Forced expression of the H11 heat shock protein can be regulated by DNA methylation and trigger apoptosis in human cells
Forced expression of the H11 heat shock protein can be regulated by DNA methylation and trigger apoptosis in human cells
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DOI：
10.1074/jbc.m303834200
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发表时间：
2003-09-26
影响因子：
4.8
通讯作者：
Aurelian, L
中科院分区：
文献类型：
--
作者：
Gober, MD;Smith, CC;Aurelian, L
H11是单纯疱疹病毒蛋白的真核同源物，具有热休克蛋白（Hsp）的晶体蛋白基序，但它与经典家族成员的不同之处在于，相对于正常对应物，在各种肿瘤组织和细胞系（即黑素瘤、前列腺癌和肉瘤）中，mRNA和蛋白水平降低。在这些细胞中，表达不能通过热休克恢复，而是通过去甲基化剂5-氮杂-2 '-脱氧胞苷（Aza-C）恢复。通过Aza-C处理强制H11表达，用H11表达载体瞬时转染，或在四环素敏感性启动子控制下逆转录病毒介导的H11递送触发细胞凋亡。这通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）和半胱天冬酶-3和p38 MAPK的活化的阳性细胞百分比的显著增加（p < 0.001）以及通过TUNEL+细胞核与增加的H11水平的共定位来证明。pancaspase抑制剂苄氧基羰基-Val-Ala-Asp-氟甲基酮或p38 MAPK抑制剂SB 203580可部分抑制细胞凋亡。它被废除的共同治疗与两种抑制剂，表明H11触发的细胞凋亡是半胱天冬酶和p38 MAPK依赖性。单位点突变体（H11-W51 C）具有与MEK/ERK激活相关的细胞保护活性，并且它阻断了H11诱导的共转染和Aza-C处理的细胞凋亡，表明它是显性负突变体。这是第一个报告的热休克蛋白具有促凋亡活性。
H11, the eukaryotic homologue of a herpes simplex virus protein, has the crystallin motif of heat shock proteins (Hsp), but it differs from canonical family members in that mRNA and protein levels were reduced in various tumor tissues and cell lines ( viz. melanoma, prostate cancer and sarcoma) relative to their normal counterparts. In these cells, expression was not restored by heat shock, but rather by the demethylating agent 5-aza-2'-deoxycytidine (Aza-C). Forced H11 expression by Aza-C treatment, transient transfection with H11 expression vectors, or retrovirus-mediated delivery of H11 under the control of a tetracycline-sensitive promoter triggered apoptosis. This is evidenced by a significant ( p < 0.001) increase in the percentage of cells positive for terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling ( TUNEL) and for activation of caspase-3 and p38MAPK and by the co-localization of TUNEL+ nuclei with increased H11 levels. Apoptosis was partially inhibited by the pancaspase inhibitor benzyloxycarbonyl-Val- Ala-Asp-fluoromethyl ketone or the p38MAPK inhibitor SB203580. It was abrogated by co-treatment with both inhibitors, suggesting that H11-triggered apoptosis is both caspase- and p38MAPK-dependent. A single site mutant (H11-W51C) had cytoprotective activity related to MEK/ERK activation, and it blocked H11-induced apoptosis in co-transfected and Aza-C-treated cells, indicating that it is a dominant negative mutant. This is the first report of a heat shock protein with proapoptotic activity.