Securin與p53調控蛋白酶體抑制劑所誘發之大腸癌細胞的凋亡

Abstract

蛋白酶體抑制藥物例如MG132及 bortezomib已經被證實具有抗癌的活性。然而，由蛋白酶體抑制劑所調控的細胞凋亡之機制仍不清楚。在本論文中，我們發現securin與p53參與調控MG132及bortezomib所誘發之人類大腸癌細胞的凋亡。MG132及bortezomib抑制大腸癌細胞中百分之五十蛋白酶體活性的濃度，分別為1.8 μM 和 43.5 nM。處理MG132及bortezomib都會增加細胞週期G2/M的分佈，減少細胞的存活率，以及誘發細胞凋亡。有趣地是蛋白酶體抑制劑會顯著誘引securin及p53蛋白的增加。處理蛋白酶體抑制劑後，在securin功能正常的大腸癌細胞會比喪失securin基因的大腸癌細胞之死亡率更高。並且蛋白酶體抑制劑在p53功能正常的大腸癌細胞比p53基因喪失或突變的大腸癌細胞，對細胞凋亡的誘發更為敏感。將大腸癌細胞中securin 的基因表現阻斷，會降低MG132所誘發的p53蛋白表現、caspase 3活化及PARP的切割。此外利用免疫沉澱分析法，發現處理蛋白酶體抑制劑會增加securin與p53之間的蛋白結合。由以上結果，我們推測人類大腸癌細胞的securin及p53的表現與蛋白交互作用，參與蛋白酶體抑制劑所誘發的細胞凋亡。

Proteasome inhibitors such as MG132 and bortezomib have been shown to exert anticancer activities; however, the mechanisms of apoptotic regulation by proteasome inhibitors remain unclear. Here, we show that securin and p53 participate in regulating apoptosis by treatment with MG132 and bortezomib in the human colon cancer. The IC50 values of inhibiting proteasome activities in colon cancer cells were 1.8 μM and 43.5 nM by treatment with MG132 and bortezomib, respectively. Both MG132 and bortezomib increased G2/M fractions, decreased cell viability, and induced apoptosis. Interestingly, securin and p53 proteins were markedly elicited by proteasome inhibitors. The securin-wild type colon cancer cells were higher on the cell death than the securin-null cells by treatment with proteasome inhibitors. Moreover, the p53-wild type colon cancer cells were more sensitive on induction of apoptosis than the p53-null and p53-mutational cells. The knockdown of securin expression reduced the p53 protein level, caspase 3 activation and poly (ADP-ribose) polymerase (PARP) cleavage in the MG132-treated colon cancer cells. Besides, treatment with MG132 increased the protein interaction of securin and p53 by immunoprecipitation assays. These findings suggest that the increase and interaction of securin and p53 may mediate apoptosis by treatment with proteasome inhibitors in the human colon cancer cells.