Department of Anesthesiology and Pain Medicine, School of Medicine, Chosun University, 309 Pilmundaero, Dong-gu, Gwangju, 501-759, Korea.
School of Medicine, Chosun University, 309 Pilmundaero, Dong-gu, Gwangju, 501-759, Korea.
Exp Mol Med. 2022 May;54(5):685-696. doi: 10.1038/s12276-022-00782-4. Epub 2022 May 27.
Cadmium (Cd) is a highly toxic environmental pollutant that can severely damage the kidneys. Here, we show that Cd-induced apoptosis is promoted by the cytoplasmic polyubiquitination of p53 (polyUb-p53), which is regulated by the polyubiquitination of SQSTM1/p62 (polyUb-p62) and autophagy in mouse kidney mesangial cells (MES13E cells). p53 was detected in monomeric and different high-molecular-weight (HMW) forms after Cd exposure. Monomeric p53 levels decreased in a concentration- and time-dependent manner. HMW-p53 transiently accumulated in the cytoplasm independent of proteasome inhibition. The expression patterns of p53 were similar to those of p62 upon Cd exposure, and the interactions between polyUb-p53 and polyUb-p62 were observed using immunoprecipitation. P62 knockdown reduced polyUb-p53 and upregulated nuclear monomeric p53, whereas p53 knockdown reduced polyUb-p62. Autophagy inhibition induced by ATG5 knockdown reduced Cd-induced polyUb-p62 and polyUb-p53 but upregulated the levels of nuclear p53. Pharmacological inhibition of autophagy by bafilomycin A1 increased polyUb-p62 and polyUb-p53 in the cytoplasm, indicating that p53 protein levels and subcellular localization were regulated by polyUb-p62 and autophagy. Immunoprecipitation and immunofluorescence revealed an interaction between p53 and LC3B, indicating that p53 was taken up by autophagosomes. Cd-resistant RMES13E cells and kidney tissues from mice continuously injected with Cd had reduced polyUb-p53, polyUb-p62, and autophagy levels. Similar results were observed in renal cell carcinoma cell lines. These results indicate that cytoplasmic polyUb-p53 is a potential biomarker for Cd-induced acute toxicity in mesangial cells. In addition, upregulation of nuclear p53 may protect cells against Cd cytotoxicity, but abnormal p53 accumulation may contribute to tumor development.
镉 (Cd) 是一种剧毒的环境污染物，可严重损害肾脏。在这里，我们表明 Cd 诱导的细胞凋亡是由细胞质多泛素化 p53（多泛素化-p53）促进的，这是由 SQSTM1/p62（多泛素化-p62）的多泛素化和自噬调节的。在 Cd 暴露后，p53 被检测为单体和不同的高分子量（HMW）形式。单体 p53 水平呈浓度和时间依赖性下降。HMW-p53 在不抑制蛋白酶体的情况下，在细胞质中短暂积累。p53 的表达模式与 Cd 暴露后的 p62 相似，并且通过免疫沉淀观察到多泛素化-p53 和多泛素化-p62 之间的相互作用。P62 敲低减少了多泛素化-p53 并上调了核单体 p53，而 p53 敲低减少了多泛素化-p62。ATG5 敲低诱导的自噬抑制减少了 Cd 诱导的多泛素化-p62 和多泛素化-p53，但上调了核 p53 的水平。用巴弗洛霉素 A1 抑制自噬会增加细胞质中多泛素化-p62 和多泛素化-p53，表明 p53 蛋白水平和亚细胞定位受多泛素化-p62 和自噬调节。免疫沉淀和免疫荧光显示 p53 和 LC3B 之间存在相互作用，表明 p53 被自噬体摄取。Cd 抗性 RMES13E 细胞和连续注射 Cd 的小鼠的肾脏组织中，多泛素化-p53、多泛素化-p62 和自噬水平降低。在肾癌细胞系中也观察到类似的结果。这些结果表明，细胞质多泛素化-p53 可能是肾小球系膜细胞中 Cd 诱导的急性毒性的潜在生物标志物。此外，核 p53 的上调可能保护细胞免受 Cd 细胞毒性，但异常的 p53 积累可能导致肿瘤发展。
