Selenoproteins are rare proteins among all kingdoms of life containing the 21 amino acid, selenocysteine. Selenocysteine resembles cysteine, differing only by the substitution of selenium for sulfur. Yet the actual advantage of selenolate- versus thiolate-based catalysis has remained enigmatic, as most of the known selenoproteins also exist as cysteine-containing homologs. Here, we demonstrate that selenolate-based catalysis of the essential mammalian selenoprotein GPX4 is unexpectedly dispensable for normal embryogenesis. Yet the survival of a specific type of interneurons emerges to exclusively depend on selenocysteine-containing GPX4, thereby preventing fatal epileptic seizures. Mechanistically, selenocysteine utilization by GPX4 confers exquisite resistance to irreversible overoxidation as cells expressing a cysteine variant are highly sensitive toward peroxide-induced ferroptosis. Remarkably, concomitant deletion of all selenoproteins in Gpx4 cells revealed that selenoproteins are dispensable for cell viability provided partial GPX4 activity is retained. Conclusively, 200 years after its discovery, a specific and indispensable role for selenium is provided.

Base 37 in tRNA, 3′-adjacent to the anticodon, is occupied by a purine base that is thought to stabilize codon recognition by stacking interactions on the first Watson-Crick base pair. If the first codon position forms an A.U or U.A base pair, the purine is likely further modified in all domains of life. One of the first base modifications found in tRNA is N6-isopentenyl adenosine (i6A) present in a fraction of tRNAs in bacteria and eukaryotes, which can be further modified to 2-methyl-thio-N6-isopentenyladenosine (ms2i6A) in a subset of tRNAs. Homologous tRNA isopentenyl transferase enzymes have been identified in bacteria (MiaA), yeast (Mod5, Tit1), roundworm (GRO-1), and mammals (TRIT1). In eukaryotes, isopentenylation of cytoplasmic and mitochondrial tRNAs is mediated by products of the same gene. Accordingly, a patient with homozygous mutations in TRIT1 has mitochondrial disease. The role of i6A in a subset of tRNAs in gene expression has been linked with translational fidelity, speed of translation, skewed gene expression, and non-sense suppression. This review will not cover the action of i6A as a cytokinin in plants or the potential function of Mod5 as a prion in yeast.

Selenocysteine is the 21st proteinogenic amino acid in mammals. The human genome contains 25 genes encoding selenoproteins, and their significance for human health is increasingly recognized through the identification of patients with inborn errors in selenoprotein biosynthetic factors or in individual selenoproteins. Mutations in selenoprotein N (SEPN1) lead to a spectrum of disorders collectively called SEPN1-related myopathy, and mutations in glutathione peroxidase 4 (GPX4) cause respiratory failure and bone defects, and mutations in thioredoxin reductase 2 (TXNRD2) are associated with familial glucocorticoid deficiency. Pathogenic mutations in selenocysteine synthase (SEPSECS) cause neurodevelopmental disorders, but also other factors epistatic to selenoprotein biosynthesis, such as SECIS-binding protein 2 (SECISBP2) and tRNA, are known to cause complex disorders. Mutations in the latter 2 genes involve impaired metabolism and action of thyroid hormones, which lead to delayed bone growth and maturation. Mutations in SECISBP2 sometimes affect nervous system development, muscle, inner ear, skin, and immune system function, underlining the significance of selenoproteins for the organism. Mouse models helped to delineate the functions of selenoproteins and explain pathomechanisms. For brevity, this review is focused on human genetic disorders associated with selenoprotein deficiency and only briefly touches on health effects of nutritional selenium deficiency.-Schweizer, U., Fradejas-Villar, N. Why 21? The significance of selenoproteins for human health revealed by inborn errors of metabolism.

Dual-assignment of codons as termination and elongation codons is used to expand the genetic code. In mammals, UGA can be reassigned to selenocysteine during translation of selenoproteins by a mechanism involving a 3΄ untranslated region (UTR) selenocysteine insertion sequence (SECIS) and the SECIS-binding protein Secisbp2. Here, we present data from ribosome profiling, RNA-Seq and mRNA half-life measurements that support distinct roles for Secisbp2 in UGA-redefinition and mRNA stability. Conditional deletions of the Secisbp2 and Trsp (tRNASec) genes in mouse liver were compared to determine if the effects of Secisbp2 loss on selenoprotein synthesis could be attributed entirely to the inability to incorporate Sec. As expected, tRNASec depletion resulted in loss of ribosome density downstream of all UGA-Sec codons. In contrast, the absence of Secisbp2 resulted in variable effects on ribosome density downstream of UGA-Sec codons that demonstrate gene-specific differences in Sec incorporation. For several selenoproteins in which loss of Secisbp2 resulted in greatly diminished mRNA levels, translational activity and Sec incorporation efficiency were shown to be unaffected on the remaining RNA. Collectively, these results demonstrate that Secisbp2 is not strictly required for Sec incorporation and has a distinct role in stabilizing mRNAs that can be separated from its effects on UGA-redefinition.

Recoding of UGA codons as selenocysteine (Sec) codons in selenoproteins depends on a selenocysteine insertion sequence (SECIS) in the 3-UTR of mRNAs of eukaryotic selenoproteins. SECIS-binding protein 2 (SECISBP2) increases the efficiency of this process. Pathogenic mutations in SECISBP2 reduce selenoprotein expression and lead to phenotypes associated with the reduction of deiodinase activities and selenoprotein N expression in humans. Two functions have been ascribed to SECISBP2: binding of SECIS elements in selenoprotein mRNAs and facilitation of co-translational Sec insertion. To separately probe both functions, we established here two mouse models carrying two pathogenic missense mutations in Secisbp2 previously identified in patients. We found that the C696R substitution in the RNA-binding domain abrogates SECIS binding and does not support selenoprotein translation above the level of a complete Secisbp2 null mutation. The R543Q missense substitution located in the selenocysteine insertion domain resulted in residual activity and caused reduced selenoprotein translation, as demonstrated by ribosomal profiling to determine the impact on UGA recoding in individual selenoproteins. We found, however, that the R543Q variant is thermally unstable in vitro and completely degraded in the mouse liver in vivo, while being partially functional in the brain. The moderate impairment of selenoprotein expression in neurons led to astrogliosis and transcriptional induction of genes associated with immune responses. We conclude that differential SECISBP2 protein stability in individual cell types may dictate clinical phenotypes to a much greater extent than molecular interactions involving a mutated amino acid in SECISBP2. This work was supported by Deutsche Forschungsgemeinschaft Grants Schw914/2 and Schw914/5 and Universitätsklinikum Bonn. The authors declare that they have no conflicts of interest with the contents of this article. This article was selected as one of our Editors' Picks. This article contains Table S1 and Figs. S1-S3. Raw and processed original data have been deposited in the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) under accession number GSE119681. 1 Supported by a BONFOR SciMed scholarship of the medical faculty.