Department of Physiology, McGill University, Montreal, QC H3G 1Y6, Canada.
Quantitative Life Sciences Program, McGill University, Montreal, QC H3G 1Y6, Canada.
Cells. 2024 May 21;13(11):885. doi: 10.3390/cells13110885.
(1) Background: Stress granules (SGs) are cytoplasmic protein-RNA condensates that assemble in response to various insults. SG production is driven by signaling pathways that are relevant to human disease. Compounds that modulate SG characteristics are therefore of clinical interest. Pifithrin-µ is a candidate anti-tumor agent that inhibits members of the hsp70 chaperone family. While hsp70s are required for granulostasis, the impact of pifithrin-µ on SG formation is unknown. (2) Methods: Using HeLa cells as model system, cell-based assays evaluated the effects of pifithrin-µ on cell viability. Quantitative Western blotting assessed cell signaling events and SG proteins. Confocal microscopy combined with quantitative image analyses examined multiple SG parameters. (3) Results: Pifithrin-µ induced SGs in the absence of exogenous stress. These SGs were dynamic; their properties were determined by the duration of pifithrin-µ treatment. The phosphorylation of eIF2α was mandatory to generate SGs upon pifithrin-µ exposure. Moreover, the formation of pifithrin-µ SGs was accompanied by profound changes in cell signaling. Pifithrin-µ reduced the activation of 5'-AMP-activated protein kinase, whereas the pro-survival protein kinase Akt was activated. Long-term pifithrin-µ treatment caused a marked loss of cell viability. (4) Conclusions: Our study identified stress-related changes in cellular homeostasis that are elicited by pifithrin-µ. These insights are important knowledge for the appropriate therapeutic use of pifithrin-µ and related compounds.
(1) 背景：应激颗粒（SGs）是细胞质蛋白-RNA 凝聚物，在响应各种刺激时会组装。SG 的产生是由与人类疾病相关的信号通路驱动的。因此，调节 SG 特征的化合物具有临床意义。Pifithrin-µ 是一种候选抗肿瘤剂，可抑制 hsp70 伴侣家族的成员。虽然 hsp70s 对于颗粒稳定是必需的，但 pifithrin-µ 对 SG 形成的影响尚不清楚。
(2) 方法：使用 HeLa 细胞作为模型系统，细胞测定法评估了 pifithrin-µ 对细胞活力的影响。定量 Western blot 评估细胞信号转导事件和 SG 蛋白。共聚焦显微镜结合定量图像分析检查了多个 SG 参数。
(3) 结果：在没有外源性应激的情况下，pifithrin-µ 诱导了 SG。这些 SG 是动态的；它们的特性取决于 pifithrin-µ 处理的持续时间。pifithrin-µ 暴露时，eIF2α 的磷酸化是生成 SG 的必要条件。此外，pifithrin-µ 形成 SG 伴随着细胞信号的深刻变化。pifithrin-µ 降低了 5'-AMP 激活蛋白激酶的激活，而促生存蛋白激酶 Akt 被激活。长期 pifithrin-µ 处理导致细胞活力明显丧失。
(4) 结论：我们的研究确定了由 pifithrin-µ 引起的细胞内稳态的应激相关变化。这些见解对于 pifithrin-µ 和相关化合物的适当治疗用途非常重要。
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