INTERACTION OF ETHYL ALCOHOL WITH CELLULAR CYSTEINE PROTEASES

某些类型的细胞半胱氨酸蛋白酶可能起作用 G-蛋白偶联活性的重要调控作用 感受器。这类酶的活性受到严格的调控。 通过细胞质和核抑制物。它们之间的相互作用 靶标蛋白水解酶的抑制剂在一定程度上是由强的 疏水相互作用。酒精暴露对人体健康的影响 半胱氨酸蛋白酶活性已在细胞培养中用一种 PC12细胞株。这些蛋白水解酶的活性似乎很强 受酒精影响。我们已经确定暴露在 酒精刺激PC12细胞96小时可导致：a) 钙刺激的蛋白酶活性，这在暴露时很明显 浓度为20 mM，以及，b)两种异构体的减少 在40 mm和80 mm处很明显。这样做的结果 著作出版(J Neurochem 1997；68：1863-9)。我们是 致力于确定酒精诱导的这种情况的机制 抑制钙激活的蛋白水解酶活性。我们已经产生了 抗细胞钙调蛋白IV结构域的多克隆抗体 蛋白水解酶活性抑制物，以及 5羟色胺-3受体。使用定量RT-PCR，我们正在确定 酒精暴露和戒断是否会改变表达 卡巴斯汀，m-钙蛋白酶。Mu-calain，和5HT3受体信息。 我们发现皮质酮与酒精接触有相互作用 同时调节相关的消息水平和蛋白质水平 靶蛋白，并正在努力确定其机制 这些观察到的影响。

Certain types of cellular cysteine proteases may play an important role in modulating the activity of G-protein coupled receptors. The activity of this class of enzymes is closely regulated by cytoplasmic and nuclear inhibitors. The interaction of these inhibitors with the target proteases is, in part, mediated by strong hydrophobic interactions. The effects of ethyl alcohol exposure on cysteine protease activity has been studied in cell culture utilizing a PC12 cell line. Activity of these proteases appears to be strongly affected by alcohol exposure. We have established that exposure of PC12 cells to ethyl alcohol for 96 hours results in: a) a decrease in calcium-stimulated protease activity, which is evident at exposure concentrations of 20 mM and, b) a decrease in both isoforms of calpain, which is evident at 40 mM and 80 mM. The results of this work were published (J Neurochem 1997;68:1863-9). We are working towards defining a mechanism for this alcohol-induced inhibition of calcium-activated protease activity. We have generated a polyclonal antibodies to Domain IV of calpastatin, a cellular inhibitor of protease activity, as well as the subunit of the the 5HT-3 receptor. Using quantitative RT-PCR, we are determining whether ethanol exposure and withdrawal changes the expression of calpastatin, m-calpain. mu-calpain, and 5HT3 receptor message. We have found that corticosterone interacts with alcohol exposure to modulate both message levels and protein levels of relevant target proteins, and are working to determine the mechansims for these observed effects.