Selenoprotein Sep15: Functional analysis and role in quality control in the endoplasmic reticulum

Selenium is an essential trace element that is incorporated into proteins in the form of selenocysteine, the 21st natural amino acid which is encoded by the UGA codon. This rare amino acid is often present at the active centers of selenium-containing enzymes. One such protein, the 15-kDa selenoprotein (Sep15), has been recently reported to reside in the endoplasmic reticulum (ER) of eukaryotic cells and occur in a complex with UDP-glucose:glycoprotein glucosyltransferase (UGGT). UGGT is an ER chaperone and a component of the quality control machinery in the ER that assists in maturation of N-linked glycoproteins. Association of Sep15 with UGGT suggests that Sep15 may also be involved in assessing the structural fidelity of glycoproteins; however, the exact role of Sep15 in this process is poorly understood. In this study, we characterized interactions between members of the Sep15 and UGGT protein families. We found that Sep15 and UGGT form a tight 1:1 complex, and their interaction is mediated by a cysteine-rich domain of Sep15. Mutation study of conserved cysteine residues showed that structural integrity of this domain is required for Sep15 to form a complex with UGGT. Next, we employed nuclear magnetic resonance (NMR) to solve the structures of Sep15 and its distant homolog selenoprotein SelM. The solution NMR structures revealed that Sep15 and SelM have a thioredoxin-like fold. In mammals, Sep15 expression was regulated by dietary selenium, and both decreased and increased expression of this selenoprotein altered cellular redox homeostasis. The presence of thioredoxin-fold, together with the measured redox potential, suggested that Sep15 may catalyze reduction or isomerization of disulfide bonds in secreted proteins. Finally, to address the role of Sep15 in protein folding, we analyzed whether expression of Sep15 is regulated by unfolded protein response in the ER. We found that Sep15 expression is affected by pharmacological ER stress, and its expression levels depend on the mechanism by which ER stress is induced. However, Sep15 deficiency did not itself result in accumulation of unfolded proteins suggesting that Sep15 may assist in maturation of a restricted group of N-linked glycoproteins and that other mechanisms may compensate for Sep15 function