Alternative transcripts and 3'UTR elements govern the incorporation of selenocysteine into selenoprotein S.
Alternative transcripts and 3'UTR elements govern the incorporation of selenocysteine into selenoprotein S.
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DOI：
10.1371/journal.pone.0062102
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Driscoll DM
中科院分区：
文献类型：
--
作者：
Bubenik JL;Miniard AC;Driscoll DM
硒蛋白S （Selenoprotein S, SelS）是一种189个氨基酸的跨膜蛋白，在未折叠蛋白反应中起重要作用，但尚未明确。有人提出，SelS可能具有还原酶的功能，其第二个硒化半胱氨酸（Sec188）残基参与了与半胱氨酸（Cys174）的硒化硫键。Sec与SelS的共译结合依赖于UGA密码子的重新编码，这需要在转录物的3'UTR中有硒代半胱氨酸插入序列（SECIS）元件。在这里，我们确定了调节SelS表达的多种机制。人类SelS基因编码两种转录本（变体1和变体2），它们的3'UTR序列不同，这是由于另一种剪接事件将SECIS元素从变体1转录本中移除。这两种转录本在人类细胞系中广泛表达，其中含有secis的变体2 mRNA更丰富。体外实验表明，变异1 3'UTR不允许读取UGA/Sec密码子。因此，该转录本将产生一个不含Sec的截断蛋白，并且不能形成硫代硒键。虽然变体2.3 ' utr确实支持Sec插入，但其活动较弱。生物信息学分析揭示了两个高度保守的茎环结构，一个位于变体23 ' utr的近端，另一个位于SECIS元件的下游。近端茎环在原生环境下促进Sec插入，但在异源mRNA中远离UGA/Sec密码子时则不会。相反，SECIS元件下游的140个核苷酸抑制Sec插入。我们还发现，除了已知的内质网定位外，内源性SelS在核周斑点处富集。我们的研究结果表明，内源性SelS的表达比以前认为的更复杂，这对过去和未来对该蛋白功能的研究具有重要意义。
Selenoprotein S (SelS) is a 189 amino acid trans-membrane protein that plays an important yet undefined role in the unfolded protein response. It has been proposed that SelS may function as a reductase, with the penultimate selenocysteine (Sec188) residue participating in a selenosulfide bond with cysteine (Cys174). Cotranslational incorporation of Sec into SelS depends on the recoding of the UGA codon, which requires a Selenocysteine Insertion Sequence (SECIS) element in the 3′UTR of the transcript. Here we identify multiple mechanisms that regulate the expression of SelS. The human SelS gene encodes two transcripts (variants 1 and 2), which differ in their 3′UTR sequences due to an alternative splicing event that removes the SECIS element from the variant 1 transcript. Both transcripts are widely expressed in human cell lines, with the SECIS-containing variant 2 mRNA being more abundant. In vitro experiments demonstrate that the variant 1 3′UTR does not allow readthrough of the UGA/Sec codon. Thus, this transcript would produce a truncated protein that does not contain Sec and cannot make the selenosulfide bond. While the variant 2 3′UTR does support Sec insertion, its activity is weak. Bioinformatic analysis revealed two highly conserved stem-loop structures, one in the proximal part of the variant 2 3′UTR and the other immediately downstream of the SECIS element. The proximal stem-loop promotes Sec insertion in the native context but not when positioned far from the UGA/Sec codon in a heterologous mRNA. In contrast, the 140 nucleotides downstream of the SECIS element inhibit Sec insertion. We also show that endogenous SelS is enriched at perinuclear speckles, in addition to its known localization in the endoplasmic reticulum. Our results suggest the expression of endogenous SelS is more complex than previously appreciated, which has implications for past and future studies on the function of this protein.
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