Laboratory of Molecular Radiology, Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.
Department of Dermatology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.
Int J Hyperthermia. 2024;41(1):2335199. doi: 10.1080/02656736.2024.2335199. Epub 2024 Apr 2.
c-Jun N-terminal kinases (JNKs) comprise a subfamily of mitogen-activated protein kinases (MAPKs). The JNK group is known to be activated by a variety of stimuli. However, the molecular mechanism underlying heat-induced JNK activation is largely unknown. The aim of this study was to clarify how JNK activity is stimulated by heat.
The expression levels of various MAPK members in HeLa cells, with or without hyperthermia treatment, were evaluated via western blotting. The kinase activity of MAPK members was assessed through kinase assays. Cell death was assessed in the absence or presence of siRNAs targeting MAPK-related members.
Hyperthermia decreased the levels of MAP3Ks, such as ASK1 and MLK3 which are JNK kinase kinase members, but not those of the downstream MAP2K/SEK1 and MAPK/JNK. Despite the reduced or transient phosphorylation of ASK1, MLK3, or SEK1, downstream JNK was phosphorylated in a temperature-dependent manner. kinase assays demonstrated that heat did not directly stimulate SEK1 or JNK. However, the expression levels of DUSP16, a JNK phosphatase, were decreased upon hyperthermia treatment. DUSP16 knockdown enhanced the heat-induced activation of ASK1-SEK1-JNK pathway and apoptosis.
JNK was activated in a temperature-dependent manner despite reduced or transient phosphorylation of the upstream MAP3K and MAP2K. Hyperthermia-induced degradation of DUSP16 may induce activation of the ASK1-SEK1-JNK pathway and subsequent apoptosis.
c-Jun N-末端激酶（JNKs）是丝裂原活化蛋白激酶（MAPKs）的一个亚家族。已知 JNK 组被各种刺激物激活。然而，热诱导 JNK 激活的分子机制在很大程度上尚不清楚。本研究旨在阐明 JNK 活性如何受到热的刺激。
通过 Western blot 评估有或没有热疗处理的 HeLa 细胞中各种 MAPK 成员的表达水平。通过激酶测定评估 MAPK 成员的激酶活性。在不存在或存在针对 MAPK 相关成员的 siRNA 的情况下评估细胞死亡。
热疗降低了 JNK 激酶激酶成员 ASK1 和 MLK3 等 MAP3K 的水平，但不降低下游 MAP2K/SEK1 和 MAPK/JNK 的水平。尽管 ASK1、MLK3 或 SEK1 的磷酸化减少或短暂，但下游 JNK 以温度依赖的方式磷酸化。激酶测定表明，热不会直接刺激 SEK1 或 JNK。然而，热疗处理后 JNK 磷酸酶 DUSP16 的表达水平降低。DUSP16 敲低增强了热诱导的 ASK1-SEK1-JNK 途径激活和细胞凋亡。
尽管上游 MAP3K 和 MAP2K 的磷酸化减少或短暂，但 JNK 仍以温度依赖的方式激活。热诱导的 DUSP16 降解可能诱导 ASK1-SEK1-JNK 途径的激活和随后的细胞凋亡。
