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New insights into how selenoprotein mRNAs elude nonsense mediated decay
Author(s) -
Berry Marla J,
Hoffmann Peter,
SmallHoward Andrea,
Jesus Lucia,
Forry Erin,
Stoytcheva Zoia
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a1473-d
Subject(s) - selenocysteine , selenoprotein , nonsense mediated decay , biology , microbiology and biotechnology , immunoprecipitation , nonsense , translation (biology) , gene , genetics , chemistry , messenger rna , rna splicing , biochemistry , rna , glutathione , glutathione peroxidase , cysteine , enzyme
Selenoproteins contain the unusual amino acid, selenocysteine, encoded by UGA. Use of UGA as a sense codon presents challenges to both the translation machinery and circumventing the nonsense mediated decay pathway. This is particularly perplexing when considering mRNAs containing as many as 18 UGA codons, as in the Xenopus selenoprotein P gene. To gain insights into how the cellular machinery has evolved to face the challenges resulting from UGA recoding, we investigated the intermolecular interactions and subcellular localization of the factors implicated in selenocysteine incorporation. Putative NLS and NES sequences were identified in several of these proteins. We epitope tagged each of these factors and carried out expression by transient transfection, coimmunoprecipitation and immunofluorescent localization studies, and heterokaryon assays. Our studies reveal significant protein‐protein interactions among all of these factors, and identify a coordinated series of associations and dissociations, resulting in the formation of several distinct complexes. Several of the factors undergo nucleocytoplasmic shuttling, cotransporting other factors. Finally, we show that overexpression of some of the factors increases levels of selenoprotein mRNAs when expressed from intron‐containing but not from intronless constructs, a hallmark of the nonsense mediated decay pathway. Our findings suggest that supra‐molecular complexes mediating selenocysteine incorporation assemble on selenoprotein mRNAs in the nucleus, allowing these mRNAs to circumvent nonsense mediated decay.