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Ribosomal protein (RP) gene haploinsufficiency is a conserved form of ribosome dysfunction across species and underlies a class of disorders known as ribosomopathies. In Drosophila, RP gene haploinsufficiency manifests as the Minute phenotype, characterized by thinner and shorter mechanosensory bristles. The development of both bristles and proprioceptive campaniform sensilla (CS) is initiated by the bHLH proneural proteins Achaete (Ac) and Scute (Sc). By analysing genetic interactions between ac sc mutants and Minute mutants of varying severity, we identified a novel bristle-promoting effect that occurs only in the strongly affected Minutes in which the average bristle length is shorter than a threshold. This threshold-dependent effect also promotes ectopic CS formation in the strong Minutes. Transcriptomic analyses comparing the sensory organ - promoting and non-promoting Minutes revealed significant differences in stress-response pathways, including differentially elevated expression of the Xrp1-Irbp18 transcriptional dimer. Notably, mutation of Xrp1 suppresses the ectopic CS phenotype, indicating a positive regulatory role. These findings reveal a previously unrecognized threshold effect in RP gene haploinsufficiency, in which excessive Xrp1 activity promotes supernumerary sensory organ formation, suggesting a compensatory mechanism that modulates neurogenesis under severe ribosomal stress.

Drosophila melanogaster Nora virus (DmNV), a positive-sense single stranded RNA virus related to picornaviruses. Given its genetic and structural similarity to neurotropic picornaviruses, such as poliovirus, we sought to determine whether DmNV could be found within the head and brain of D. melanogaster. RNA was extracted from heads of chronically DmNV-infected stocks, as well as from uninfected controls, and assayed using reverse transcription-polymerase chain reaction (RT-PCR) for DmNV open reading frame 1 (ORF1). The results showed that DmNV genomic material can be isolated from the heads of DmNV-infected D. melanogaster, which suggests that the virus reaches the head during the course of infection. To determine whether DmNV infects the brain tissue itself, small-molecule RNA fluorescence in situ hybridization (smRNA FISH) experiments on whole brains dissected from DmNV-infected and uninfected D. melanogaster were done. The smRNA FISH detection method was validated by identifying DmNV RNA in gut tissue, but there was no evidence of DmNV localization in any brain specimens examined. These findings suggest an alternative explanation for why DmNV may be present in dissected head specimens. Additionally, we highlight the effectiveness of smRNA FISH as a highly specific and accessible method for detecting RNA viruses in Drosophila, offering an alternative to antibody-based or transgenic fluorescence approaches. Together, our results refine the understanding of DmNV tissue tropism and provide methodological insights for future studies using insect RNA viruses.

Angelman syndrome (AS) is a rare neurogenetic disorder characterized by developmental delay, speech impairment, ataxia, epilepsy, and in some cases hyperphagic feeding behavior. AS is caused by loss of function mutations, loss of expression, or maternal allele deletion of the E3 ubiquitin ligase UBE3A. Recent work has identified a connection between UBE3A and the mechanosensitive ion channel PIEZO2, raising the possibility that UBE3A may regulate PIEZO-dependent satiety signaling. In this study, we investigated the role of the Drosophila UBE3A ortholog, Dube3a, in Piezo-associated feeding behaviors. Single-cell RNA-sequencing data revealed overlapping expression of Dube3a and Piezo within crop and enterocyte populations of the gut, identifying a relevant cellular context for this pathway to occur. We developed a novel feeding assay using GFP-expressing yeast to quantify food intake and gut distention in vivo. Dube3a loss-of-function (Dube3a^15b) flies exhibited hyperphagia and gut distention nearly identical to Piezo knockout flies. Analysis of chromosomal deficiency lines spanning the Dube3a locus further supported a requirement for Dube3a in normal satiety signaling. Finally, biochemical analyses demonstrated that Dube3a knockdown results in decreased Piezo protein levels, consistent with an indirect regulatory relationship. Together, these findings identify Dube3a as a critical regulator of Piezo-dependent satiety pathways and suggest that dysregulation of mechanosensory signaling may contribute to hyperphagia observed in AS. Further work is needed to define the intermediate factors linking UBE3A activity to Piezo stability and function.

A conserved cohort of signalling pathways orchestrate development and adult homoeostasis. Deregulation of these pathways underlies many diseases. A key set of signals is the family of Wnt ligands. Members of this family are conserved, but a clear understanding of the unique and redundant roles is lacking. Previous efforts to study Wnt ligand function in Drosophila have been hampered by the difficulty of generating , functional transgenes. To address this, we have created a complete set of synthesized constructs in an insulated expression system, integrated into the same genomic location, enabling reliable gain-of-function analyses across multiple tissues. Distinct phenotypic outcomes were observed, reflecting both shared and unique features of individual ligands. To define the canonicity of Wnt signalling, we monitored canonical targets such as Dfz3, notum, and the 'naked cuticle' phenotype in developing tissues and the adult gut. Our findings revealed strong evidence of canonical responses from not only wg, but also DWnt6 and DWnt10 in the embryo, wing disc, larval gut, and adult gut. In addition, DWnt2, DWnt4, and WntD produced phenotypes distinct from the control with DWnt2 and DWnt4, showing context-dependent evidence of some canonical activity. While previous studies have suggested regulatory features between wg and DWnt6, our work provides functional evidence that Wg, DWnt6, and DWnt10 each induce expression of canonical signalling reporters in vivo. These findings refine our understanding of redundancy and specificity within the Drosophila Wnt family and demonstrate that multiple Wnt ligands can act similarly within the canonical pathway depending on tissue context.

Drosophila melanogaster is a highly versatile model organism that has profoundly advanced our understanding of human diseases. With more than 60% of its genes having human homologs, Drosophila provides an invaluable system for modelling a wide range of pathologies, including neurodegenerative disorders, cancer, metabolic diseases, as well as cardiac and muscular conditions. This review highlights key developments in utilizing Drosophila for disease modelling, emphasizing the genetic tools that have transformed research in this field. Technologies such as the GAL4/UAS system, RNA interference (RNAi) and CRISPR-Cas9 have enabled precise genetic manipulation, with CRISPR-Cas9 allowing for the introduction of human disease mutations into orthologous Drosophila genes. These approaches have yielded critical insights into disease mechanisms, identified novel therapeutic targets and facilitated both drug screening and toxicological studies. Articles were selected based on their relevance, impact and contribution to the field, with a particular focus on studies offering innovative perspectives on disease mechanisms or therapeutic strategies. Our findings emphasize the central role of Drosophila in studying complex human diseases, underscoring its genetic similarities to humans and its effectiveness in modelling conditions such as Alzheimer's disease, Parkinson's disease and cancer. This review reaffirms Drosophila's critical role as a model organism, highlighting its potential to drive future research and therapeutic advancements.

UCH-L1 (Ubiquitin Carboxyl-terminal Hydrolase - L1) is a protein that plays a critical role in the ubiquitin-proteasome system. Previous studies have demonstrated a link between UCH-L1 and various diseases, including neurodegenerative disorders, diabetes, and cancer. However, the role of UCH-L1 in development remains unclear. To investigate the functions of UCH-L1 in a living organism, taking advantage of the Drosophila model, and to explore the correlation between Drosophila UCH (dUCH) and human UCH-L1, we established a GAL4/UAS-targeted expression system to examine the effect of dUCH on Drosophila eye development. We found that knockdown of dUCH resulted in a rough eye phenotype associated with the MAPK pathway. In this study, for the first time, we revealed that loss of dUCH function leads to a reduction in EGFR protein levels. Additionally, dUCH knockdown downregulated Spitz (spi), a ligand of EGFR, as well as Draf, a key component of the MAPK pathway. Furthermore, under dUCH knockdown conditions, several genes known to play critical roles in eye cell differentiation were affected, including the downregulation of sens, salm, lz, barth1/2, and salm, which are essential for the differentiation of R2/5, R3/4, and R1/6 photoreceptor cells. Interestingly, dUCH was found to be involved not only in the MAPK pathway but also in the regulation of pros, lz, barth1/2, and sev gene expression, suggesting its role in R7 photoreceptor differentiation. Taken together, these findings highlight the important role of dUCH in regulating genes associated with eye cell differentiation and its involvement in EGFR signalling in Drosophila melanogaster.

We are utilizing an adult penetrating traumatic brain injury (PTBI) model in Drosophila to investigate regenerative mechanisms after damage to the central brain. Here, we focus on cell proliferation as an early event in the regenerative process. To identify pathways that could trigger cell proliferation following PTBI, we utilized bulk RNA-Seq. We find that transcript levels for components of both Toll and Immune Deficiency (Imd) innate immunity pathways are rapidly and highly upregulated post-PTBI. We then tested mutants for the NF-κB transcription factors of the Toll and Imd pathways, Dorsal-related immunity factor (Dif) and Relish (Rel), respectively. We find that loss of either Dif or Rel results in loss of cell proliferation after injury and identify tissue-specific requirements for Dif and Rel. In addition, while the canonical downstream targets of Drosophila innate immune signalling, the antimicrobial peptides (AMPs), are upregulated following PTBI, their levels revert to near baseline within 24 hr. Taken together, these results indicate that the innate immunity pathways play an integral role in the regenerative response and that this response may not require the antimicrobial peptides. Innate immunity previously has been implicated as both a potentiator and an inhibitor of regenerative processes. Our work suggests that modulation of innate immunity may be essential to prevent adverse outcomes. Thus, this work is likely to inform future experiments to dissect regenerative mechanisms in higher organisms as well as in Drosophila.

Three decades of research aimed at understanding the basis for autosomal recessive primary microcephaly (MCPH), a human clinical disorder defined by a significant reduction in head and brain size, has uncovered a suite of ~30 genes that participate in this process. Work in both vertebrate and invertebrate model systems have been instrumental in attempting to link MCPH gene function to the brain growth phenotype. However, we still lack definitive evidence as to what these functions are for many of these genes. In this review, we summarize recent work in Drosophila aimed at overcoming these limitations in our knowledge of MCPH gene function that may be applicable to humans. We discuss the clinical features of MCPH, parallels between human and Drosophila neurogenesis modes with a particular focus on the fly optic lobe, and highlight four of the most well-studied Drosophila MCPH orthologs: abnormal spindle (asp)/MCPH5, Microcephalin/MCPH1, WD Repeat-Containing Protein 62 (Wdr62)/MCPH2, and Ankryin Repeat-and LEM Domain- Containing Protein 2 (ANKLE2)/MCPH16. We focus on the multifunctional roles for these proteins that may underlie the microcephaly phenotype and advocate for the use of flies as a relevant model for human MCPH.

Neurodegenerative diseases are devastating conditions characterized by progressive cognitive decline with few available treatments. Neurodegeneration can be quantified in vertebrate and invertebrate models of disease by analysis of vacuolation - the formation of empty spaces within brain tissue. Previous approaches for quantifying this phenotype have required time-consuming methods such as manual counting and measuring of vacuole dimensions, which can be subjective. Here we describe VacQuant, a novel application that can be paired with existing machine learning software to automatically measure the area of vacuolation in brain tissue. Using Drosophila brain sections from tauopathy model flies, a well-described model of dementia-related neurodegeneration, we quantified a significant increase in brain vacuolation at several timepoints in adult flies with the aid of VacQuant. When compared with quantification by five blinded volunteers, the machine learning method positively correlated with their group average, confirming its accuracy and functionality. This automated method developed with VacQuant removes human bias and measurement variation, providing a consistent threshold for all brain sections and experiments. This automated pipeline will be particularly useful for high-throughput screening for genetic modifiers or therapeutic compounds in animal models of neurodegeneration.

The bipartite GAL4/UAS system is the most widely used method for targeted gene expression in Drosophila melanogaster and facilitates rapid in vivo genetic experimentation. Defining precise gene expression patterns for tissues and/or cell types under GAL4 control will continue to evolve to suit experimental needs. However, the precise spatial and temporal expression patterns for some commonly used muscle tissue promoters are still unclear. This missing information limits the precise timing of experiments during development. Here, we focus on three muscle-enriched GAL4 drivers (Mef2-GAL4, C57-GAL4 and G7-GAL4) to better inform selection of the most appropriate muscle promoter for experimental needs. Specifically, C57-GAL4 and G7-GAL4 turn on in the first or second instar larval stages, respectively, and can be used to bypass myogenesis for studies of muscle function after development.