Selenium is recognized as a chemopreventive agent against cancer and heart disease, as an antiviral agent and it likely plays a role in delaying the aging process and in delaying the progression of AIDS in HIV positive patients. The beneficial effects of this element are due, at least in part, to its presence in selenoproteins as the amino acid, selenocysteine (Sec). Our program focuses on the means by which Sec is incorporated into protein, the role of specific selenoproteins in human health and the role of selenium in HIV infection. As Sec tRNA is the key molecule in selenoprotein biosynthesis, we have produced transgenic mice encoding from 2 to 20 wild type or 2 to 40 isopentenyladenosine (i6A) mutant Sec tRNA transgenes to examine the effect of their expression on selenium metabolism. Over-expression of wild type Sec tRNA does not affect selenoprotein synthesis demonstrating that Sec tRNA is not limiting in protein translation. Expression of i6A mutant Sec tRNA, however, selectively inhibits endogenous Sec tRNA maturation and translation of specific selenoproteins. DNA microarray analysis of the global effects of selenoprotein deficiency in these i6A mutant Sec tRNA mice show that many other enzymes involved in redox reactions are substantially reduced. These data suggest an intracellular connection between selenoprotein expression and other enzymes involved in redox reactions. In addition, we participated (with MJB, see collaborators) in identifying the specific elongation factor that is responsible for binding with selenocysteyl-tRNA for insertion of Sec into protein. In our studies on examining the role of specific selenoproteins in human health, we found that a 15 kDa selenoprotein (Sep15) is localized on chromosome 1p31 at a site commonly mutated or deleted in human cancers. The gene spans 51 kb consisting of 5 exons and 4 introns. It exists as two alleles in the human population that differ by two single nucleotide polymorphic sites. One of the polymorphisms appears to require higher levels of selenium for Sep15 expression. Sep15 gene expression occurs in highest levels in prostate, liver, testes, kidney and brain and the level of expression was reduced dramatically in the corresponding malignant tissues or cells. Each of these observations suggests that Sep15 has a role in cancer development. Interestingly, Sep15 exists in the endoplasmic reticulum of mammalian cells in a complex with UDP-glucose:glycoprotein glucosyltransferase suggesting a role of selenium in quality control of protein folding. In a related study on the effects of selenium on human health, we have examined the means by which HIV infection and the subsequent development of AIDS results in selenium and glutathione peroxidase (a selenoprotein) deficiency in afflicted patients. Rhesus monkeys that are inoculated with simian immunodeficiency virus become characteristically selenium and glutathione peroxidase deficient in the progression of the resulting disease as is found in HIV positive patients. This observation provides us with a model for studying selenium deficiency associated with AIDS. We previously observed that HIV infected cells do not label the selenoprotein population as efficiently as uninfected control cells. We have completely ruled out a role of the HIV TAT protein in this process. However, we observed that induction of HIV expression in chronically infected HIV cells results in the distribution of the Sec tRNA isoforms that is normally seen in selenium deficient tissues, while the Sec tRNA distribution in the corresponding uninduced cells is like that observed in selenium sufficient tissues. Thus, the Sec tRNA population appears to have a direct role in selenium deficiency in this disease. To better understand the role of tRNA in ribosomal frameshifting in HIV and other mammalian retroviruses, we examined the ability of phenylalanine tRNA lacking the highly modified wybutoxosine base on the 3 side of its anticodon to stimulate the frameshift event. This hypomodified tRNA dramatically enhances frameshifting suggesting an alternative avenue for inhibiting expression of some retroviruses by maintaining fully modified tRNAs intracellularly.
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