Decrease in MAP3Ks expression enhances the cell death caused by hyperthermia
Decrease in MAP3Ks expression enhances the cell death caused by hyperthermia
复制标题
DOI：
10.1080/02656736.2021.2024281
复制
发表时间：
2022-12-31
影响因子：
3.1
通讯作者：
Miyagawa，Kiyoshi
中科院分区：
文献类型：
--
作者：
Enomoto，Atsushi;Fukasawa，Takemichi;Miyagawa，Kiyoshi
目的热疗是一种很有前途的抗肿瘤治疗方法。然而，肿瘤细胞热敏感性的分子机制在很大程度上是未知的。本研究的目的是澄清如何由热刺激antimicrobial activity.Methods和materialsThe各种MAPK成员在HeLa细胞中的表达水平与或无热疗引发的生化变化进行了评估，通过Western印迹和RT-PCR。用CaTM-2 AM数字成像仪监测细胞内Ca ~（2+）浓度[Ca ~（2+）] i。采用体外切割实验检测钙依赖性蛋白酶calpain是否切割MAPK组分。细胞增殖和克隆形成进行了评估的siRNA靶向MAPK members.ResultsHyperthermia的存在或不存在的情况下，降低了水平的MAP 3 K TAK 1，RAF 1和MEKK 2，但不是下游的MAP 2K和MAPK成员。蛋白酶体抑制剂MG 132或钙蛋白酶抑制剂ALLN可挽救高血压诱导的TAK 1和MEKK 2降解;然而，RAF 1不受抑制剂的影响。热诱导RAF 1下调。热疗增加[Ca ~（2+）] i和calpain I表达。钙离子载体A23187降低TAK 1和MEKK 2水平。体外切割实验证明TAK 1和MEKK 2是钙蛋白酶I的底物。TAK 1，RAF 1和MEKK 2的敲低抑制细胞增殖和clonogenicity.ConclusionsHyperthermia降低了MAP 3 K TAK 1，RAF 1和MEKK 2的水平，而不减少的下游成分的MAP 3 K-MAP 2K-MAPK级联反应，钙蛋白酶依赖性降解途径或转录调控。TAK 1、RAF 1和/或MEKK 2在细胞增殖和克隆形成中起关键作用，并且是热疗的潜在分子靶点。
PurposeHyperthermia is a promising anticancer treatment modality. However, the molecular mechanism underlying the thermal sensitivity of tumor cells is largely unknown. The aim of this study was to clarify how biochemical changes triggered by heat stimulate antitumor activity.Methods and materialsThe expression levels of various MAPK members in HeLa cells with or without hyperthermia were evaluated by western blotting and RT-PCR. The intracellular Ca2+concentration [Ca2+]iwas monitored by digital imaging using CaTM-2 AM. Anin vitrocleavage assay was used to determine whether calcium-dependent protease calpain cleaves MAPK components. Cell proliferation and clonogenicity were assessed in the absence or presence of siRNAs targeting MAPK members.ResultsHyperthermia decreased the levels of MAP3K TAK1, RAF1 and MEKK2 but not of the downstream MAP2K and MAPK members. The hyperthermia-induced degradation of TAK1 and MEKK2 was rescued by either the proteasome inhibitor MG132 or the calpain inhibitor ALLN; however, RAF1 was not affected by the inhibitors. Heat induced down regulation ofRAF1. Hyperthermia increased [Ca2+]iand calpain I expression. The calcium ionophore A23187 decreased TAK1 and MEKK2 levels. Anin vitrocleavage assay demonstrated that TAK1 and MEKK2 are calpain I substrates. Knockdown of TAK1, RAF1 and MEKK2 suppressed cell proliferation and clonogenicity.ConclusionsHyperthermia decreased the levels of MAP3K TAK1, RAF1 and MEKK2, without reduction of the downstream components in the MAP3K-MAP2K-MAPK cascade, by a calpain-dependent degradation pathway or transcriptional regulation. TAK1, RAF1 and/or MEKK2 play crucial roles in cell proliferation and clonogenicity and are potential molecular targets for hyperthermia.