Geley S, Hartmann B L, Hattmannstorfer R, Löffler M, Ausserlechner M J, Bernhard D, Sgonc R, Strasser-Wozak E M, Ebner M, Auer B, Kofler R
Institute for General and Experimental Pathology, Division of Molecular Pathophysiology, University of Innsbruck, Austria.
Oncogene. 1997 Nov 13;15(20):2429-37. doi: 10.1038/sj.onc.1201399.
The tumor suppressor p53 has been implicated in apoptosis induction and is mutated in human T-ALL CCRF-CEM cells. To investigate possible consequences of wild-type p53 loss, we reconstituted CEM-C7H2, a subclone of CCRF-CEM, with a temperature-sensitive p53 allele (p53ts). Stably transfected lines expressed high levels of p53ts and shift to the permissive temperature (32 degrees C) caused rapid induction of p53-regulated genes, such as p21(CIP1/WAF1), mdm-2 and bax. This was followed by extensive apoptosis within 24 h to 36 h, supporting the notion that mutational p53 inactivation contributed to the malignant phenotype. p53-dependent apoptosis was preceded by digestion of poly(ADP-ribose) polymerase, a typical target of interleukin-1beta-converting enzyme (ICE)-like proteases/caspases, and was markedly resistant to the ICE/caspase-1 and FLICE/caspase-8 inhibitor acetyl-Tyr-Val-Ala-Asp.chloromethylketone (YVAD), but sensitive to the CPP32/caspase-3 inhibitor benzyloxycarbonyl-Asp-Glu-Val-Asp.fluoromethylketone (DEVD) and benzyloxycarbonyl-Val-Ala-Asp.fluoromethylketone (zVAD), a caspase inhibitor with broader specificity. This indicated an essential involvement of caspases, but argued against a significant role of ICE/caspase-1 or FLICE/caspase-8. Actinomycin D or cycloheximide prevented cell death, suggesting that, in this system, p53-induced apoptosis depends upon macromolecule biosynthesis. Introduction of functional p53 into CEM cells enhanced their sensitivity to the DNA-damaging agent doxorubicin, but not to the tubulin-active compound vincristine. Thus, mutational p53 inactivation in ALL might entail relative resistance to DNA-damaging, but not to tubulin-destabilizing, chemotherapy.
肿瘤抑制因子p53与细胞凋亡诱导有关，且在人T细胞急性淋巴细胞白血病（T-ALL）CCRF-CEM细胞中发生突变。为了研究野生型p53缺失可能产生的后果，我们用一个温度敏感型p53等位基因（p53ts）对CCRF-CEM的一个亚克隆CEM-C7H2进行了重建。稳定转染的细胞系表达高水平的p53ts，转移到允许温度（32摄氏度）会导致p53调控基因如p21（CIP1/WAF1）、mdm-2和bax的快速诱导。随后在24小时至36小时内发生广泛的细胞凋亡，支持了突变型p53失活导致恶性表型的观点。p53依赖的细胞凋亡之前会发生聚（ADP-核糖）聚合酶的切割，聚（ADP-核糖）聚合酶是白细胞介素-1β转换酶（ICE）样蛋白酶/半胱天冬酶的典型作用靶点，并且对ICE/半胱天冬酶-1和FLICE/半胱天冬酶-8抑制剂乙酰-Tyr-Val-Ala-Asp.氯甲基酮（YVAD）具有明显抗性，但对CPP32/半胱天冬酶-3抑制剂苄氧羰基-Asp-Glu-Val-Asp.氟甲基酮（DEVD）和苄氧羰基-Val-Ala-Asp.氟甲基酮（zVAD，一种具有更广泛特异性的半胱天冬酶抑制剂）敏感。这表明半胱天冬酶起着重要作用，但反对ICE/半胱天冬酶-1或FLICE/半胱天冬酶-8起显著作用。放线菌素D或放线菌酮可防止细胞死亡，表明在该系统中，p53诱导的细胞凋亡依赖于大分子生物合成。将功能性p53导入CEM细胞可增强其对DNA损伤剂阿霉素的敏感性，但对微管活性化合物长春新碱不敏感。因此，ALL中突变型p53失活可能导致对DNA损伤化疗药物的相对抗性，但对微管破坏化疗药物不产生抗性。
