Analysis of regulatory mechanism for signal transduction responding to oxidative stress

众所周知，PKC在影响多种细胞反应的受体介导的信号转导中起着至关重要的作用。我们已经证明氧化二酰基甘油（DAG-OOH）激活大鼠脑PKC的效率与苯酚酯（PMA）一样高，后者是PKC的人工强效活化剂。这一结果表明，DAG-OOH可能在氧化应激中作为生物信使，有效地改变pkc依赖的信号转导系统，其程度与PMA相似。此外，DAG-OOH通过过度激活PKC δ和MAP激酶来损伤培养的神经元。在这些研究中，人们怀疑大鼠脑中存在两种PKC异构体（PKC- x：比其他PKC异构体更容易受到DAG- ooh的影响，PKC DSV: PKC δ剪接变体，只有DAG结合结构域而没有激酶结构域）。PKC DSV被认为是DAG-OOH和PKC δ诱导的神经元细胞损伤的调节分子。本研究通过细胞过表达PKC DSV、PKC δ和PKC α基因来研究PKC DSV的作用。2006年，制备了这些基因的腺病毒表达载体。用这些表达载体转染PC12细胞。2007年，PKC dsv转染的细胞被添加PKC激活剂如PMA刺激。转染PKC DSV基因可抑制PMA刺激诱导的生长信号。这些结果表明PKC DSV调节DAG-OOH刺激引起的PKC δ的过度激活。

It is well known that PKC plays a crucial role in receptor-mediated signal transduction affecting diverse range of cellular responses. We have demonstrated that oxidized-diacylglycerol (DAG-OOH) activates rat brain PKC as efficiently as phorbol ester (PMA), artificial and powerful activator for PKC. This result markedly suggested that DAG-OOH might act as a biological messenger in oxidative stress to efficiently alter the PKC-dependent signal transduction system to a similar extent to PMA. Furthermore, DAG-OOH injured the cultured neurons with the over-activation of PKC delta and MAP kinase. Among these studies, it has been suspected that two types of PKC isoforms (PKC-X: more susceptible to DAG-OOH than other PKC isoforms, PKC DSV: PKC delta splicing variant, which has only DAG binding domain but not kinase domain) exist in rat brain. PKC DSV is thought to act as a regulatory molecule for the DAG-OOH and PKC delta-induced neuronal cell injury. In the present study, the role of PKC DSV has been examined by means of the cells over-expressed the genes for PKC DSV, PKC delta and PKC alpha. In 2006, the adenovirus expression vectors for these genes were prepared. PC12 cells were transfected with these expression vectors. In 2007, PKC DSV-transfected cells were stimulated by the addition of PKC activator such as PMA. The growth signal induced by PMA irritation has been suppressed by the transfection of PKC DSV gene. These results suggested that PKC DSV regulate the over-activation of PKC delta provoked by DAG-OOH stimulation.