Miyaji E N, Menck C F
Departamento de Biologia, Universidade de São Paulo, Brazil.
Photochem Photobiol. 1998 Nov;68(5):719-24.
We have introduced the human bcl-2 gene under the control of the human metallothionein MTIIA promoter into the rat kangaroo PtK2 cell line. Two independent clones were obtained in which the levels of Bcl-2 protein expression can be controlled by the addition of metals in the culture medium. These cell lines were employed to investigate the effects of this protein in UV-induced apoptosis. Overexpression of Bcl-2 in PtK2 cells resulted in a delay in the appearance of apoptosis markers, such as chromatin condensation and internucleosomal DNA fragmentation. However, colony survival after UV was not affected, suggesting that Bcl-2 did not impose a definitive block for cell death. The elimination of cyclobutane pyrimidine dimers through photoreactivation 24 h after irradiation in cells overexpressing Bcl-2 did not affect apoptosis. This indicates that irreversible events in the signaling pathway of apoptosis occur in the period between irradiation and photoreactivation even in the presence of high levels of Bcl-2 protein can delay the onset of UV-induced apoptosis in these marsupial cells, early events triggered by the pyrimidine dimers, upstream from the Bcl-2 action, lead the cell to a state committed to die.
我们已将受人类金属硫蛋白MTIIA启动子控制的人类bcl-2基因导入大鼠袋鼠PtK2细胞系。获得了两个独立的克隆，其中Bcl-2蛋白表达水平可通过在培养基中添加金属来控制。利用这些细胞系研究该蛋白在紫外线诱导的细胞凋亡中的作用。PtK2细胞中Bcl-2的过表达导致凋亡标志物（如染色质浓缩和核小体间DNA片段化）出现延迟。然而，紫外线照射后的集落存活率未受影响，这表明Bcl-2并未对细胞死亡形成决定性的阻滞。在过表达Bcl-2的细胞中，照射后24小时通过光复活消除环丁烷嘧啶二聚体并不影响细胞凋亡。这表明，即使存在高水平的Bcl-2蛋白，凋亡信号通路中的不可逆事件在照射和光复活之间的时间段内仍会发生，Bcl-2可延迟这些有袋类细胞中紫外线诱导的细胞凋亡的起始，嘧啶二聚体引发的早期事件在Bcl-2作用的上游，使细胞进入注定死亡的状态。
