通过影响TOP2A解连接功能促进结直肠癌进展的机制研究

We found chromosome 1p36.22MIIP deletion was strongly linked to colorectal cancer (CRC) progression, but the mechanism is unknown. In the preliminary study, we found that sister chromosome mis-segregation which was the characteristic phenotype of chromosome instability (CIN) occurred in CRC cells with MIIP deletion and the decatenation ability of topoisomerase IIα (TOP2A) which regulated sister chromosome segregation was decreased due to MIIP deletion. Furthermore, our co-IP assay showed MIIP could bind to TOP2A. Therefore, we hypothesize that 1p36.22MIIP deletion may contribute to CRC progression by affecting TOP2A decatenation ability. To prove this hypothesis, we will: 1) further validate the relationship of MIIP deletion and CIN in CRC and the role of MIIP deletion in CRC progression; 2) study how MIIP regulates the decatenation function of TOP2A, discover the accurate binding site of MIIP and TOP2A, study molecular mechanism of MIIP inducing CIN and finnally explain how 1p36.22MIIP deletion contributes to CRC progression.

我们发现位于染色体1p36.22的MIIP基因缺失与结直肠癌进展密切相关，但机制尚未阐明。预实验结果显示MIIP基因缺失的肠癌细胞中出现有丝分裂期姊妹染色体分离显著异常这一染色体不稳定性的特征表型，进一步发现调控这一过程的关键酶TOP2A与MIIP相互结合且解连接能力显著下降，提示MIIP基因缺失可能与TOP2A介导的结直肠癌染色体不稳定性相关。为此，我们提出假说：1p36.22MIIP基因缺失通过影响TOP2A的解连接功能及其介导的染色体不稳定性促进结直肠癌的进展。为了验证这一假说，本项目拟：1）通过体内外实验，进一步验证MIIP基因缺失与结直肠癌染色体不稳定的相关性及其在肿瘤进展中的作用；2）通过分子实验，确定MIIP对TOP2A的调控方式，并解析MIIP与TOP2A相互作用的结构基础，最终阐明1p36.22MIIP基因缺失通过影响TOP2A的解连接功能促进结直肠癌进展的机制。

迁移侵袭抑制蛋白（migration and invasion inhibitory protein, MIIP）大小为45kDa，最初因其可以抑制胶质瘤细胞迁移和侵袭故而得名。我们发现MIIP蛋白可以通过与TOP2A结合，并影响TOP2A这一解连接酶的稳定性从而影响肠癌细胞有丝分裂过程，导致肿瘤细胞染色体不稳定和结直肠癌进展。成果已经发表于Journal of Pathology 杂志（J Path. 2017;241(1):67-79.）。我们深入研究了MIIP作为重要分子参与了表皮生长因子（EGF）诱导的NF-κB信号通路的激活。并通过一系列实验数据证明MIIP S303磷酸化与结直肠癌转移密切相关，PKCε磷酸化MIIP可以保护RelA不被HDAC6去乙酰化，从而增强EGF刺激的RelA转录活性和结肠肿瘤侵袭转移。在EGFR信号通路激活的条件下，PKCε磷酸化MIIP的保守位点Ser303，促使MIIP与NF-κΒ家族RelA结合，保护RelA不被HDAC6去乙酰化，从而增加下游基因（如Twist、MMP2）表达，最终促进结直肠癌转移。这一成果揭示了MIIP调控肿瘤转移的新机制，已发表于Nature Communications杂志（Nat Commun. 2017;8(1):939. ）此外，项目负责人专注于结直肠肿瘤的转移（特别是早期转移）这一科学问题和临床难点，在该项国家自然科学基金的资助下，开展了相关基础和临床研究，结果发表于Clinical Gastroenterology and Hepatology、Cancer Communications等杂志。