Regulation of Selenoprotein Synthesis by SECIS-binding Proteins

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DESCRIPTION (provided by applicant): Selenium is an essential micronutrient that exerts many important health benefits. The element is incorporated into selenoproteins as selenocysteine (Sec), the 21st amino acid. The mammalian selenoproteins perform important functions in anti-oxidant defense, thyroid hormone metabolism, male reproduction, and development. Sec is encoded by a UGA codon, which is normally read as a stop codon. The receding of UGA as Sec requires a Sec Insertion Sequence (SECIS) element in the 3' untranslated region of the selenoprotein mRNA. We previously identified two SECIS-binding proteins, SECIS Binding Protein 2 (SBP2) and ribosomal protein L30, which play critical roles in the receding mechanism. Although much progress has been made in understanding the Sec incorporation machinery, less is known about the regulation of this pathway. During selenium deficiency, the element is preferentially utilized in the brain and endocrine organs. There is also a hierarchy of expression of individual selenoproteins. Even under selenium adequate conditions, certain selenoproteins are essential for life and health whereas others are not. The central hypothesis of our proposal is that cis-acting sequences in the SECIS recruit multiple trans-acting factors, which prioritize the utilization of selenium is a tissue-specific and selenoprotein-dependent manner. We recently discovered two additional SECIS-binding proteins: a 110 kDa protein which was purified by RNA affinity chromatography and identified as nucleolin by mass spectrometry analysis, and an unknown protein of - 45 kDa, referred to here as SBP45. Unlike SBP2 and L30, nucleolin and SBP45 can distinguish between selenoprotein mRNAs. The two proteins have distinct binding specificities, with nucleolin preferentially binding to SECIS elements from selenoproteins that are essential for normal health and development. We also show that nucleolin specifically regulates UGA receding in vitro. In this project, we propose to use a variety of biochemical, cell biology, and molecular biology approaches to: 1) identify functionally important interactions between nucleolin and the SECIS element; 2) elucidate the function of nucleolin in regulating selenoprotein mRNA translation, and 3) identify SBP45 and determine its function in regulating selenoprotein expression. The successful pursuit of these aims may identify limiting factors and regulatory pathways that could be used therapeutically to modulate selenoprotein expression in vivo.