Redox Regulation of Cysteine-Dependent Peroxidases and Signal Transduction Pathways

半胱氨酸依赖性过氧化物酶和信号转导途径的氧化还原调节

• 批准号: 10548745
• 负责人: LESLIE B POOLE
• 金额: $ 38.75万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10548745
• 关键词: Acetylation; Active Sites; Aging; Antioxidants; Apoptosis; Area; Biophysics; Cell Cycle Regulation; Cell Signaling Process; Cells; Chemicals; Collaborations; Communicable Diseases; Cysteine; Data; Degenerative Disorder; Development; Disease; Effectiveness; Enzymatic Biochemistry; Enzymes; Eukaryota; Excision; Family; Future; Growth Factor; Health; Hot Spot; Human; Hydrogen Peroxide; Immune system; Impairment; Infectious Agent; Intervention; KRAS2 gene; Malignant Neoplasms; Mediating; Medical; NADPH Oxidase; Organism; Oxidants; Oxidation-Reduction; Paper; Peroxidases; Peroxides; Phosphorylation; Play; Post-Translational Protein Processing; Prevention; Proliferating; Proteins; Publishing; Regulation; Research Personnel; Resistance; Role; Science; Second Messenger Systems; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Source; Structure; Sulfenic Acids; System; TXN gene; Therapeutic Agents; Toxin; Work; combat; cytokine; follow-up; human disease; human pathogen; insight; mutant; nitration; novel therapeutics; oxidation; oxidative damage; peroxiredoxin; receptor internalization; response; spatiotemporal; structural determinants; tool; tumor

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.

概括 虽然过氧化氢长期以来被认为是人类免疫系统用来杀死细菌的毒素 传染性生物体，直到最近才被广泛接受，它是传染性生物体的第二信使 真核生物，响应生长因子、细胞因子和免疫系统效应物而产生，并促进或 调节下游信号转导途径。 PI 是一种半胱氨酸依赖性过氧化物还原酶，称为过氧化还原蛋白 (Prxs)，也具有 从相对默默无闻到被广泛认可，不仅仅是作为主要的氧化剂去除之一 几乎所有生物体中的系统，也是细胞信号传导途径的关键调节剂。 来自多种生物体的 Prxs 的酶学、生物物理属性和结构做出了巨大贡献 2003 年，PI Poole 和 安迪·卡普拉斯 (Andy Karplus) 合作者发表了一篇《科学》论文，其中指出 Prxs 敏感性的结构决定因素 确定了过氧化物介导的活性位点半胱氨酸的过度氧化，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”的潜在好处； 在过氧化物水平开始升高的条件下（例如 NADPH 氧化酶激活），Prx 失活会促进 来源附近过氧化物的局部积累，允许替代蛋白质目标的氧化。 普尔还处于开发化学工具的前沿，通过以下方式评估细胞中的蛋白质氧化： 这些探针现在针对的是过氧化物介导的氧化的直接产物次磺酸（R-SOH）。 商业上有售，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证明了这一点。 癌症相关生长因子引起内化受体附近蛋白质氧化的“热点”， 此处提出的未来研究将建立在我们现有的基础上。 具体来说，我们建议研究额外的翻译后的影响。 我们还将对 Prx 结构和活性进行修饰，包括硝化、乙酰化和磷酸化。 研究硫氧还蛋白调节和被人类Prxs界面调节的机制。 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 存在显着差距；我们计划跟进我们的数据，表明 Prx 失活和过氧化氢上升 最后，我们目前正在合作研究一个新领域。 与 Sharon Campbell 合作的是 KRAS 致癌 G12C 突变体的氧化敏感性，该突变体具有 这些努力将解决可能严重限制最近开发的治疗药物的有效性的问题。 对 Prx 功能和蛋白质氧化很重要的领域，从而使人们对这些领域的理解达到新的水平 可以设想采取医学和生物学相关的干预措施。