Ionizing radiation (IR) confers a survival advantage in tongue squamous cell carcinoma (TSCC), however, IR resistance limits its efficacy. Although Yin Yang 1 (YY1) has been reported to play a role in genotoxic drug resistance by accelerating DNA repair, its role in TSCC radioresistance remains unclear. In this study, we examined YY1 mRNA and protein expression in human tongue cancer samples using qRT-PCR and western blotting, respectively. DNA array data identified YY1 mRNA expression in IR sensitivity or resistance cell lines and tissues. Tongue carcinoma primary cells and CAL27 cells with YY1 stably overexpressed or knocked-down were exposed to IR and evaluated for cell proliferation and apoptosis by CCK8-assay and caspase-3 assay, respectively. We also examined DNA damage- or repair-related indicators, such as YY1, p-H2AX, nuclear PTEN, p-PTEN, and Rad51 through Western blot analysis. Additionally, we explored the mechanism of IR-induced PTEN nuclear translocation by introducing a series of PTEN phosphorylation site mutations and co-IP assay. We observed that YY1 mRNA and protein are highly expressed in TSCC tissues, which was correlated with worse overall survival. Moreover, higher expression of YY1 and Rad51 was observed in radioresistant cells and tissues, overexpression of YY1 led to IR resistance in TSCC cells, whereas YY1 knockdown sensitized TSCC cells to IR. The underlying mechanism showed that the overexpression of YY1 upregulated nuclear PTEN and Rad51 expression, which is essential for DNA repair. IR upregulated YY1, nuclear PTEN, and Rad51; thus, knockdown of YY1 completely blocked IR-induced upregulation of nuclear PTEN/Rad51. IR upregulated PTEN phosphorylation, and mutation of the phosphorylation site of Ser380 nearly completely blocked IR-induced PTEN nuclear translocation. Furthermore, the phosphatase PP2A negatively regulated pS380-PTEN, and knockdown of YY1 completely blocked IR-induced pS380-PTEN through PP2A. In conclusion, knockdown of YY1 enhanced TSCC radiosensitivity through PP2A-mediated dephosphorylation of PTEN Ser380; thus, antagonizing the IR-induced nuclear PTEN/Rad51 axis and targeting YY1 may reverse IR resistance in TSCC.

中文翻译：

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YY1介导的PTEN去磷酸化可拮抗IR诱导的DNA修复，从而促进舌鳞状细胞癌的放射增敏作用。

电离辐射（IR）在舌鳞状细胞癌（TSCC）中具有生存优势，但是，IR耐药性限制了其疗效。尽管据报道，Yin Yang 1（YY1）通过加速DNA修复在遗传毒性药物抗性中起作用，但其在TSCC放射抗性中的作用仍不清楚。在这项研究中，我们分别使用qRT-PCR和Western blotting检测了人舌癌样品中YY1 mRNA和蛋白的表达。DNA阵列数据确定了IR敏感性或耐药细胞系和组织中的YY1 mRNA表达。将稳定表达过高或敲低的YY1舌癌原代细胞和CAL27细胞暴露于红外下，分别通过CCK8测定和caspase-3测定评估细胞增殖和凋亡。我们还检查了与DNA损伤或修复相关的指标，例如YY1，p-H2AX，核PTEN，p-PTEN，Western blot分析显示Rad和Rad51。此外，我们通过引入一系列PTEN磷酸化位点突变和co-IP分析探索了IR诱导的PTEN核易位的机制。我们观察到，YY1 mRNA和蛋白质在TSCC组织中高度表达，这与较差的总体生存率相关。此外，在放射抗性细胞和组织中观察到YY1和Rad51的更高表达，YY1的过表达导致TSCC细胞中的IR抗性，而YY1敲低使TSCC细胞对IR敏感。潜在的机制表明，YY1的过表达上调了核PTEN和Rad51的表达，这对于DNA修复至关重要。IR上调YY1，PTEN核和Rad51；因此，YY1的敲低完全阻断了IR诱导的核PTEN / Rad51的上调。IR上调PTEN磷酸化，Ser380磷酸化位点的突变和突变几乎完全阻断了IR诱导的PTEN核易位。此外，磷酸酶PP2A对pS380-PTEN负调控，而YY1的敲低则完全阻断了IR通过PP2A诱导的pS380-PTEN。总之，YY1的敲低通过PP2A介导的PTEN Ser380的去磷酸化增强了TSCC的放射敏感性。因此，拮抗IR诱导的核PTEN / Rad51轴并靶向YY1可逆转TSCC中的IR抵抗力。