The study of modulating effects on X-ray induced cell mutability by chaperons and SUMO.

我们以前报道过，人血清因子对细胞变异性的调节与蛋白酶活性水平的增加有关。目前的工作旨在研究细胞易变性与伴侣如GRP78、GRP94、HSP27和SUMO-3之间的关系。：(1) grp94；通过对x射线诱导蛋白酶的分析，其中一个伴侣子GRP94被鉴定为中介分子的候选基因。利用siRNA对GRP94 mRNA进行集落形成实验，提示GRP94参与细胞x射线抗性。(2)通过对紫外光诱导蛋白酶的分析，我们还发现了其他的伴侣蛋白HSP27。当抑制HSP27的表达时，紫外线照射后6-4光产物的去除活性降低。我们进一步发现GRP78也参与了紫外线损伤的修复机制。因此，伴侣子对细胞突变的调节似乎是通过蛋白酶信号传导介导的。(3)其中一种泛素样蛋白SUMO-3被鉴定为x射线照射后DNA合成活性异常的候选基因。已知SUMO-3的功能受特定蛋白酶的调节。x射线照射后DNA合成活性升高，SUMO-3表达降低。我们表达了GFP-SUMO-3融合蛋白，观察到x射线照射后细胞核中有明显浓度的绿色荧光。评价人血清氧化应激新方法的应用。采用血清过氧化氢法监测氧化应激。部分患者和重度吸烟者血清过氧化氢水平高于健康志愿者。

We previously reported that modulation of cell mutability by human serum factors is associated with increased levels of protease activity. The present work is designed to examine the relationships between cell mutability and chaperons such as GRP78,GRP94,HSP27 and SUMO-3. : (1)GRP94 ; One of the chaperons, GRP94, was identified as a candidate gene for the mediator molecule by analysis of X-ray-induced protease. GRP94 was suggested to be involved in cellular X-ray resistance by the colony forming assay using siRNA for GRP94 mRNA.(2)We also identified other chaperons, HSP27, by analysis of UV-induced protease. The activity of removing 6-4 photo products after UV irradiation was decreased when expression of HSP27 was suppressed. We further identified that GRP78 is also involved in the repair mechanism for the UV-induced damage. Thus, the modulation of cell mutability by chaperons seems to be mediated though protease signaling.(3)One of ubiquitin-like proteins, SUMO-3, was identified as a candidate gene for abnormal DNA synthesis activity after X-ray irradiation. It is known that the function of SUMO-3 is regulated by specific proteases. The activity of DNA synthesis after X-ray irradiation was increased when expression of SUMO-3 was decreased. We expressed GFP-SUMO-3 fusion protein and observed a marked concentration of green fluorescence in the nuclear after X-ray irradiation.Application of a new method to evaluate oxidative stress in human serum. A test to monitor the oxidative stress by evaluating serum hydroperoxide was carried out. Serum hydrpperoxide of some patients and heavy-smokers showed higher levels than healthy volunteers.