The Werner syndrome protein suppresses telomeric instability caused by chromium (VI) induced DNA replication stress.
The Werner syndrome protein suppresses telomeric instability caused by chromium (VI) induced DNA replication stress.
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DOI：
10.1371/journal.pone.0011152
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发表时间：
2010-06-16
期刊：
影响因子：
3.7
通讯作者：
Opresko PL
中科院分区：
文献类型：
--
作者：
Liu FJ;Barchowsky A;Opresko PL
端粒保护染色体末端，由被特殊蛋白质包裹的富含鸟嘌呤的重复序列组成。极短的端粒与疾病、衰老和癌症有关。端粒复制缺陷可导致端粒丢失，端粒丢失可通过端粒酶介导的端粒延长或Werner综合征解旋酶/外切酶蛋白（WRN）的活性来预防。端粒酶和WRN都能减弱环境致癌物六价铬（Cr(VI)）诱导的细胞毒性，从而促进复制应激和DNA聚合酶阻滞。然而，尚不清楚Cr（VI）诱导的复制胁迫是否会影响端粒完整性。在这里，我们报告了Cr（VI）暴露于人成纤维细胞诱导端粒损伤，通过端粒病灶磷酸化的H2AX （γH2AX）表明。诱导的γ - h2ax灶发生在s期细胞中，表明复制分叉停止或崩溃。中期染色体的端粒荧光原位杂交（FISH）显示，Cr（VI）暴露导致端粒丢失和姐妹染色单体融合增加，这些染色体被端粒酶活性拯救。在Cr（VI）暴露后恢复过程中，通过γ - h2ax聚焦显示，缺乏WRN蛋白的人类细胞表现出端粒和非端粒损伤的延迟减少，这与WRN在修复受损复制叉中的作用一致。染色体扩散的端粒FISH结果显示，WRN对Cr（VI）诱导的端粒丢失和下游染色体融合具有保护作用，但不阻止染色体融合，保留端粒序列在融合点。我们的研究表明，环境诱导的复制胁迫会导致端粒丢失和畸变，而端粒酶介导的端粒延伸或复制叉恢复中的WRN功能会抑制端粒丢失和畸变。
Telomeres protect the chromosome ends and consist of guanine-rich repeats coated by specialized proteins. Critically short telomeres are associated with disease, aging and cancer. Defects in telomere replication can lead to telomere loss, which can be prevented by telomerase-mediated telomere elongation or activities of the Werner syndrome helicase/exonuclease protein (WRN). Both telomerase and WRN attenuate cytotoxicity induced by the environmental carcinogen hexavalent chromium (Cr(VI)), which promotes replication stress and DNA polymerase arrest. However, it is not known whether Cr(VI)-induced replication stress impacts telomere integrity. Here we report that Cr(VI) exposure of human fibroblasts induced telomeric damage as indicated by phosphorylated H2AX (γH2AX) at telomeric foci. The induced γH2AX foci occurred in S-phase cells, which is indicative of replication fork stalling or collapse. Telomere fluorescence in situ hybridization (FISH) of metaphase chromosomes revealed that Cr(VI) exposure induced an increase in telomere loss and sister chromatid fusions that were rescued by telomerase activity. Human cells depleted for WRN protein exhibited a delayed reduction in telomeric and non-telomeric damage, indicated by γH2AX foci, during recovery from Cr(VI) exposure, consistent with WRN roles in repairing damaged replication forks. Telomere FISH of chromosome spreads revealed that WRN protects against Cr(VI)-induced telomere loss and downstream chromosome fusions, but does not prevent chromosome fusions that retain telomere sequence at the fusion point. Our studies indicate that environmentally induced replication stress leads to telomere loss and aberrations that are suppressed by telomerase-mediated telomere elongation or WRN functions in replication fork restoration.