Project Details
Description
Abstract
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. 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. We propose to marry
our vast library of molecular tools from prior work with new model systems including the first use of a
genetically modified zebrafish system to study Sec incorporation as well as CRISPR/Cas9 edited cell lines.
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
production of a specific 50 kDa selenoprotein, likely SEPHS2, during early development. 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)
Determine the mechanism of SBP2-independent Sec incorporation and the role of SECIS binding proteins
in responding to cellular stress; 3) Utilize a zebrafish model system to determine the function of
SECISBP2L and the regulation of selenoprotein synthesis by oxidative stress. 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.
Status | Finished |
---|---|
Effective start/end date | 1/6/06 → 1/31/23 |
Funding
- National Institute of General Medical Sciences: $259,002.00
- National Institute of General Medical Sciences: $24,000.00
- National Institute of General Medical Sciences: $144,013.00
- National Institute of General Medical Sciences: $314,475.00
- National Institute of General Medical Sciences: $10,205.00
- National Institute of General Medical Sciences: $40,494.00
- National Institute of General Medical Sciences: $318,834.00
- National Institute of General Medical Sciences: $291,980.00
- National Institute of General Medical Sciences: $7,110.00
- National Institute of General Medical Sciences: $258,790.00
- National Institute of General Medical Sciences: $7,020.00
- National Institute of General Medical Sciences: $291,865.00
- National Institute of General Medical Sciences: $266,840.00
- National Institute of General Medical Sciences: $336,363.00
- National Institute of General Medical Sciences: $34,014.00
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