Role of Selenium in Cancer and Health

硒是人类和其他哺乳动物饮食中的一种基本微量营养素，已知在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方面具有作用。此外，它还作为一种抗病毒药物，在延缓衰老过程、延缓HIV阳性患者的艾滋病进展、免疫功能、哺乳动物发育和雄性生殖方面发挥作用。人们对硒的作用机制知之甚少，但我们认为这在很大程度上是由于硒蛋白中含有硒半胱氨酸(Sec)这种元素。因此，我们的计划侧重于SEC被结合到蛋白质中的方法以及特定的硒蛋白在人类健康和发育中的作用。由于SEC tRNA是唯一已知的控制一整类蛋白质--硒蛋白表达的tRNA，我们开发了几个转基因、基因敲除、条件基因敲除和基因敲除的细胞和小鼠株系，它们扰乱SEC tRNA的合成，进而扰乱硒蛋白的表达，以此作为模型来更好地了解硒和硒蛋白的代谢及其在健康中的作用。在过去的一年里，我们的计划重点是利用RNAi技术敲除几种硒蛋白和其他参与SEC生物合成的蛋白(例如SLA、SECp43、PSTK、SPS1和2)，以阐明它们在硒代谢中的作用，并确定该元件是如何结合到蛋白质中的。本课题组进一步研究了SECp43和SLA在SEC生物合成及其与蛋白质结合中的作用。此外，我们开发了一种新的方法来研究硒蛋白的功能，包括敲除/敲入，并正在使用这一技术来研究许多以前不容易接近的硒蛋白功能的新参数。利用loxP-Cre技术，我们的目标是去除小鼠肝脏中的硒蛋白表达，然后分别用一个野生型或两个不同的突变型SEC tRNA转基因基因完全或部分取代硒蛋白群体。突变的SEC tRNA转基因基因涉及tRNA的突变，该突变阻止在反密码子的摆动位置37位添加甲基2‘-O-甲基核糖(标记为Um34)。Um34的缺失导致与胁迫相关的硒蛋白的表达丧失，而看家的硒蛋白几乎完全表达。我们早些时候通过使用野生型和突变型转基因拯救标准基因敲除小鼠中的硒蛋白群体表明，Um34参与了与应激相关的硒蛋白的合成。然而，我们现在针对特定器官或组织的能力将使我们能够专注于硒和硒蛋白在给定细胞类型中的作用。应该进一步注意的是，这种新的蛋白质表达调控发生在翻译水平上，并表现出组织特异性的模式。我们还在扩大我们对不同组织和器官中硒蛋白表达的定向敲除，并明确显示了硒蛋白在发育和疾病预防中的作用。在过去的一年里，我们在这一领域的主要关注之一是硒蛋白在免疫功能中的作用。我们的目标是去除T细胞中硒蛋白的表达，观察到CD8群体减少了约50%，mIL-13在雄性基因敲除小鼠中增加了55倍以上，在雌性基因敲除小鼠中增加了80倍。在目标小鼠中，IgG2a增加了约两倍。到目前为止，这些研究表明T细胞中的硒蛋白与炎性过敏免疫反应有关，在T细胞的发育/分化/成熟和功能中起着重要作用，是决定Th1和Th2反应的不可或缺的因素。

Selenium is an essential micronutrient in the diet of humans and other mammals and is known to have roles in preventing various forms of cancer (e.g., colon, prostate, lung and liver), heart disease and other cardiovascular and muscle disorders. In addition, it serves as an antiviral agent and plays a role in delaying the aging process, in delaying the progression of AIDS in HIV positive patients, in immune function, in mammalian development and in male reproduction. The means by which selenium exerts these many health benefits are poorly understood, but we have proposed that they are due largely to the presence of this element in selenoproteins as the amino acid, selenocysteine (Sec). Our program therefore focuses on the means by which Sec is incorporated into protein and the role of specific selenoproteins in human health and development. Since Sec tRNA is the only known tRNA that governs the expression of an entire class of proteins, the selenoproteins, we have developed several transgenic, knockout, conditional knockout and knockdown cell and mouse lines that perturb Sec tRNA synthesis which in turn perturbs selenoprotein expression as models to better understand selenium and selenoprotein metabolism and their roles in health. This past year, our program has focused on using RNAi technology to knockdown several selenoproteins and other proteins involved in Sec biosynthesis (e.g., SLA, SECp43, PSTK, SPS1 and 2) to elucidate their roles in selenium metabolism and to determine how this element is incorporated into protein. Our group further characterized the roles of SECp43 and SLA in the biosyntheisis of Sec and its incorporation into protein. In addition, we developed a novel assay for studying selenoprotein function that involved knockdown/knock-in and are using this technique to study many new parameters of selenoprotein function that could not readily be approached previously. Using loxP-Cre technology, we targeted the removal of selenoprotein expression in mouse liver and then replaced the selenoprotein population either fully or partially with a wild type or two different mutant Sec tRNA transgenes, respectively. The mutant Sec tRNA transgenes involve mutations in the tRNA that prevent the addition of a methyl group, 2'-O-methylribose (designated Um34), at position 37 which is the wobble position of the anticodon. The absence of Um34 results in the loss of expression of stress-related selenoproteins, while the housekeeping selenoproteins are virtually completely expressed. We had shown earlier through rescue of the selenoprotein population in a standard knockout mouse using wild type and mutant transgenes that Um34 was involved in synthesis of stress-related selenoproteins. However, our ability now to target a specific organ or tissue will permit us to focus on the role of selenium and selenoproteins in a given cell type. It should be further noted that this novel regulation of protein expression occurred at the level of translation and manifested a tissue-specific pattern. We are also expanding our targeted knockout of selenoprotein expression in various tissues and organs and have shown unequivocally a role of selenoproteins in development and disease prevention. One of our major focuses in this area over the last year has been on the role of selenoproteins in immune function. We targeted the removal of selenoprotein expression in T-cells and observed about a 50% reduction in the CD8 population and that mIL-13 was enriched more than 55 fold in males and 80 fold in females of knockout mice. IgG2a was increased about two-fold in targeted mice. These studies show thus far that selenoproteins in T-cells are associated with inflammatory allergic immune responses, have an essential role in the development/differentiation/maturation and function of T-cells and are indespensible factors in determining Th1 vs Th2 responses.