Redox Regulation of Cysteine-Dependent Peroxidases and Signal Transduction Pathways

总结 虽然过氧化氢一直被认为是一种毒素，被人体免疫系统用来杀死 直到最近，它才被广泛接受，它作为第二信使， 真核生物，响应于生长因子、细胞因子和免疫系统效应物而产生，并促进或 调节下游信号转导通路。通过部分由联合国工作人员的工作提供的见解， PI，一个半胱氨酸依赖性过氧化物还原酶家族，被称为过氧化物氧化还原酶（Prxs）， 从相对默默无闻的情况出现，成为广泛认可的不仅仅是作为一个主要的氧化剂去除 在几乎所有生物体中，它不仅是细胞信号传导途径的关键调节剂，而且是细胞信号传导途径的关键调节剂。PI Poole的工作 来自各种生物体的Prxs的酶学、生物物理属性和结构极大地有助于 了解这个高度丰富的酶家族的机制和调节。2003年，PI Poole和 合作者安迪·卡普拉斯发表了一篇《科学》杂志的论文， 对过氧化物介导的活性部位半胱氨酸的过氧化作用进行了鉴定。这导致了我们的建议， “闸门假说”解释了这种过氧化物介导的“关闭开关”的潜在益处; 在过氧化物水平开始升高（例如NADPH氧化酶活化）的条件下，Prx失活将促进 过氧化物在来源附近的局部积累，允许替代蛋白质靶的氧化。博士 普尔还一直处于开发化学工具的最前沿，以评估细胞中的蛋白质氧化， 靶向次磺酸（R-SOH），过氧化物介导的氧化的直接产物。这些探测器现在 其可商购获得，并已被研究氧化还原调节和信号传导的研究人员广泛使用， 以高时空精度评估蛋白质氧化。PI Poole的实验室使用这些工具表明， 癌症相关生长因子引起邻近内化受体的蛋白质氧化的“热点”， 为闸门假说提供了支持未来的研究将建立在我们现有的基础上。 优势和合作。具体来说，我们建议研究额外的翻译后效应， 修饰，包括硝化、乙酰化和磷酸化对Prx结构和活性的影响。我们还将 研究硫氧还蛋白可以调节和被人Prxs调节的机制。接口 Prx功能与涉及蛋白质氧化的信号转导途径的调节是另一个领域 我们计划跟进我们的数据，这些数据表明Prx失活和过氧化物升高 水平是细胞周期调节的关键。最后，我们目前正在合作研究的一个新领域 与Sharon坎贝尔合作的是KRAS的致癌G12 C突变体的氧化敏感性，该突变体具有 可能严重限制最近开发的治疗剂的有效性。这些努力将解决 对Prx功能和蛋白质氧化很重要的区域，导致了一个新的理解水平， 可以设想与医学和生物学相关的干预措施。

SUMMARY While hydrogen peroxide has long been understood as a toxin used by the human immune system to kill infectious organisms, only recently has it become well accepted that it serves as a second messenger in eukaryotes, produced in response to growth factors, cytokines and immune system effectors and promoting or modulating downstream signal transduction pathways. Through insights contributed in part by the work of the PI, a family of cysteine-dependent, peroxide-reducing enzymes known as the peroxiredoxins (Prxs) have also emerged from relative obscurity to become widely recognized not just as one of the primary oxidant removal systems in almost all organisms, but also as key modulators of cell signaling pathways. PI Poole's work on the enzymology, biophysical attributes and structures of Prxs from a variety of organisms has contributed greatly to understanding the mechanism and regulation of this highly abundant family of enzymes. In 2003, PI Poole and collaborator Andy Karplus published a Science paper in which structural determinants of the sensitivity of Prxs toward peroxide-mediated hyperoxidation of the active site cysteine were identified. This led to our proposal of the “floodgate hypothesis” explaining the potential benefits of such a peroxide-mediated “off switch”; under conditions where peroxide levels begin to rise (e.g. NADPH oxidase activation), Prx inactivation would promote the local accumulation of peroxide near the source, allowing for the oxidation of alternative protein targets. Dr. Poole has also been at the forefront of developing chemical tools to evaluate protein oxidation in cells through targeting sulfenic acid (R-SOH), the direct product of peroxide-mediated oxidation. These probes are now commercially available and have been used widely by researchers studying redox regulation and signaling to evaluate protein oxidation with high spatiotemporal precision. PI Poole's lab used these tools to show that cancer-associated growth factors elicit “hot spots” of protein oxidation proximal to the internalized receptors, providing support for the floodgate hypothesis. Future studies proposed here will build upon our existing strengths and collaborations. Specifically, we propose to investigate the effects of additional posttranslational modifications, including nitration, acetylation and phosphorylation, on Prx structure and activity. We will also investigate the mechanism by which thioredoxin can regulate and be regulated by human Prxs. The interface of Prx function with the regulation of signal transduction pathways involving protein oxidation is another area with significant gaps; we plan to follow up on our data suggesting that Prx inactivation and rising peroxide levels are key to cell cycle regulation. Finally, a new area that we are currently investigating in collaboration with Sharon Campbell is the oxidation sensitivity of the cancer-causing G12C mutant of KRAS, which has the potential to severely limit the effectiveness of recently developed therapeutic agents. These efforts will address areas important to Prx function and protein oxidation, leading to a new level of understanding through which medically-and biologically-relevant interventions could be envisioned.