Effects of tRNA[Ser]Sec anticodon stem loop modifications on selenoprotein expression
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA762342 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR15844694.fastq.gz | Fastqsanger.gz | |||
| SRR15844695.fastq.gz | Fastqsanger.gz | |||
| SRR15844696.fastq.gz | Fastqsanger.gz | |||
| SRR15844697.fastq.gz | Fastqsanger.gz | |||
| SRR15844698.fastq.gz | Fastqsanger.gz |
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Project description:Expression of selenoproteins requires the co-translational incorporation of selenocysteine (Sec) in response to an in-frame UGA codon. The machinery of UGA/Sec re-coding is complex and many factors affect the hierarchy of expression among selenoproteins, including modification of tRNA[Ser]Sec. Its hyper-modification in the anticodon stem loop is influenced by selenium bioavailability, and a mutation in adenosine 37 (A37) that abrogates isopentenylation, has a profound effect on selenoprotein expression in mice. Patients with mutations in tRNA-isopentenyl-transferase (TRIT1) show a severe neurological disorder and hence we wondered whether mutations in TRIT1 negatively affected the expression of selenoproteins. Fibroblasts from a patient carrying a pathogenic R323Q mutation in TRIT1 in homozygosity did not show decreased selenoprotein expression, although recombinant TRIT1R323Q had significantly reduced activity in vitro towards anticodon stem-loop substrates. We thus engineered mice conditionally deficient in Trit1 in hepatocytes and neurons. Selenoprotein expression as assessed by western blotting, 75Se metabolic labeling, and ribosomal profiling was not decreased despite the general reduction of N6-isopentenyl-adenosine in tRNAs. We show that 5-methylcarboxymethylation and 2’O-methylation of U34 occur independently of isopentenylation of A37 in tRNA[Ser]Sec. Reanalyzing previously published ribosomal profiling datasets, we demonstrate that (i) failure of 5-carboxymethylation at U34 is associated with reduced expression of GPX1, but not GPX4, and that (ii) FTSJ1 is not the elusive U34-2’O-methyltransferase involved in the methylation of tRNA[Ser]Sec.
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Project description:Comparative analysis of gene expression in bone marrow-derived macrophages (BMDM) from trsp knockout mice (Trspfl/fl-LysM-Cre+/-) and Control (Trspfl/fl-LysM-Cre-/-) mice. Selenium, a micronutrient whose deficiency in the diet causes immune dysfunction and inflammatory disorders, exerts its physiological effects partly in the form of selenium-containing proteins (selenoproteins). Incorporation of selenium into the amino acid selenocysteine (Sec), and subsequently into selenoproteins, is mediated by Sec tRNA[Ser]Sec. To identify macrophage-specific selenoprotein function, we generated mice with the Sec tRNA[Ser]Sec gene specifically deleted in myeloid cells. These mutant mice were devoid of the selenoproteome in macrophages, yet exhibited largely normal inflammatory responses. However, selenoprotein deficiency led to aberrant expression of extracellular matrix-related genes, and diminished migration of macrophages in a protein gel matrix. Therefore, selenium status may affect immune defense and tissue homeostasis through its effect on selenoprotein expression and the trafficking of tissue macrophages.
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Project description:Comparative analysis of gene expression in bone marrow-derived macrophages (BMDM) from trsp knockout mice (Trspfl/fl-LysM-Cre+/-) and Control (Trspfl/fl-LysM-Cre-/-) mice. Selenium, a micronutrient whose deficiency in the diet causes immune dysfunction and inflammatory disorders, exerts its physiological effects partly in the form of selenium-containing proteins (selenoproteins). Incorporation of selenium into the amino acid selenocysteine (Sec), and subsequently into selenoproteins, is mediated by Sec tRNA[Ser]Sec. To identify macrophage-specific selenoprotein function, we generated mice with the Sec tRNA[Ser]Sec gene specifically deleted in myeloid cells. These mutant mice were devoid of the selenoproteome in macrophages, yet exhibited largely normal inflammatory responses. However, selenoprotein deficiency led to aberrant expression of extracellular matrix-related genes, and diminished migration of macrophages in a protein gel matrix. Therefore, selenium status may affect immune defense and tissue homeostasis through its effect on selenoprotein expression and the trafficking of tissue macrophages. We have generated mice in which we have selectively removed the selenocysteine tRNA gene (trsp) in macrophages under the control of LysM-Cre promoter. Microarray analysis was performed on RNA samples taken from bone marrow-derived macrophages in knockout and control mice. 1. Control unstimulated 2. Knockout unstimulated 3. Control lipopolysaccharide (LPS) stimulated (4h) 4. Knockout LPS stimulated (4h). Three replicates for each condition. Thus, a total of 12 samples.
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2023-10-02 | PXD038242 | Pride
Project description:In the yeast genera Saccharomycopsis and Ascoidea, nuclear genes use a non-standard genetic code in which CUG codons are translated as serine instead of leucine, due to the presence of a tRNA-Ser with the unusual anticodon CAG. However, some species in this ‘CUG-Ser2’ clade also contain an ancestral tRNA-Leu gene with the same anticodon. One of these species, Ascoidea asiatica, has been shown to have a stochastic proteome in which proteins contain approximately 50% Ser and 50% Leu at CUG codon sites, whereas previously examined Saccharomycopsis species translate CUG only as Ser. Here, we investigated the presence, conservation, and possible functionality of the tRNA-Leu(CAG) gene in the genus Saccharomycopsis. We analyzed the genomes of 33 strains, including almost all known species of Saccharomycopsis, and found that most of them contain both tRNA-Ser(CAG) and tRNA-Leu(CAG) genes. The tRNA-Leu(CAG) gene is evolving faster than tRNA-Ser(CAG) and it has been lost in two species, S. microspora and S. synnaedendra. We deleted the single tRNA-Leu(CAG) gene in S. capsularis and found that it is not essential. Bioinformatic analysis suggested that some CUG codon sites in Saccharomycopsis species may be translated as Leu, specifically in genes with functions in meiosis or sporulation, but mass spectrometry of sporulating S. capsularis and S. fermentans cultures showed only CUG-Ser translation. Cloverleaf structures of tRNA-Leu(CAG) from all Saccharomycopsis species contain mutations that are likely to make them non-functional in translation, but the evolutionary conservation of the gene leads us to propose that it has been retained for an unknown non-translational role.
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Project description:In the yeast genera Saccharomycopsis and Ascoidea, nuclear genes use a non-standard genetic code in which CUG codons are translated as serine instead of leucine, due to the presence of a tRNA-Ser with the unusual anticodon CAG. However, some species in this ‘CUG-Ser2’ clade also contain an ancestral tRNA-Leu gene with the same anticodon. One of these species, Ascoidea asiatica, has been shown to have a stochastic proteome in which proteins contain approximately 50% Ser and 50% Leu at CUG codon sites, whereas previously examined Saccharomycopsis species translate CUG only as Ser. Here, we investigated the presence, conservation, and possible functionality of the tRNA-Leu(CAG) gene in the genus Saccharomycopsis. We analyzed the genomes of 33 strains, including almost all known species of Saccharomycopsis, and found that most of them contain both tRNA-Ser(CAG) and tRNA-Leu(CAG) genes. The tRNA-Leu(CAG) gene is evolving faster than tRNA-Ser(CAG) and it has been lost in two species, S. microspora and S. synnaedendra. We deleted the single tRNA-Leu(CAG) gene in S. capsularis and found that it is not essential. Bioinformatic analysis suggested that some CUG codon sites in Saccharomycopsis species may be translated as Leu, specifically in genes with functions in meiosis or sporulation, but mass spectrometry of sporulating S. capsularis and S. fermentans cultures showed only CUG-Ser translation. Cloverleaf structures of tRNA-Leu(CAG) from all Saccharomycopsis species contain mutations that are likely to make them non-functional in translation, but the evolutionary conservation of the gene leads us to propose that it has been retained for an unknown non-translational role.
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Project description:In the yeast genera Saccharomycopsis and Ascoidea, nuclear genes use a non-standard genetic code in which CUG codons are translated as serine instead of leucine, due to the presence of a tRNA-Ser with the unusual anticodon CAG. However, some species in this ‘CUG-Ser2’ clade also contain an ancestral tRNA-Leu gene with the same anticodon. One of these species, Ascoidea asiatica, has been shown to have a stochastic proteome in which proteins contain approximately 50% Ser and 50% Leu at CUG codon sites, whereas previously examined Saccharomycopsis species translate CUG only as Ser. Here, we investigated the presence, conservation, and possible functionality of the tRNA-Leu(CAG) gene in the genus Saccharomycopsis. We analyzed the genomes of 33 strains, including almost all known species of Saccharomycopsis, and found that most of them contain both tRNA-Ser(CAG) and tRNA-Leu(CAG) genes. The tRNA-Leu(CAG) gene is evolving faster than tRNA-Ser(CAG) and it has been lost in two species, S. microspora and S. synnaedendra. We deleted the single tRNA-Leu(CAG) gene in S. capsularis and found that it is not essential. Bioinformatic analysis suggested that some CUG codon sites in Saccharomycopsis species may be translated as Leu, specifically in genes with functions in meiosis or sporulation, but mass spectrometry of sporulating S. capsularis and S. fermentans cultures showed only CUG-Ser translation. Cloverleaf structures of tRNA-Leu(CAG) from all Saccharomycopsis species contain mutations that are likely to make them non-functional in translation, but the evolutionary conservation of the gene leads us to propose that it has been retained for an unknown non-translational role.
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Project description:In the yeast genera Saccharomycopsis and Ascoidea, nuclear genes use a non-standard genetic code in which CUG codons are translated as serine instead of leucine, due to the presence of a tRNA-Ser with the unusual anticodon CAG. However, some species in this ‘CUG-Ser2’ clade also contain an ancestral tRNA-Leu gene with the same anticodon. One of these species, Ascoidea asiatica, has been shown to have a stochastic proteome in which proteins contain approximately 50% Ser and 50% Leu at CUG codon sites, whereas previously examined Saccharomycopsis species translate CUG only as Ser. Here, we investigated the presence, conservation, and possible functionality of the tRNA-Leu(CAG) gene in the genus Saccharomycopsis. We analyzed the genomes of 33 strains, including almost all known species of Saccharomycopsis, and found that most of them contain both tRNA-Ser(CAG) and tRNA-Leu(CAG) genes. The tRNA-Leu(CAG) gene is evolving faster than tRNA-Ser(CAG) and it has been lost in two species, S. microspora and S. synnaedendra. We deleted the single tRNA-Leu(CAG) gene in S. capsularis and found that it is not essential. Bioinformatic analysis suggested that some CUG codon sites in Saccharomycopsis species may be translated as Leu, specifically in genes with functions in meiosis or sporulation, but mass spectrometry of sporulating S. capsularis and S. fermentans cultures showed only CUG-Ser translation. Cloverleaf structures of tRNA-Leu(CAG) from all Saccharomycopsis species contain mutations that are likely to make them non-functional in translation, but the evolutionary conservation of the gene leads us to propose that it has been retained for an unknown non-translational role.
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