Biology subjects

Moehring, A. J.

Publications and source records attributed to Moehring, A. J..

6 recordsLinked to original sources

bioRxiv2026

The role of the fruitless P3 transcript in development in Drosophila melanogaster

The generation of multiple transcripts from a single gene through alternative promoter usage and alternative splicing is a major source of functional diversity in eukaryotic genomes. The fruitless (fru) gene of Drosophila melanogaster is a classic example of transcript complexity, producing at least 15 transcript isoforms through the use of alternative promoters and terminal exons. While transcripts derived from the sex-specifically spliced P1 promoter have been extensively studied for their roles in sex determination and courtship behaviour, the functions of most non-sex-specific fru transcripts remain poorly understood. Previous studies suggested that transcripts originating from the P3 and/or P4 promoters may contribute to developmental viability, but their individual roles could not be resolved because available mutations disrupted multiple transcript classes simultaneously. Here, we used transcript-specific RNA interference (RNAi) and the Gal4/UAS system to investigate the function of fru P3 transcripts. Loss of P3 expression significantly increased developmental time to pupariation and resulted in complete lethality during pupal development despite normal survival to pupariation. Most mortality occurred during early and mid-pupal stages, indicating a critical requirement for P3-derived transcripts during metamorphosis. Notably, P3 knockdown did not phenocopy complete loss of fruitless, as knockdown individuals survived substantially later into development than fru null mutants, suggesting that additional fruitless transcript classes contribute essential functions during earlier developmental stages. Together, these findings establish an independent developmental role for the fru P3 transcript class and demonstrate that transcript-specific functions contribute substantially to the biological complexity of the fruitless locus.

genetics↗

bioRxiv2026

Adaptation of fluorescence rescue reporters of mistranslation for use in Saccharomyces cerevisiae

Specific mutations to tRNA genes can cause mistranslation - the incorporation of amino acids mismatched to the mRNA code during translation. The effects of mistranslation are hugely varied, depending on the form of mistranslation and various contextual factors. A key aspect of mutant tRNA mistranslation investigation is the reliable detection of mistranslation events. While mistranslation confirmation and quantification are commonly accomplished via mass spectrometry, others have devised a means of detecting mistranslation via fluorescent signal. Here, we successfully adapt this method for use in Saccharomyces cerevisiae, a model in which a great deal of mistranslation research is currently conducted. We demonstrate that mistranslation fluorescence rescue is detectable using both microscopy as well as fluorescent plate reader measurement, and that fluorescence intensity varies with the form of mistranslation assessed. Interestingly, the quantity of mistranslation measured by fluorescence rescue differs from measurements by standard mass spectrometry methods, potentially due to capturing a different timepoint of proteins that contain mistranslation errors. As such, this tool has high utility in detecting mutant tRNA mistranslation presence in yeast models, while its quantification of mistranslation frequency requires further investigation.

molecular biology↗

bioRxiv2026

Varied forms of mutant tRNA mistranslation produce distinct phenotypes across multiple model organisms

Mutant tRNA mistranslation is a phenomena in which specific tRNA gene mutations cause translational errors wherein amino acids incorporated during translation differ from those coded by mRNA sequences. The biological consequences of mutant tRNA mistranslation are highly complex and contextual, but are often deleterious. Importantly, many variables exist that are likely to affect the outcomes of any one mutant mistranslating tRNA; biochemical characteristics of the exchanged amino acids, frequency of use, identity of mistranslated products, and so on. Here, we generate a carefully curated array of tRNA mutants to assess which characteristics of mistranslating tRNA variants are most predictive of deleterious phenotypes. We find that toxicity often arises from mutant tRNAs that induce dramatic changes in biochemical properties between exchanged amino acids, as well as mistranslation that occurs more frequently. Importantly, exceptions are also observed to each of these general rules, implying instead that some deliriousness may arise from more granular, product-specific mechanisms. Additionally, quantification of mistranslation via mass spectrometry demonstrates a weak relationship between quantity of mistranslated products and severity of toxic phenotypes. Lastly, despite previous establishment of mutant mistranslating tRNA models in Drosophila melanogaster, incorporation of higher frequency mistranslating tRNA variants was largely unsuccessful, and incorporation of lower frequency mistranslating tRNA variants produces no detectable developmental phenotypes. These experiments support the notion that although general rules may be capable of reasonably predicting their consequences, each mistranslating tRNA variant warrants individual consideration and investigation for thorough understanding of its biological outcomes and precise mechanisms of toxicity.

genetics↗

bioRxiv2024

Mistranslating tRNA variants have anticodon- and sex-specific impacts on Drosophila melanogaster

Transfer RNAs (tRNAs) are vital in determining the specificity of translation. Mutations in tRNA genes can result in the misincorporation of amino acids into nascent polypeptides in a process known as mistranslation. Since mistranslation has different impacts, depending on the type of amino acid substitution, our goal here was to compare the impact of different mistranslating tRNASer variants on fly development, lifespan, and behaviour. We established two mistranslating fly lines, one with a tRNASer variant that misincorporates serine at valine codons (VS) and the other that misincorporates serine at threonine codons (TaS). While both mistranslating tRNAs increased development time and developmental lethality, the severity of the impacts differed depending on amino acid substitution and sex. The VS variant extended embryonic, larval, and pupal development whereas the TS only extended larval and pupal development. Females, but not males, containing either mistranslating tRNA presented with significantly more anatomical deformities than controls. Mistranslating females also experienced extended lifespan whereas mistranslating male lifespan was unaffected. In addition, mistranslating flies from both sexes showed improved locomotion as they aged, suggesting delayed neurodegeneration. Therefore, although mistranslation causes detrimental effects, we demonstrate that mistranslation also has positive effects on complex traits such as lifespan and locomotion. This has important implications for human health given the prevalence of tRNA variants in humans. PLAIN LANGUAGE SUMMARYMutant tRNA genes can cause mistranslation, the misincorporation of amino acids into proteins, and are associated with several human diseases. This study investigated the role of two tRNA variants that cause threonine-to-serine (TS) or valine-to-serine (VS) substitution. Both variants caused developmental delays and lethality in both sexes and increase prevalence of deformities in females. Surprisingly, female TS and VS flies experienced increased lifespan and mistranslating males and females showed improved locomotion. These results suggest that mistranslation has both positive and negative effects that depend on the tRNA variant and sex of the fly.

genetics↗

bioRxiv2024

Impact of tRNA-induced proline-to-serine mistranslation on the transcriptome of Drosophila melanogaster

Mistranslation is the misincorporation of an amino acid into a polypeptide. Mistranslation has diverse effects on multicellular eukaryotes and is implicated in several human diseases. In Drosophila melanogaster, a serine transfer RNA (tRNA) that misincorporates serine at proline codons (P[->]S) affects male and female flies differently. The mechanisms behind this discrepancy are currently unknown. Here, we compare the transcriptional response of male and female flies to P[->]S mistranslation to identify genes and cellular processes that underlie sex-specific differences. Both males and females downregulate genes associated with various metabolic processes in response to P[->]S mistranslation. Males downregulate genes associated with extracellular matrix organization and response to negative stimuli such as wounding, whereas females downregulate aerobic respiration and ATP synthesis genes. Both sexes upregulate genes associated with gametogenesis, but females also upregulate cell cycle and DNA repair genes. These observed differences in the transcriptional response of male and female flies to P[->]S mistranslation have important implications for the sex-specific impact of mistranslation on disease and tRNA therapeutics. ARTICLE SUMMARYProline-to-serine mistranslation affects male and female flies differently, but the mechanisms underlying this discrepancy are unknown. We present a transcriptomic analysis of male and female flies showing that mistranslation disrupts metabolic pathways and gametogenesis in both sexes, whereas processes such as DNA repair and cell cycle regulation are affected only in one sex. This is the first analysis that characterizes sex-specific effects of mistranslation and provides intriguing avenues for future research to understand how mistranslation affects males and females.

genetics↗

bioRxiv2021

A Novel Mistranslating tRNA Model in Drosophila melanogaster has Diverse, Sexually Dimorphic Effects

Transfer RNAs (tRNAs) are the adaptor molecules required for reading of the genetic code and the accurate production of proteins. tRNA variants can lead to genome-wide mistranslation, the misincorporation of amino acids not specified by the standard genetic code into nascent proteins. While genome sequencing has identified putative mistranslating tRNA variants in human populations, little is known regarding how mistranslation affects multicellular organisms. Here, we create a Drosophila melanogaster model for mistranslation by integrating a serine tRNA variant that mistranslates serine for proline (tRNASerUGG, G26A) into the fly genome. Using mass spectrometry, we find that tRNASerUGG, G26A misincorporates serine for proline at a frequency of [~] 0.6% per codon. We find that mistranslation extends development time and decreases the number of flies that reach adulthood. Adult flies containing tRNASerUGG, G26A present with more morphological deformities and worse climbing performance than flies expressing only wild type tRNA. Female flies with the serine tRNA variant have more deformities and experience a faster decline in climbing performance than males, suggesting sex-specific effects. This model will enable studies into the synergistic effects of mistranslating tRNA variants and disease-causing alleles.

molecular biology↗