Department of Surgery, James Graham Brown Cancer Center, Louisville, KY, USA.
Cancer Biol Ther. 2012 Sep;13(11):1091-101. doi: 10.4161/cbt.21143. Epub 2012 Jul 24.
The transcription factor E2F-1 plays a crucial role in the control of cell proliferation. E2F-1 has tumor suppressive properties by inducing apoptosis and autophagy. In this study, E2F-1 and its truncated form (E2Ftr), lacking the transactivation domain (TAD), were compared for their ability to induce autophagy. In Gaussia luciferase-based assays, both E2F-1 and E2Ftr induced the proteolytic cleavage of the autophagic marker LC3. In addition, LC3 and autophagy protein 5 (Atg5) were upregulated by E2F-1 and E2Ftr. Likewise, both E2F proteins induced a punctate pattern of GFP-tagged LC3, indicating autophagosome formation. The presence of double-membrane autophagic vesicles induced by E2F-1 and E2Ftr was confirmed by transmission electron microscopy (TEM). The application of z-VAD-fmk, a caspase inhibitor, partially blocked both E2F-1 and E2Ftr-mediated cytotoxicity. Moreover, Atg5 (-/-) cells were more resistant to the E2F-1 or E2Ftr-induced cell killing effect than Atg5 wt cells. The TAD of E2F-1 is not essential for induction of autophagy; apoptosis and autophagy cooperate for an efficient cancer cell killing effect induced by E2F-1 or E2Ftr. E2Ftr-induced autophagy is a promising approach to destroy tumors that are resistant to conventional treatments.
转录因子 E2F-1 在控制细胞增殖中起着至关重要的作用。E2F-1 通过诱导细胞凋亡和自噬发挥肿瘤抑制作用。在这项研究中，比较了 E2F-1 及其截断形式（E2Ftr），后者缺乏转录激活结构域（TAD），以比较它们诱导自噬的能力。在基于 Gaussia 荧光素酶的测定中，E2F-1 和 E2Ftr 均诱导自噬标志物 LC3 的蛋白水解裂解。此外，E2F-1 和 E2Ftr 上调 LC3 和自噬蛋白 5（Atg5）。同样，两种 E2F 蛋白均诱导 GFP 标记的 LC3 形成点状模式，表明自噬体形成。透射电子显微镜（TEM）证实了由 E2F-1 和 E2Ftr 诱导的双层自噬囊泡的存在。应用半胱氨酸天冬氨酸蛋白酶抑制剂 z-VAD-fmk 部分阻断了 E2F-1 和 E2Ftr 介导的细胞毒性。此外，与 Atg5 wt 细胞相比，Atg5（-/-）细胞对 E2F-1 或 E2Ftr 诱导的细胞杀伤作用更具抗性。E2F-1 的 TAD 对于诱导自噬并非必需；凋亡和自噬协同作用，可有效诱导 E2F-1 或 E2Ftr 诱导的癌细胞杀伤作用。E2Ftr 诱导的自噬是破坏对传统治疗方法具有抗性的肿瘤的一种很有前途的方法。
