Regulation of Homocysteine-dependent Redox Homeostasis

描述（由申请人提供）：氧化应激引起的氧化还原稳态的扰动似乎是连接各种复杂和多因素疾病（如心血管疾病、帕金森病、阿尔茨海默病、关节炎和某些癌症）病因的共同线索。谷氨酸是真核生物细胞内抗氧化剂库的关键组分，其合成中的限制性试剂被认为是半胱氨酸。将高半胱氨酸转化为半胱氨酸的转硫途径是肝脏中细胞内半胱氨酸库的定量显著贡献者，谷胱甘肽中约50%的半胱氨酸是通过该途径获得的。同型半胱氨酸是一种含硫氨基酸，其水平升高与许多多因素疾病相关，如心血管疾病、神经管缺陷和阿尔茨海默病。然而，这种氨基酸的细胞内调节和谷胱甘肽的主要细胞氧化还原缓冲池的代谢联系知之甚少。该建议侧重于三个关键位点的重要性，在调节同型半胱氨酸浓度：蛋氨酸合成酶，蛋氨酸合成酶还原酶和胱硫醚β-合成酶。这些酶中的每一种的突变都与遗传性高同型半胱氨酸血症相关，高同型半胱氨酸血症是一种常染色体隐性遗传疾病。通过结合生物物理学、细胞生物学和小鼠模型对胱硫醚β-合酶缺乏症的研究，我们将：（i）表征甲硫氨酸合成酶还原酶中的多态性变异和遗传突变如何影响高半胱氨酸的甲硫氨酸脱氢酶依赖性转甲基化的氧化还原活化，（ii）阐明甲硫氨酸合酶的翻译调节机制，和（iii）阐明同型半胱氨酸代谢的氧化还原调节机制及其对谷胱甘肽稳态的影响。这些研究将为高半胱氨酸代谢的多水平调节提供重要的见解，并将评估在细胞培养和小鼠中调节转硫途径对谷胱甘肽依赖性抗氧化防御的影响。

DESCRIPTION (provided by applicant): Perturbations of redox homeostasis incurred by oxidative stress appear to be a common thread connecting the etiologies of various complex and multifactorial diseases, such as cardiovascular diseases, Parkinson's disease, Alzheimer's disease, arthritis and some cancers. Glutathione is a key component of the intracellular arsenal of antioxidants in eukaryotes, and the limiting reagent in its synthesis is believed to cysteine. The transsulfuration pathway which converts homocysteine to cysteine is a quantitatively significant contributor to the intracellular cysteine pool in the liver and -50 percent of the cysteine in glutathione is derived via this pathway. Homocysteine is a sulfur containing amino acid whose elevated levels are correlated with a number of multifactorial diseases such as cardiovascular diseases, neural tube defects and Alzheimer's disease. However, intracellular regulation of this amino acid and of the metabolic link to the major cellular redox buffer pool of glutathione are poorly understood. This proposal focuses on three key loci important in regulation of homocysteine concentrations: methionine synthase, methionine synthase reductase and cystathionine beta-synthase. Mutations in each of these enzymes is correlated with hereditary hyperhomocystenemia which is inherited as an autosomal recessive disorder. Using a combination of biophysical, cell biological and mouse model studies for cystathionine beta-synthase deficiency, we will; (i) characterize how polymorphic variations and hereditary mutations in methionine synthase reductase influence redox activation of methionine synthase-dependent transmethyiation of homocysteine, (ii) elucidate the mechanism of translational regulation of methionine synthase and (iii) elucidate the mechanism of redox regulation of homocysteine metabolism and its influence on glutathione homeostasis. These studies will provide important insights into multiple levels of regulation of homocysteine metabolism and will assess the effects of modulating the transsulfuration pathway on glutathione-dependent antioxidant defense in cell culture and in mice.