Cancer Sciences Unit, Southampton University Faculty of Medicine, General Hospital, Southampton, UK.
Cell Cycle. 2013 Feb 1;12(3):430-41. doi: 10.4161/cc.23285. Epub 2012 Feb 1.
Recent studies have highlighted an apparently paradoxical link between self-renewal and senescence triggered by DNA damage in certain cell types. In addition, the finding that TP53 can suppress senescence has caused a re-evaluation of its functional role in regulating these outcomes. To investigate these phenomena and their relationship to pluripotency and senescence, we examined the response of the TP53-competent embryonal carcinoma (EC) cell line PA-1 to etoposide-induced DNA damage. Nuclear POU5F1/OCT4A and P21CIP1 were upregulated in the same cells following etoposide-induced G 2M arrest. However, while accumulating in the karyosol, the amount of OCT4A was reduced in the chromatin fraction. Phosphorylated CHK2 and RAD51/γH2AX-positive nuclear foci, overexpression of AURORA B kinase and moderate macroautophagy were evident. Upon release from G 2M arrest, cells with repaired DNA entered mitoses, while the cells with persisting DNA damage remained at this checkpoint or underwent mitotic slippage and gradually senesced. Reduction of TP53 using sh- or si-RNA prevented the upregulation of OCT4A and P21CIP1 and increased DNA damage. Subsequently, mitoses, micronucleation and senescence were all enhanced after TP53 reduction with senescence confirmed by upregulation of CDKN2A/P16INK4A and increased sa-β-galactosidase positivity. Those mitoses enhanced by TP53 silencing were shown to be multicentrosomal and multi-polar, containing fragmented and highly deranged chromosomes, indicating a loss of genome integrity. Together, these data suggest that TP53-dependent coupling of self-renewal and senescence pathways through the DNA damage checkpoint provides a mechanism for how embryonal stem cell-like EC cells safeguard DNA integrity, genome stability and ultimately the fidelity of self-renewal.
最近的研究强调了在某些细胞类型中，DNA 损伤引发的自我更新和衰老之间存在一种明显的悖论关系。此外，TP53 可以抑制衰老的发现，促使人们重新评估其在调节这些结果中的功能作用。为了研究这些现象及其与多能性和衰老的关系，我们检查了具有 TP53 功能的胚胎癌细胞系 PA-1 对依托泊苷诱导的 DNA 损伤的反应。在依托泊苷诱导的 G2M 期阻滞后，核内 POU5F1/OCT4A 和 P21CIP1 上调。然而，虽然 OCT4A 在核质中积累，但在染色质部分的含量减少。磷酸化的 CHK2 和 RAD51/γH2AX 阳性核斑、AURORA B 激酶的过表达和适度的巨自噬是明显的。从 G2M 期阻滞中释放后，修复 DNA 的细胞进入有丝分裂，而持续存在 DNA 损伤的细胞则停留在这个检查点，或发生有丝分裂滑步，逐渐衰老。使用 sh-或 si-RNA 减少 TP53 的表达可防止 OCT4A 和 P21CIP1 的上调，并增加 DNA 损伤。随后，随着 TP53 减少导致的衰老的确认，即通过 CDKN2A/P16INK4A 的上调和增加的 sa-β-半乳糖苷酶阳性率，有丝分裂、微核形成和衰老都增强。用 TP53 沉默增强的有丝分裂显示为多中心体和多极的，包含断裂和高度紊乱的染色体，表明基因组完整性的丧失。总的来说，这些数据表明，TP53 依赖的自我更新和衰老途径通过 DNA 损伤检查点的耦合，为胚胎干细胞样 EC 细胞如何保护 DNA 完整性、基因组稳定性和最终自我更新的保真度提供了一种机制。
