Selenium in Health and its incorporation into Protein as Selenocysteine

硒是人类和其他哺乳动物饮食中必不可少的微量营养素。许多健康益处都归功于硒，包括预防各种形式的癌症(如结肠癌、前列腺癌、肺癌和肝癌)、心脏病和其他心血管和肌肉疾病。近年来，人们进行了许多人类临床试验，以评估这种元素在预防癌症、延缓艾滋病进展等方面的作用，耗资数亿美元，但人们对硒如何在哺乳动物的代谢水平上将硒结合到蛋白质中的机制知之甚少。我们几年前提出，硒的健康益处很大程度上是由于硒蛋白中这种元素的存在，即含硒氨基酸SEC。在接下来的几年里，我们在真核生物和古生物中建立了SEC的生物合成途径，重点研究了SEC tRNA的两种亚型，管家蛋白和与胁迫相关的硒蛋白的合成；并继续研究了甲基酶，称为Um34甲基酶，它在SEC tRNA的34号核苷酸的核糖部分的2‘-O位合成甲基。我们提供了强有力的证据表明，Um34与5-甲基羧甲基尿苷(McmU)加成形成5-甲基羧甲基-2‘-O-甲基尿苷(McmUm)需要mcmU与SEC氨酰化，即执行这一反应的甲基酶(命名为Um34甲基酶)的底物是硒半胱基-tRNA。此外，我们的计划侧重于建立小鼠模型，以评估管家和压力相关硒蛋白两个亚类中所有含硒蛋白的作用，以及单个硒蛋白在预防和促进癌症以及哺乳动物发育中的作用。在过去的一年里，我们完成并发表了以下项目：1)分析了在一个家庭中发现的第一个硒半胱氨酸(Sec)tRNA突变，其中父母是65位G-C碱基改变的杂合子，先证者是纯合子。患者是一名8岁男性，患有腹痛、肌肉无力和疲劳。与正常tRNA和先证者合成的应激相关硒蛋白相比，突变的SEC tRNA水平显著降低：2)膳食硒调节哺乳动物中唯一含有多个SEC残基的硒蛋白P中SEC的含量，并用半胱氨酸取代SEC；3)硒蛋白谷胱甘肽过氧化物酶4的丧失比小鼠肝脏中整个硒蛋白种群的丧失更有害，维生素E可以弥补其损失；4)对人类肝脏和肺肿瘤中的抗氧化蛋白进行了检测，并将它们与相应周围正常组织中的抗氧化蛋白进行了比较，发现肝脏更依赖硫氧还蛋白或谷胱甘肽系统来推动肿瘤的发生，而肺癌似乎主要依赖硫氧还蛋白系统；5)使用条件基因敲除小鼠模型分析了必需蛋白质硒磷酸合成酶1在调节哺乳动物氧化还原动态平衡中的作用；6)观察到小鼠结肠癌细胞系中硫氧还蛋白还原酶1或15 kDa硒蛋白(Sep15)的缺失逆转了肿瘤的几个恶性特性，但令人惊讶的是，这两种抗氧化性硒蛋白的同时丢失逆转了抗癌作用：7)小鼠肝脏中SECp43(功能未知)的敲除被发现是胚胎致死的，但尽管已知它与SEC合成酶和SEC tRNA形成复合体，但在肝脏中对硒蛋白的表达没有明显的作用；8)观察到膳食硒水平对免疫功能的影响不仅限于肝脏，并进一步确定IL-6和干扰素-γ途径对膳食硒摄入量有反应；9)两项研究涉及长肝细胞中Sep15的缺失，抑制了G1期细胞的增殖和运动，以及细胞骨架重塑和膜起泡。此外，《自然结构》杂志正在修改一份关于分析游离体中广泛的核糖体移码的手稿。比奥尔。我们的书《硒：它的分子生物学和在人类健康中的作用》第四版包括50章，已经完成，将于2016年出版。

Selenium is an essential micronutrient in the diet of humans and other mammals. Many health benefits have been attributed to selenium that include preventing various forms of cancer (e.g., colon, prostate, lung and liver cancers), heart disease and other cardiovascular and muscle disorders. Numerous human clinical trials have been undertaken in recent years to assess the role of this element in cancer prevention, delaying the progression of AIDS, etc., at a cost of hundreds of millions of dollars, but little was known about the mechanism of how selenium acts at the metabolic level in mammals to incorporate selenium into protein. We proposed several years ago that the health benefits of selenium are due largely to the presence of this element in selenoproteins as the selenium-containing amino acid, Sec. In the ensuing years, we established the biosynthetic pathway of Sec in eukaryotes and Archaea and focused on the two Sec tRNA isoforms that we demonstrated were responsible for the synthesis of the two subclasses of selenoproteins, housekeeping and stress-related selenoproteins; and pursued studying the methylase, designated Um34 methylase, that synthesizes the methyl group at the 2'-O-postion on the ribosyl moiety at nucleotide 34 of Sec tRNA. We provided strong evidence that addition of Um34 to the isoform, 5-methylcarobxymethyl-uridine (mcmU), to form 5-methylcarboxymethyl-2'-O-methyluridine (mcmUm) requires that mcmU is aminoacylated with Sec, i.e., that the substrate for the methylase (designated Um34 methylase) which carries out this reaction is selenocysteyl-tRNA. In addition, our program focused on developing mouse models to assess the role of all selenium-containing proteins within the two subclasses, housekeeping and stress-related selenoproteins, and on individual selenoproteins in preventing and promoting cancer and in mammalian development. In the past year, we have completed and published the following projects: 1) analysis of the first selenocysteine (Sec) tRNA mutation found in a family, wherein the parents were heterozygous for a G-C base change at position 65 and the proband was homozygous. The patient, an eight year old male, suffers from abdominal pain, muscle weakness and fatigue. The level of mutant Sec tRNA was reduced substantially compared to normal tRNA and the proband synthesized stress-related selenoproteins poorly; 2) dietary selenium regulates the Sec content in selenoprotein P, the only selenium-containing protein in mammals that contains multiple Sec residues, and the replacement of Sec by cysteine; 3) loss of the glutathione peroxidase 4, a selenoprotein, was far more detrimental than the loss of the entire selenoprotein population in mouse liver, and vitamin E could compensate for its loss; 4) an examination of the antioxidant proteins in human liver and lung tumors and comparing them to antioxidant proteins in the corresponding surrounding normal tissues and to each other revealed a greater dependence of liver on either the thioredoxin or glutathione system to drive the malignancy, while lung cancer appeared to depend primarily on the thioredoxin system; 5) an analysis of the role of selenophosphate synthetase 1, an essential protein, in regulating redox homeostasis in mammals using a conditional knockout mouse model; 6) the observation that the loss of thioredoxin reductase 1 or the 15kDa selenoprotein (Sep15) in a mouse colon cancer cell line reversed several malignant properties of the cancerous properties, but surprisingly the simultaneous loss of both of these antioxidant selenoproteins reversed the anticancer effects; 7) knockout of SECp43 (function unknown) in mouse liver was found to be embryonic lethal, but had no apparent role in selenoprotein expression in liver even though it was known to form a complex with Sec synthase and Sec tRNA; 8) observation that the effects of dietary selenium levels on immune function are not limited to liver and further identify the IL-6 and interferon-gamma pathways as being responsive to dietary selenium intake; and 9) two studies involving loss of Sep15 in Chang liver cells inhibiting cell proliferation and motility at the G1 phase and cytoskeleton remodeling and membrane blebbing. In addition, a manuscript on an analysis of extensive ribosomal frameshifting in Euplotes is under revision at Nature Struct. Biol. and the 4th edition of our book entitled "Selenium: Its Molecular Biology and Role in Human Health", containing 50 chapters, was completed and will be published in 2016.