Cheng T J, Chen T M, Chen C H, Lai Y K
Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan, Republic of China.
J Cell Biochem. 1998 May 1;69(2):221-31. doi: 10.1002/(sici)1097-4644(19980501)69:2<221::aid-jcb12>3.0.co;2-h.
We herein demonstrate that sodium fluoride (NaF) acts as a stress response inducer on HeLa and 9L rat brain tumor cells. NaF is only slightly cytotoxic, and inhibitory to Ser/Thr-phosphatases but not to Tyr-phosphatases in both cell lines. After treatment with 5 mM NaF for 2 h, the phosphorylation levels of vimentin and an alkali-resistant 65-kDa phosphoprotein were enhanced, a common phenomenon detected in cells under a variety of stress conditions. Under an identical treatment protocol, in which the cells were treated with 5 mM NaF for 2 h and then allowed to recover under normal growing conditions for up to 12 h, NaF differentially induced the cytoplasmic/nuclear heat-shock protein70s (including both the inducible and the constitutively expressed members of this protein family) in HeLa cells and the endoplasmic reticulum residing heat-shock protein70 (the glucose-regulated protein with an apparent molecular weight of 78 kDa) in 9L cells. Electrophoretic mobility shift assays (EMSA) using probes containing well-characterized regulatory elements revealed the activation of the heat-shock factor in HeLa but not in 9L cells; this is in good agreement with the stress protein induction pattern. Additional differential induction of binding activities toward EMSA probes individually containing NF-KB, AP-2, and CRE-like elements were detected in NaF-treated cells. The possible involvement of these binding sites as well as the corresponding factors in the stress response are discussed.
我们在此证明，氟化钠（NaF）对HeLa细胞和9L大鼠脑肿瘤细胞起到应激反应诱导剂的作用。NaF仅有轻微的细胞毒性，并且在这两种细胞系中均抑制丝氨酸/苏氨酸磷酸酶，但不抑制酪氨酸磷酸酶。用5 mM NaF处理2小时后，波形蛋白和一种耐碱的65 kDa磷蛋白的磷酸化水平增强，这是在各种应激条件下细胞中检测到的常见现象。在相同的处理方案下，即细胞用5 mM NaF处理2小时，然后在正常生长条件下恢复长达12小时，NaF在HeLa细胞中差异性地诱导细胞质/细胞核热休克蛋白70（包括该蛋白家族的诱导型和组成型表达成员），在9L细胞中诱导内质网驻留热休克蛋白70（表观分子量为78 kDa的葡萄糖调节蛋白）。使用含有特征明确的调控元件的探针进行的电泳迁移率变动分析（EMSA）显示，热休克因子在HeLa细胞中被激活，但在9L细胞中未被激活；这与应激蛋白诱导模式高度一致。在NaF处理的细胞中还检测到对分别含有NF-κB、AP-2和CRE样元件的EMSA探针的结合活性的额外差异性诱导。讨论了这些结合位点以及相应因子在应激反应中的可能作用。
