Glutamine missense suppression transfer RNAs inhibit polyglutamine aggregation
Ontology highlight
ABSTRACT:
INSTRUMENT(S):
ORGANISM(S): Mus Musculus (mouse)
TISSUE(S): Stem Cell
DISEASE(S): Huntington Disease
SUBMITTER: 
Kyle Hoffman
LAB HEAD: Patrick O'Donoghue
PROVIDER: PXD053115 | Pride | 2025-01-15
REPOSITORIES: Pride
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Project description:Transfer RNAs (tRNAs) play an essential role in protein synthesis by linking the nucleic acid sequences of gene products to the amino acid sequences of proteins. Although only 32 tRNAs are needed to decode all 61 sense codons in the genetic code, there are > 400 functional tRNA genes in the humans. Adding to this diversity, there are many single nucleotide polymorphisms in tRNAs across our population, including anticodon variants that mistranslate the genetic code. In human genomes, we identified three alanine tRNA (tRNAAla) variants with non-synonymous anticodon mutations: tRNAAlaCGC G35T, tRNAAlaUGC G35A, and tRNAAlaAGC C36T. Since alanyl-tRNA synthetase (AlaRS) does not recognize the anticodon, we hypothesized that these human tRNAAla variants will mis-incorporate Ala at glutamate (Glu), valine (Val), and threonine (Thr) codons. We found that human cells expressing the naturally occurring tRNAAla variants were characterized by defects in protein production and cell growth. Using mass spectrometry, we confirmed and estimated Ala mis-incorporation levels at Glu (0.7%), Val (5%) and Thr (0.1%) codons. Although Ala mis-incorporation was higher at Val codons, cells mis-incorporating Ala at Glu codons had the most severe defect in protein production. Significant growth defects were observed in cells mistranslating Glu and Val codons, while the low level of mis-incorporation at Thr codons was well-tolerated in human cells. The data demonstrate the ability of natural human tRNAAla variants to generate mistranslation leading to a phenotypic response that depends on the nature of the amino acid replacement and the level of mis-incorporation.
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Project description:“Inaccurate” expression of the genetic code, also known as mistranslation, is an emerging paradigm in microbial studies. Growing evidence indicates that many microbial pathogens can deliberately mistranslate their genetic code to help invade a host or evade host immune responses. However, discovering new capacities for deliberate mistranslation remains a challenge because each group of pathogens typically employs a unique mistranslation mechanism. In this study, we address this problem by studying duplicated genes of aminoacyl-tRNA synthetases. Using bacterial prolyl-tRNA synthe¬tase (ProRS) genes as an example, we identify an anomalous ProRS isoform, ProRSx, and a corre¬sponding tRNA, tRNAProA, that are predominately found in plant pathogens from Streptomyces species. We then show that tRNAProA has an unusual hybrid structure that allows this tRNA to mistranslate alanine codons as proline. We finally provide biochemical, genetic, and mass-spectro-metric evidence that cells which express ProRSx and tRNAProA can translate GCU alanine codons both as alanine and proline. This dual use of alanine codons creates a hidden proteome diversity due to stochastic Ala→Pro mutations in protein sequences. Thus, we show that important plant pathogens are equipped with a tool to alter the identity of their sense codons. This finding reveals the first example of a natural tRNA synthetase/tRNA pair for dedicated mistranslation of sense codons.
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Project description:The correct coupling of amino acids with transfer RNAs (tRNAs) is vital for translating genetic information into functional proteins. Errors during this process lead to mistranslation, where a codon is translated using the wrong amino acid. While unregulated and prolonged mistranslation is often toxic, growing evidence suggests that organisms, from bacteria to humans, can induce and use mistranslation as a mechanism to overcome unfavorable environmental conditions. Most known cases of mistranslation are caused by translation factors with poor substrate specificity or when substrate discrimination is sensitive to molecular changes such as mutations or post-translational modifications. Here we report two novel families of tRNAs, encoded by bacteria from the Streptomyces and Kitasatospora genera, that adopted dual identities by integrating the anticodons AUU (for Asn) or AGU (for Thr) into the structure of a distinct proline tRNA. These tRNAs are typically encoded next to a full-length or truncated version of a distinct isoform of bacterial-type prolyl-tRNA synthetase. Using two protein reporters, we showed that these tRNAs translate asparagine and threonine codons with proline. Moreover, when expressed in Escherichia coli, the tRNAs cause varying growth defects due to global Asn-to-Pro and Thr-to-Pro mutations. Yet, proteome-wide substitutions of Asn with Pro induced by tRNA expression increased cell tolerance to the antibiotic carbenicillin, indicating that Pro mistranslation can be beneficial under certain conditions. Collectively, our results significantly expand the catalog of organisms known to possess dedicated mistranslation machinery and provide support for the concept that mistranslation is a mechanism for cellular resiliency against environmental stress.
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