Jarvis W D, Johnson C R, Fornari F A, Park J S, Dent P, Grant S
Department of Medicine, Medical College of Virginia, Richmond, Virginia, USA.
J Pharmacol Exp Ther. 1999 Sep;290(3):1384-92.
We recently demonstrated that physiological induction of apoptosis by cytotoxic sphingolipid messengers proceeds via activating protein-1 (AP1)-dependent and AP1-independent mechanisms in U937 human monoblastic leukemia cells. Here we examine involvement of the stress-activated protein kinase (SAPK) cascade and AP1 in the initiation of apoptosis in U937 cells by podophyllotoxin-derived inhibitors of topoisomerase II. Induction of apoptotic cell death and DNA damage by treatment of U937 cells with etoposide (100 microM) was associated with phosphorylation and activation of the c-Jun NH(2)-terminal kinase (JNK1) SAPK enzymes p46 and p54-JNK2 and transient increases in expression of the transcription factor c-Jun, a primary JNK substrate. These responses were accompanied by a modest, but sustained, recruitment of the mitogen-activated protein kinases p42-extracellular signal receptor-activated kinase (ERK)1 and p44-extracellular signal receptor-activated kinase 2. The capacity of etoposide to promote double-stranded DNA degradation and cell death was unaffected by manipulations that interfere with SAPK signaling outflow through c-Jun/AP1, including: 1) pharmacological inhibition of AP1 activity by diferuloylmethane and 2) molecular ablation of normal c-Jun function by the Jun dominant-negative mutant TAM-67. Cytotoxicity of the structurally related compound teniposide was similarly unaffected. In parallel trials, the lethal actions of ceramide (but not of sphingosine) were markedly diminished by pretreatment with diferuloylmethane or expression of TAM-67, confirming the effectiveness of these interventions in suppression of SAPK/AP1-dependent apoptosis. The involvement of AP1 in the proapoptotic actions of other inhibitors of topoisomerase II activity was also evaluated. Induction of cell death by the anthracyclines daunorubicin, daunorubicin, and idarubicin was found to be insensitive to pretreatment with diferuloylmethane or expression of TAM-67. Collectively, the present data indicate that induction of apoptosis by etoposide and related inhibitors of topoisomerase II is mediated through a cell death pathway that does not require SAPK-dependent recruitment of AP1. These findings additionally suggest that activation of the SAPK represents a consequence, rather than an underlying cause, of etoposide-induced apoptosis in myeloid leukemia cells.
我们最近证明，在U937人单核细胞白血病细胞中，细胞毒性鞘脂信使诱导的细胞凋亡通过激活蛋白-1（AP1）依赖性和非AP1依赖性机制进行。在此，我们研究了应激激活蛋白激酶（SAPK）级联和AP1在鬼臼毒素衍生的拓扑异构酶II抑制剂诱导U937细胞凋亡起始过程中的作用。用依托泊苷（100 microM）处理U937细胞诱导凋亡性细胞死亡和DNA损伤，与c-Jun NH(2)-末端激酶（JNK1）SAPK酶p46和p54-JNK2的磷酸化和激活以及转录因子c-Jun（一种主要的JNK底物）表达的短暂增加相关。这些反应伴随着丝裂原活化蛋白激酶p42-细胞外信号受体激活激酶（ERK）1和p44-细胞外信号受体激活激酶2适度但持续的募集。依托泊苷促进双链DNA降解和细胞死亡的能力不受干扰通过c-Jun/AP1的SAPK信号流出的操作的影响，这些操作包括：1）二阿魏酰甲烷对AP1活性的药理学抑制和2）Jun显性负突变体TAM-67对正常c-Jun功能的分子消除。结构相关化合物替尼泊苷的细胞毒性同样不受影响。在平行试验中，用二阿魏酰甲烷预处理或表达TAM-67可显著降低神经酰胺（而非鞘氨醇）的致死作用，证实了这些干预措施在抑制SAPK/AP1依赖性细胞凋亡中的有效性。还评估了AP1在其他拓扑异构酶II活性抑制剂促凋亡作用中的参与情况。发现柔红霉素、阿霉素和伊达比星等蒽环类药物诱导的细胞死亡对用二阿魏酰甲烷预处理或表达TAM-67不敏感。总体而言，目前的数据表明，依托泊苷和相关拓扑异构酶II抑制剂诱导的细胞凋亡是通过一条不需要SAPK依赖性募集AP1的细胞死亡途径介导的。这些发现还表明，SAPK的激活是髓系白血病细胞中依托泊苷诱导的细胞凋亡的结果，而非根本原因。
