Lehman-McKeeman L D, Andrews G K, Klaassen C D
Department of Pharmacology, University of Kansas Medical Center, Kansas City 66103.
Toxicol Appl Pharmacol. 1988 Jan;92(1):1-9. doi: 10.1016/0041-008x(88)90221-9.
The purpose of this study was to determine the role of transcription, translation, and protein degradation on the accumulation of metallothionein-I (MT-I) and metallothionein-II (MT-II) in rat liver following induction of these proteins by Zn. The time course of MT induction indicated that concentrations of MT-I and MT-II, quantitated by high-performance liquid chromatography, were similar at 6 hr after administration of 1000 mumol Zn/kg (sc), but thereafter the concentration of MT-II was always higher than that of MT-I. By 24 hr after Zn administration, the concentration of MT-II in liver was more than two times that of MT-I. This difference increased with time such that by 96 hr the concentration of MT-II was more than five times that of MT-I. MT-I and MT-II mRNA levels, measured by Northern blot hybridization with mouse cRNA probes, increased coordinately following administration of Zn. MT mRNAs increased to maximum levels 6-9 hr after Zn administration, at which times MT-II mRNA was about two times more abundant than MT-I mRNA. MT mRNA levels remained elevated above control for as long as 36 hr after Zn administration. The relative rates of synthesis for MT-I and MT-II were determined by quantitating incorporation of [35S]cysteine into MTs during a 2-hr period. For both proteins, the maximum relative rates of synthesis were observed 6-9 hr after administration of Zn, in parallel with the increase in mRNA levels. When MT synthesis was at maximal levels, there was approximately two times more [35S]cysteine incorporated into MT-II than MT-I, but at no other times were differences observed. In contrast to MT mRNA levels, MT synthesis returned to control levels by 24 hr after administration of Zn. Half-lives of the isometallothioneins, determined by pulse-labeling experiments, were calculated to be 12.2 +/- 0.8 and 21.9 +/- 3.0 hr for MT-I and MT-II, respectively. Thus, Zn treatment increases transcription of both MT-I and MT-II genes and the synthesis of MT-I and MT-II. However, Zn-induced MT-II is more stable than MT-I. These results suggest that differences in the rate of synthesis and degradation of MT-I and MT-II lead to a greater and more prolonged induction of MT-II following administration of Zn.
本研究的目的是确定转录、翻译和蛋白质降解在锌诱导大鼠肝脏中金属硫蛋白-I（MT-I）和金属硫蛋白-II（MT-II）积累过程中的作用。MT诱导的时间进程表明，通过高效液相色谱法定量，在给予1000 μmol锌/千克（皮下注射）后6小时，MT-I和MT-II的浓度相似，但此后MT-II的浓度始终高于MT-I。在给予锌后24小时，肝脏中MT-II的浓度是MT-I的两倍多。这种差异随时间增加，以至于在96小时时，MT-II的浓度是MT-I的五倍多。通过用小鼠cRNA探针进行Northern印迹杂交测量，MT-I和MT-II mRNA水平在给予锌后协同增加。MT mRNA在给予锌后6 - 9小时增加到最高水平，此时MT-II mRNA的丰度约为MT-I mRNA的两倍。在给予锌后长达36小时内，MT mRNA水平一直高于对照水平。通过在2小时内定量[35S]半胱氨酸掺入MTs来确定MT-I和MT-II的相对合成速率。对于这两种蛋白质，在给予锌后6 - 9小时观察到最大相对合成速率，与mRNA水平的增加平行。当MT合成处于最高水平时，掺入MT-II的[35S]半胱氨酸比MT-I多约两倍，但在其他任何时候都未观察到差异。与MT mRNA水平相反，在给予锌后24小时，MT合成恢复到对照水平。通过脉冲标记实验确定，MT-I和MT-II的异金属硫蛋白半衰期分别计算为12.2±0.8小时和21.9±3.0小时。因此，锌处理增加了MT-I和MT-II基因的转录以及MT-I和MT-II的合成。然而，锌诱导的MT-II比MT-I更稳定。这些结果表明，MT-I和MT-II合成和降解速率的差异导致在给予锌后MT-II的诱导更大且更持久。
