Cao Y, Matsumoto T, Motomura K, Ohtsuru A, Yamashita S, Kosaka M
Department of Environmental Physiology, Institute of Tropical Medicine, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan.
Pflugers Arch. 1998 Dec;437(1):15-20. doi: 10.1007/s004240050740.
Cells of a temperature-sensitive mutant line (tsFT101) derived from a mouse mammary carcinoma cell line (FM3A) become multinucleated at a non-permissive temperature of 39 degrees C because of disturbed cytokinesis. To explore how this relates to thermotolerance, we examined the proliferative activity of, and heat shock protein (HSP) expression in, FM3A and tsFT101 cells cultured at 37 degrees C and 39 degrees C after heat shock pretreatment (15 min exposure at 45 degrees C). FM3A cells developed thermotolerance when cultured at both 37 degrees C and 39 degrees C, but whereas tsFT101 cells developed thermotolerance at 37 degrees C, this was markedly reduced at 39 degrees C. Western blot analysis showed similar degrees of expression of constitutive HSP70 (HSP73) in FM3A and tsFT101 cells after heat shock pretreatment at both 37 degrees C and 39 degrees C. However, expression of inducible HSP70 (HSP72) was reduced in tsFT101 cells at 39 degrees C compared to 37 degrees C and to FM3A cells at both 37 degrees C and 39 degrees C. Heat shock pretreatment activated DNA binding of heat shock transcription factor (HSF) in FM3A cells at 37 degrees C and 39 degrees C, but only at 37 degrees C in tsFT101 cells. These results indicate that (1) multinucleation caused by disturbed cytokinesis increases temperature sensitivity, (2) HSP70 is critical for the development of thermotolerance in both FM3A and tsFT101 cells, and (3) decreased expression of inducible HSP70 parallels deficient development of thermotolerance in tsFT101 cells cultured at a non-permissive temperature.
源自小鼠乳腺癌细胞系（FM3A）的温度敏感突变株系（tsFT101）的细胞，在39℃的非允许温度下会因胞质分裂受扰而形成多核。为探究这与热耐受性的关系，我们检测了热休克预处理（45℃暴露15分钟）后，在37℃和39℃培养的FM3A和tsFT101细胞的增殖活性及热休克蛋白（HSP）表达。FM3A细胞在37℃和39℃培养时均产生了热耐受性，但tsFT101细胞虽在37℃产生了热耐受性，在39℃时却明显降低。蛋白质印迹分析表明，在37℃和39℃进行热休克预处理后，FM3A和tsFT101细胞中组成型HSP70（HSP73）的表达程度相似。然而，与37℃时相比，以及与37℃和39℃时的FM3A细胞相比，tsFT101细胞在39℃时诱导型HSP70（HSP72）的表达降低。热休克预处理在37℃和39℃时激活了FM3A细胞中热休克转录因子（HSF）的DNA结合，但在tsFT101细胞中仅在37℃时激活。这些结果表明：（1）胞质分裂受扰导致的多核化增加了温度敏感性；（2）HSP70对FM3A和tsFT101细胞热耐受性的产生至关重要；（3）在非允许温度下培养的tsFT101细胞中，诱导型HSP70表达的降低与热耐受性发育缺陷平行。
