Selenium is an essential trace element that is incorporated into 25 human proteins as the amino acid selenocysteine (Sec). The proteins that contain Sec (selenoproteins) are essential for many cellular functions including combatting oxidative stress, thyroid hormone production and protein folding. Sec is incorporated at specific UGA codons that would otherwise signal translation termination. A specialized set of factors are known to be required for Sec incorporation: a specialized elongation factor that delivers the Sec-tRNASec to the ribosome and unique RNA binding proteins that bind to a Sec insertion sequence (SECIS) in selenoprotein mRNA 3' UTRs. This SECIS-protein complex signals the ribosome to incorporate Sec instead of translation termination. Our prior work has provided molecular characterization of each of the required factors, but the mechanism by which they interact with each other and other cellular components to allow Sec incorporation remains unknown. In addition, we provide preliminary evidence that the processive incorporation of 10 Sec residues into the selenium transport protein Selenoprotein P (SELENOP) requires a unique mechanism and additional factors. The overall goals for this proposal are to determine the mechanism by which SECIS binding proteins promote single and multiple Sec incorporation events. All vertebrates possess two SECIS binding proteins encoded by separate genes: SECISBP2 (SBP2) and SECISBP2L. While the mechanism of action for SBP2 is coming into focus, the role for SECISBP2L in Sec incorporation has not been deciphered. Our preliminary data shows that SECISBP2L is essential for the processive incorporation of Sec into Selenoprotein P. As such, we have established three model systems to study the synthesis of SELENOP: in vitro translation, expression in transfected mammalian cells and a zebrafish system that will allow unprecedented access to the role of selenoprotein function during development. These are also leveraged and combined with structural biology and transcriptomics to determine how synthesis of the entire selenoproteome is regulated by SECIS binding proteins. In this proposal we propose to 1) Decipher the mechanism by which SECIS elements and SECIS binding proteins enable processive Sec incorporation into the selenium transport protein, SELENOP; 2) Utilize a zebrafish model system to determine the function of SECISBP2L and the mechanism of SELENOP synthesis in vivo; 3) Determine the molecular basis for differential selenoprotein expression. The successful completion of these aims will bring us significantly closer to our long term goal of developing reagents that will permit selective activation or inhibition of selenoprotein synthesis in vivo.

Public Health Relevance

of this project to public health lies in the importance of dietary selenium for the prevention of cancer, male infertility immune disorders, hypothyroidism and heart disease. As a trace element that is essential for life, it is critical to understand how selenium biology intersects with human health and disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM077073-16
Application #
10102132
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Reddy, Michael K
Project Start
2006-01-06
Project End
2023-01-31
Budget Start
2021-02-01
Budget End
2022-01-31
Support Year
16
Fiscal Year
2021
Total Cost
Indirect Cost
Name
Rbhs-Robert Wood Johnson Medical School
Department
Biochemistry
Type
Schools of Medicine
DUNS #
078795875
City
Piscataway
State
NJ
Country
United States
Zip Code
08854
Shetty, Sumangala; Marsicano, John R; Copeland, Paul R (2018) Uptake and Utilization of Selenium from Selenoprotein P. Biol Trace Elem Res 181:54-61
Carlson, Bradley A; Lee, Byeong Jae; Tsuji, Petra A et al. (2018) Selenocysteine tRNA[Ser]Sec, the Central Component of Selenoprotein Biosynthesis: Isolation, Identification, Modification, and Sequencing. Methods Mol Biol 1661:43-60
Shetty, Sumangala P; Copeland, Paul R (2018) The Selenium Transport Protein, Selenoprotein P, Requires Coding Sequence Determinants to Promote Efficient Selenocysteine Incorporation. J Mol Biol 430:5217-5232
Shetty, Sumangala; Copeland, Paul R (2018) Molecular mechanism of selenoprotein P synthesis. Biochim Biophys Acta Gen Subj :
Carlson, Bradley A; Gupta, Nirupama; Pinkerton, Mark H et al. (2017) The utilization of selenocysteine-tRNA[Ser]Sec isoforms is regulated in part at the level of translation in vitro. Translation (Austin) 5:e1314240
Dobosz-Bartoszek, Malgorzata; Pinkerton, Mark H; Otwinowski, Zbyszek et al. (2016) Crystal structures of the human elongation factor eEFSec suggest a non-canonical mechanism for selenocysteine incorporation. Nat Commun 7:12941
Dubey, Aditi; Copeland, Paul R (2016) The Selenocysteine-Specific Elongation Factor Contains Unique Sequences That Are Required for Both Nuclear Export and Selenocysteine Incorporation. PLoS One 11:e0165642
Shetty, Sumangala P; Copeland, Paul R (2015) Selenocysteine incorporation: A trump card in the game of mRNA decay. Biochimie 114:97-101
French, Rachel L; Gupta, Nirupama; Copeland, Paul R et al. (2014) Structural asymmetry of the terminal catalytic complex in selenocysteine synthesis. J Biol Chem 289:28783-94
Shetty, Sumangala P; Shah, Ravi; Copeland, Paul R (2014) Regulation of selenocysteine incorporation into the selenium transport protein, selenoprotein P. J Biol Chem 289:25317-26

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