Similar Datasets
Project description:Mistranslation and developmental defects in an allelic series of tRNA isodecoder deletions [Ribo-seq]
| PRJNA944706 | ENA
Project description:The number of tRNA isodecoders has increased dramatically in mammals, but the specific molecular and physiological reasons for this expansion remain elusive. To address this fundamental question we used CRISPR editing to knockout the seven-membered phenylalanine tRNA gene family in mice, both individually and combinatorially. Using ATAC-Seq, RNA-seq and proteomics we observed distinct molecular consequences of individual tRNA deletions. We show that tRNA-Phe-1-1 is required for neuronal function and its loss is partially compensated by increased expression of other tRNAs but results in mistranslation. In contrast, the other tRNA-Phe isodecoder genes buffer the loss of each of the remaining six tRNA-Phe genes. In the tRNA-Phe gene family, the expression of at least six tRNA-Phe alleles is required for embryonic viability and tRNA-Phe-1-1 is most important for development and survival. Our results reveal that the multi-copy configuration of tRNA genes is required to buffer translation and viability in mammals.
2023-03-05 | GSE200783 | GEO
Project description:The number of tRNA isodecoders has increased dramatically in mammals, but the specific molecular and physiological reasons for this expansion remain elusive. To address this fundamental question we used CRISPR editing to knockout the seven-membered phenylalanine tRNA gene family in mice, both individually and combinatorially. Using ATAC-Seq, RNA-seq and proteomics we observed distinct molecular consequences of individual tRNA deletions. We show that tRNA-Phe-1-1 is required for neuronal function and its loss is partially compensated by increased expression of other tRNAs but results in mistranslation. In contrast, the other tRNA-Phe isodecoder genes buffer the loss of each of the remaining six tRNA-Phe genes. In the tRNA-Phe gene family, the expression of at least six tRNA-Phe alleles is required for embryonic viability and tRNA-Phe-1-1 is most important for development and survival. Our results reveal that the multi-copy configuration of tRNA genes is required to buffer translation and viability in mammals.
2023-03-05 | GSE223662 | GEO
Project description:Mistranslation, the mis-incorporation of an amino acid not specified by the “standard” genetic code, occurs in all cells. tRNA variants that increase mistranslation arise spontaneously and engineered tRNAs can achieve mistranslation frequencies approaching 10% in yeast and bacteria. The goal of this study was to detect mistranslation from two different tRNA variants. The first variant, tRNA-Pro-G3:U70, has a mutation in its acceptor stem creating a G3:U70 base pair which is the key identity element for the alanine tRNA synthetase. This tRNA should be charged with alanine and mis-incorporate alanine at proline codons. The second variant, tRNA-Ser-UCU,G26A, is a serine tRNA with an arginine anticodon and a G26A secondary mutation to dampen function and prevent lethal levels of mistranslation. This tRNA should mis-incorporate serine at arginine codons.
2021-05-17 | PXD025934 | Pride
Project description:Orofacial clefts are the most common form of congenital craniofacial malformations worldwide. The etiology of these birth defects is multifactorial, involving genetic and environmental factors. In most cases, however, the underlying causes remain unexplained, precluding molecular understanding of disease mechanisms. Here, we integrated genome-wide association data, targeted re-sequencing of case and control cohorts, cell type-specific epigenomic profiling, and genome architecture analyses, to functionally and molecularly dissect a genomic locus associated with an increased risk of non-syndromic orofacial cleft. We found that common and rare risk variants associated with orofacial cleft intersect with a conserved enhancer (e2p24.2) that becomes activated in cranial neural crest cells—the embryonic cell type responsible for sculpting the craniofacial complex. We mapped e2p24.2 long-range interactions to a topologically associated domain harboring MYCN and DDX1 and demonstrated that both MYCN and DDX1 are required for craniofacial development in chicken embryos. We found that e2p24.2 regulates the expression of MYCN, but not DDX1, in cranial neural crest cells. In turn, DDX1 is a target of the MYC family of transcription factors and a component of the tRNA splicing complex. The loss of DDX1 in cranial neural crest cells resulted in the accumulation of unspliced tRNA fragments, depletion of the mature pool of intron-containing tRNAs, and ribosome stalling at codons decoded by these tRNAs. These effects were accompanied by defects in both global protein synthesis and cranial neural crest cell migration. We further showed that the induction of tRNA fragments is sufficient to disrupt craniofacial development. Together, these results uncovered a molecular mechanism in which impaired tRNA splicing affects neural crest and craniofacial development and positioned MYCN, DDX1, and tRNA processing defects as risk factors in the pathogenesis of orofacial clefts.
2025-04-17 | GSE269561 | GEO
Project description:Transfer RNAs (tRNAs) maintain translational fidelity through strict charging by their cognate aminoacyl-tRNA synthetase and codon:anticodon base pairing with the mRNA at the ribosome. Mistranslation occurs when an amino acid not specified by the genetic code is incorporated into a protein. Since alanyl-tRNA synthetase uniquely recognizes a G3:U70 base pair in alanine tRNAs and the anticodon plays no role in charging, alanine tRNA variants with anticodon mutations have the potential to mistranslate alanine. Our goal was to quantify mis-incorporation of alanine into proteins in Saccharomyces cerevisiae strains expressing one of 57 different alanine tRNA anticodon variants. Using mass spectrometry, we observed mistranslation for 45 of the variants when expressed on single-copy plasmids.
2023-10-02 | PXD038242 | Pride
Project description:High-fidelity translation and a strictly accurate proteome were originally assumed as essential to life and cellular viability. Yet recent studies in bacteria and eukaryotic model organisms suggest that proteome-wide mistranslation can provide selective advantages and is tolerated in the cell at higher levels than previously thought (one error in 6.9 × 10-4 in yeast) with a limited impact on phenotype. Previously, we selected a tRNAPro containing a single mutation that induces mistranslation with alanine at proline codons in yeast. Yeast tolerate the mistranslation by inducing a heat-shock response and through the action of the proteasome. Here we found a homologous human tRNAPro (G3:U70) mutant that is not aminoacylated with proline, but is an efficient alanine acceptor. In live human cells, we visualized mistranslation using a green fluorescent protein reporter that fluoresces in response to mistranslation at proline codons. In agreement with measurements in yeast, quantitation based on the GFP reporter suggested a mistranslation rate of up to 2-5% in HEK 293 cells. Our findings suggest a stress-dependent phenomenon where mistranslation levels increased during nutrient starvation. Human cells did not mount a detectable heat-shock response and tolerated this level of mistranslation without apparent impact on cell viability. Because humans encode ∼600 tRNA genes and the natural population has greater tRNA sequence diversity than previously appreciated, our data also demonstrate a cell-based screen with the potential to elucidate mutations in tRNAs that may contribute to or alleviate disease.
| S-EPMC6103679 | biostudies-literature
Project description:Mistranslation, the mis-incorporation of an amino acid not specified by the “standard” genetic code, occurs in all cells. tRNA variants that increase mistranslation arise spontaneously and engineered tRNAs can achieve mistranslation frequencies approaching 10% in yeast and bacteria. The goal of this study was to detect and quantify mistranslation from a serine tRNA variant with proline UGG anticodon and G26A secondary mutation engineered in yeast
2022-08-12 | PXD032063 | Pride
Project description:The number of tRNA isodecoders has increased dramatically in mammals, but the specific molecular and physiological reasons for this expansion remain elusive. To address this fundamental question we used CRISPR editing to knockout the seven-membered phenylalanine tRNA gene family in mice, both individually and combinatorially. Using ATAC-Seq, RNA-seq, ribo-profiling and proteomics we observed distinct molecular consequences of single tRNA deletions. We show that tRNA-Phe-1-1 is required for neuronal function and its loss is partially compensated by increased expression of other tRNAs but results in mistranslation. In contrast, the other tRNA-Phe isodecoder genes buffer the loss of each of the remaining six tRNA-Phe genes. In the tRNA-Phe gene family, the expression of at least six tRNA-Phe alleles is required for embryonic viability and tRNA-Phe-1-1 is most important for development and survival. Our results reveal that the multi-copy configuration of tRNA genes is required to buffer translation and viability in mammals.
| S-EPMC10113395 | biostudies-literature
Project description:Several classes of small noncoding RNAs are key players in cellular metabolism including mRNA decoding, RNA processing, and mRNA stability. Here we show that a tRNA(Asp) isodecoder, corresponding to a human tRNA-derived sequence, binds to an embedded Alu RNA element contained in the 3' UTR of the human aspartyl-tRNA synthetase mRNA. This interaction between two well-known classes of RNA molecules, tRNA and Alu RNA, is driven by an unexpected structural motif and induces a global rearrangement of the 3' UTR. Besides, this 3' UTR contains two functional polyadenylation signals. We propose a model where the tRNA/Alu interaction would modulate the accessibility of the two alternative polyadenylation sites and regulate the stability of the mRNA. This unique regulation mechanism would link gene expression to RNA polymerase III transcription and may have implications in a primate-specific signal pathway.
| S-EPMC3189049 | biostudies-literature