Wild-type Flies Research Articles

A63: Exercise Improves Appetite and Heart Function in High Fat Drosophila

Purpose: High-fat diets cause obesity and disease leading to excess appetite and cardiovascular disease. At present, there is literature showing the improvement effect of exercise on obesity and related diseases. To explore more deeply the mechanism of action of exercise on appetite improvement and heart function in high-fat diets, we used fruit fly motility models to reveal this aspect of function. Methods: A total of 300 wild-type W1118 virgin flies that matured within 12 hours were collected. They were randomly divided into 100 animals in the normal diet control group (NFD), 100 in the high-fat diet group (HFD), and 100 in the high-fat diet exercise group (HFD+E). Exercise intervention for 7-day-old fruit flies for 5 consecutive days. An EM-CCD high-speed camera was used to record the heartbeat of fruit flies (video at 130 fps, the 30 s), and HC Image software was used to record the cardiogram data. Semi-Automated Optical Heartbeat Analysis (SOHA) quantifies Heart Rate (HR), Heart Period (HP), Diastolic Intervals (DI), Systolic Intervals (SI), Arrhythmia Index (AI), Diastolic Diameter (DD), Systolic Diameter (SD), Fractional Shortening (FS), and Fibrillations (FL). Fruit fly uptake was measured using the FlyPAD high-throughput Drosophila quantitative feeding system. All fruit flies needed to be fed on normal medium for 5 days first and then transferred to fresh normal medium or the high-fat medium on the 6th day for another 2 days. NFD flies are placed in a constant temperature and humidity incubator (25 °C, 50% humidity, 12 h day and night cycle), HFD flies are housed in incubators at 22-24 °C and 50% relative humidity to make high-fat medium by mixing 30% coconut oil and 70% standard medium. Results: The HFD group had an increase in AI, HR and HP, constant SD, decreased DD, and decreased FS. After exercise, HFD+E group had a decrease in HR, an increase in HP, an improvement in AI, an improvement in DD, and no change in SD. The food intake and sipping frequency of fruit flies in the HFD group were significantly higher than those in the NFD group, and during the same time period, the food intake and number of sipping times in the HFD+E group and the HFD group decreased significantly after exercise. Conclusion: Exercise improved excess appetite and cardiac dysfunction in high-fat diets.

Stress Pathways Induced by Volatile Anesthetics and Failure of Preconditioning in a Mitochondrial Complex I Mutant.

Carriers of mutations in the mitochondrial electron transport chain are at increased risk of anesthetic-induced neurotoxicity. To investigate the neurotoxicity mechanism and to test preconditioning as a protective strategy, this study used a Drosophila melanogaster model of Leigh syndrome. Model flies carried a mutation in ND23 (ND2360114) that encodes a mitochondrial electron transport chain complex I subunit. This study investigated why ND2360114 mutants become susceptible to lethal, oxygen-modulated neurotoxicity within 24 h of exposure to isoflurane but not sevoflurane. This study used transcriptomics and quantitative real-time reverse transcription polymerase chain reaction to identify genes that are differentially expressed in ND2360114 but not wild-type fly heads at 30 min after exposure to high- versus low-toxicity conditions. This study also subjected ND2360114 flies to diverse stressors before isoflurane exposure to test whether isoflurane toxicity could be diminished by preconditioning. The ND2360114 mutation had a greater effect on isoflurane- than sevoflurane-mediated changes in gene expression. Isoflurane and sevoflurane did not affect expression of heat shock protein (Hsp) genes (Hsp22, Hsp27, and Hsp68) in wild-type flies, but isoflurane substantially increased expression of these genes in ND2360114 mutant flies. Furthermore, isoflurane and sevoflurane induced expression of oxidative (GstD1 and GstD2) and xenobiotic (Cyp6a8 and Cyp6a14) stress genes to a similar extent in wild-type flies, but the effect of isoflurane was largely reduced in ND2360114 flies. In addition, activating stress response pathways by pre-exposure to anesthetics, heat shock, hyperoxia, hypoxia, or oxidative stress did not suppress isoflurane-induced toxicity in ND2360114 mutant flies. Mutation of a mitochondrial electron transport chain complex I subunit generates differential effects of isoflurane and sevoflurane on gene expression that may underlie their differential effects on neurotoxicity. Additionally, the mutation produces resistance to preconditioning by stresses that protect the brain in other contexts. Therefore, complex I activity modifies molecular and physiologic effects of anesthetics in an anesthetic-specific manner.

Parallel evolution and enhanced virulence upon in vivo passage of an RNA virus in Drosophila melanogaster.

Virus evolution is strongly affected by antagonistic co-evolution of virus and host. Host immunity positively selects for viruses that evade the immune response, which in turn may drive counter-adaptations in host immune genes. We investigated how host immune pressure shapes virus populations, using the fruit fly Drosophila melanogaster and its natural pathogen Drosophila C virus (DCV), as a model. We performed an experimental evolution study in which DCV was serially passaged for ten generations in three fly genotypes differing in their antiviral RNAi response: wild-type flies and flies in which the endonuclease gene Dicer-2 was either overexpressed or inactivated. All evolved virus populations replicated more efficiently in vivo and were more virulent than the parental stock. The number of polymorphisms increased in all three host genotypes with passage number, which was most pronounced in Dicer-2 knockout flies. Mutational analysis showed strong parallel evolution, as mutations accumulated in a specific region of the VP3 capsid protein in every lineage in a host genotype-independent manner. The parental tyrosine at position ninety-five of VP3 was substituted with either one of five different amino acids in fourteen out of fifteen lineages. However, no consistent amino acid changes were observed in the viral RNAi suppressor gene 1A, nor elsewhere in the genome in any of the host backgrounds. Our study indicates that the RNAi response restricts the sequence space that can be explored by viral populations. Moreover, our study illustrates how evolution towards higher virulence can be a highly reproducible, yet unpredictable process.

The identification of regulatory ceRNA network involved in Drosophila Toll immune responses

Non-coding RNAs play important roles in the innate immunity of Drosophila, with various lncRNAs and miRNAs identified to maintain Drosophila innate immune homeostasis by regulating protein functions. However, it remains unclear whether interactions between lncRNAs and miRNAs give rise to a ceRNA network. In our previous study, we observed the highest differential expression levels of lncRNA-CR11538, lncRNA-CR33942, and lncRNA-CR46018 in wild-type flies after Gram-positive bacterial infection, prompting us to investigate their role in the regulation of Drosophila Toll immune response through RNA-seq analysis. Herein, our comprehensive bioinformatics analysis revealed that lncRNA-CR11538, lncRNA-CR33942, and lncRNA-CR46018 are involved in defense mechanisms and stimulus response. Moreover, lncRNA-CR11538 and lncRNA-CR46018 can also participate in the metabolic recovery processes following Gram-positive bacterial infection. Subsequently, we employed GSEA screening and RT-qPCR to identify seven miRNAs (miR-957, miR-1015, miR-982, miR-993, miR-1007, miR-193, and miR-978) that may be regulated by these three lncRNAs. Furthermore, we predicted the potential target genes in the Toll signaling pathway for these miRNAs and their interaction with the three lncRNAs using TargetScan and miRanda software and preliminary verification. As a result, we established a potential ceRNA regulatory network for Toll immune responses in Drosophila, comprising three lncRNAs and seven miRNAs. This study provides evidence of a ceRNA regulatory network in Drosophila Toll immune responses and offers novel insights into understanding the regulatory networks involved in the innate immunity of other animals.

Toxicological assessment of sodium benzoate in Drosophila melanogaster.

Sodium benzoate (SB), the sodium salt of benzoic acid, is a food preservative with wide applications in the food, cosmetic and pharmaceutical industries due to its ability to kill many microorganisms effectively. Experimental evidence however suggests that excessive intake of SB poses detrimental health risks among consumers in the population. The present study investigated the toxic effects of various concentrations of SB using Drosophila melanogaster as a model. Adult wild-type flies of Canton S strain (1- to 3-daysold) was orally exposed to SB (0, 0.5, 1.0, 2.0 and 5.0 mg/5 g diet) to evaluate survival rates for 21days. Thereafter, we evaluated markers of oxidative stress, antioxidant status and behavioral activity in D. melanogaster exposed to SB for seven (7) days. We observed that SB (2.0 and 5.0 mg/5 g diet) decreased the survival of D. melanogaster. Also, SB inhibited glutathione-S-transferase activity and depleted total thiols and nonprotein thiols contents. Moreover, SB (5 mg/5 g diet) increased nitric oxide (nitrite/nitrate) level and reduced flies' emergence rate. Conclusively, findings from this study revealed that exposure to high concentrations of SB reduced survival rate and induced toxicity via the induction of oxidative stress and inhibition of antioxidant enzymes in D. melanogaster.

Immunodetection of Truncated Forms of the α6 Subunit of the nAChR in the Brain of Spinosad Resistant Ceratitis capitata Phenotypes.

The α6 subunit of the nicotinic acetylcholine receptor (nAChR) has been proposed as the target for spinosad in insects. Point mutations that result in premature stop codons in the α6 gene of Ceratitis capitata flies have been previously associated with spinosad resistance, but it is unknown if these transcripts are translated and if so, what is the location of the putative truncated proteins. In this work, we produced a specific antibody against C. capitata α6 (Ccα6) and validated it by ELISA, Western blotting and immunofluorescence assays in brain tissues. The antibody detects both wild-type and truncated forms of Ccα6 in vivo, and the protein is located in the cell membrane of the brain of wild-type spinosad sensitive flies. On the contrary, the shortened transcripts present in resistant flies generate putative truncated proteins that, for the most part, fail to reach their final destination in the membrane of the cells and remain in the cytoplasm. The differences observed in the locations of wild-type and truncated α6 proteins are proposed to determine the susceptibility or resistance to spinosad.

The effect of cigarette smoke extract exposure on the size and sexual behaviour of Drosophila melanogaster

Drosophila melanogaster is a widely used animal model in human diseases and to date it has not been applied to the study of the impact of tobacco use on human sexual function. Hence, this report examines the effects of different concentrations of cigarette smoke extract (CSE) exposure on the size and sexual behavior of D. melanogaster. Wild-type flies were held in vials containing CSE-infused culture media at concentrations of 10%, 25%, and 50% for three days, and their offspring were reared under the same conditions before measuring their body size and mating behavior. CSE exposure during development reduced the tibia length and body mass of emerging adult flies and prolonged the time required for successful courtship copulation success, while courtship behaviors (wing extension, tapping, abdomen bending, attempted copulation) remained largely unchanged. Our findings indicate that CSE exposure negatively affects the development of flies and their subsequent reproductive success. Future experiments should investigate the CSE effect on male female fertility.

Laboratory-adapted and wild-type black soldier flies express differential plasticity in bioconversion and nutrition when reared on urban food waste streams.

The black soldier fly (BSF) offers a potential solution to address shortages of feed and food sources; however, selecting effective rearing substrates remains a major hurdle in BSF farming. In an urban area like Singapore, current practice is based on rearing BSF on homogeneous waste streams (e.g., spent brewery grains or okara) because heterogeneous food wastes (e.g., mixed kitchen/canteen waste or surplus cooked food) present several operational challenges with respect to the standardization of development, nutritional content, and harvesting. In this study, we compared two genetic strains of BSF larvae (wild-type and laboratory-adapted line) in a bioconversion experiment with diverse types of food waste (homogeneous/heterogeneous; plant/meat) and we quantified the phenotypic plasticity. Our results demonstrate different plasticity in bioconversion performance, larval growth and larval nutrition between the two BSF lines. This difference may be attributed to the selective breeding the laboratory-adapted line has experienced. Notably, larval lipid content displayed little to no genetic variation for plasticity compared with larval protein and carbohydrate content. Despite variation in larval development, heterogeneous food wastes can produce better performance in bioconversion, larval growth, and larval nutrient content than homogeneous food waste. All-meat diets result in high larvae mortality but larval survival could be rescued by mixing meat with plant-based food wastes. Overall, we suggest using mixed meals for BSF larvae feeding. Targeted breeding may be a promising strategy for the BSF industry but it is important to consider the selection effects on plasticity in larval nutrition carefully. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Ex Vivo Calcium Imaging for Drosophila Model of Epilepsy.

Epilepsy is a neurological disorder characterized by recurrent seizures, partially correlated with genetic origin, affecting over 70 million individuals worldwide. Despite the clinical importance of epilepsy, the functional analysis of neural activity in the central nervous system is still to be developed. Recent advancements in imaging technology, in combination with stable expression of genetically encoded calcium indicators, such as GCaMP6, have revolutionized the study of epilepsy at both brain-wide and single-cell resolution levels. Drosophila melanogaster has emerged as a tool for investigating the molecular and cellular mechanisms underlying epilepsy due to its sophisticated molecular genetics and behavioral assays. In this study, we present a novel and efficient protocol for ex vivo calcium imaging in GCaMP6-expressing adult Drosophila to monitor epileptiform activities. The whole brain is prepared from cac, a well-known epilepsy gene, knockdown flies for calcium imaging with a confocal microscope to identify the neural activity as a follow-up to the bang-sensitive seizure-like behavior assay. The cac knockdown flies showed a higher rate of seizure-like behavior and abnormal calcium activities, including more large spikes and fewer small spikes than wild-type flies. The calcium activities were correlated to seizure-like behavior. This methodology serves as an efficient methodology in screening the pathogenic genes for epilepsy and exploring the potential mechanism of epilepsy at the cellular level.

Iditarod, a Drosophila homolog of the Irisin precursor FNDC5, is critical for exercise performance and cardiac autophagy

Mammalian FNDC5 encodes a protein precursor of Irisin, which is important for exercise-dependent regulation of whole-body metabolism. In a genetic screen in Drosophila, we identified Iditarod (Idit), which shows substantial protein homology to mouse and human FNDC5, as a regulator of autophagy acting downstream of Atg1/Atg13. Physiologically, Idit-deficient flies showed reduced exercise performance and defective cold resistance, which were rescued by exogenous expression of Idit. Exercise training increased endurance in wild-type flies, but not in Idit-deficient flies. Conversely, Idit is induced upon exercise training, and transgenic expression of Idit in wild-type flies increased endurance to the level of exercise trained flies. Finally, Idit deficiency prevented both exercise-induced increase in cardiac Atg8 and exercise-induced cardiac stress resistance, suggesting that cardiac autophagy may be an additional mechanism by which Idit is involved in the adaptive response to exercise. Our work suggests an ancient role of an Iditarod/Irisin/FNDC5 family of proteins in autophagy, exercise physiology, and cold adaptation, conserved throughout metazoan species.

Drosophila genotypes can be predicted from their exploration locomotive trajectories using supervised machine learning

This study employs supervised machine learning algorithms to test whether locomotive features during exploratory activity in open field arenas can serve as predictors for the genotype of fruit flies. Because of the nonlinearity in locomotive trajectories, traditional statistical methods that are used to compare exploratory activity between genotypes of fruit flies may not reveal all insights. 10-minute-long trajectories of four different genotypes of fruit flies in an open-field arena environment were captured. Turn angles and step size features extracted from the trajectories were used for training supervised learning models to predict the genotype of the fruit flies. Using the first five minute locomotive trajectories, an accuracy of 83% was achieved in differentiating wild-type flies from three other mutant genotypes. Using the final 5 min and the entire ten minute duration decreased the performance indicating that the most variations between the genotypes in their exploratory activity are exhibited in the first few minutes. Feature importance analysis revealed that turn angle is a better predictor than step size in predicting fruit fly genotype. Overall, this study demonstrates that features of trajectories can be used to predict the genotype of fruit flies through supervised machine learning methods.

Two clip-domain serine protease homologs, cSPH35 and cSPH242, act as a cofactor for prophenoloxidase-1 activation in Drosophila melanogaster.

Insect phenoloxidases (POs) catalyze phenol oxygenation and o-diphenol oxidation to form reactive intermediates that kill invading pathogens and form melanin polymers. To reduce their toxicity to host cells, POs are produced as prophenoloxidases (PPOs) and activated by a serine protease cascade as required. In most insects studied so far, PPO activating proteases (PAPs) generate active POs in the presence of a high Mr cofactor, comprising two serine protease homologs (SPHs) each with a Gly residue replacing the catalytic Ser of an S1A serine protease (SP). These SPHs have a regulatory clip domain at the N-terminus, like most of the SP cascade members including PAPs. In Drosophila, PPO activation and PO-catalyzed melanization have been examined in genetic analyses but it is unclear if a cofactor is required for PPO activation. In this study, we produced the recombinant cSPH35 and cSPH242 precursors, activated them with Manduca sexta PAP3, and confirmed their predicted role as a cofactor for Drosophila PPO1 activation by MP2 (i.e., Sp7). The cleavage sites and mechanisms for complex formation and cofactor function are highly similar to those reported in M. sexta. In the presence of high Mr complexes of the cSPHs, PO at a high specific activity of 260 U/μg was generated in vitro. To complement the in vitro analysis, we measured hemolymph PO activity levels in wild-type flies, cSPH35, and cSPH242 RNAi lines. Compared with the wild-type flies, only 4.4% and 18% of the control PO level (26 U/μl) was detected in the cSPH35 and cSPH242 knockdowns, respectively. Consistently, percentages of adults with a melanin spot at the site of septic pricking were 82% in wild-type, 30% in cSPH35 RNAi, and 53% in cSPH242 RNAi lines; the survival rate of the control (45%) was significantly higher than those (30% and 15%) of the two RNAi lines. These data suggest that Drosophila cSPH35 and cSPH242 are components of a cofactor for MP2-mediated PPO1 activation, which are indispensable for early melanization in adults.

The Effects of SKF-835666 on Levodopa-Induced Dyskinesia in Drosophila melanogaster

Although levodopa is considered the "gold standard" medication for Parkinson's disease, it is flawed as almost 90% of patients contract a side effect known as Levodopa Induced Dyskinesia (LID) after 10 years of treatment. LID manifests as involuntary movements and muscle spasms and presently lacks a cure. Previous studies on Drosophila melanogaster models have found that epigenetically inhibiting the ADCY2 gene reduces LID symptoms as represented by abnormal involuntary movements. Consequently, this study sought to assess the therapeutic potential of SKF-83566, an ADCY2 inhibitor, in ameliorating LID symptoms within a D. melanogaster model. The primary objective was to quantify the drug's impact by implementing Abnormal Involuntary Movements (AIMs) assay. This study found that wild-type flies on a diet of 10 mM of levodopa had a statistically significant increase in AIMs scores when compared to normal flies, implying that they successfully modeled LID. Furthermore, the data from this study supports that SKF-83566 alleviates LID symptoms, as flies with LID had lower AIMs scores when given SKF-83566. Moreover, findings from this study indicate that the locomotion of healthy D. melanogaster remained unaffected when exposed to a 5 mM dosage of SKF-83566 over a 7-day period. This study presents a novel administration procedure of SKF-83566 which could be applied in future research with D. melanogaster. The demonstrated effectiveness of the compound in reducing LID symptoms within a D. melanogaster model highlights its potential as a treatment option for LID, which could be researched further in the future.

Identifying genetic factors that increase cognitive reserve: A theoretical approach

Studies have demonstrated that some individuals display pathological hallmarks of Alzheimer's disease (AD) but are not afflicted with cognitive decline. The ability to maintain cognitive function despite the presence of pathology is referred to as cognitive reserve. This project aims to identify the molecular pathways involved in cognitive reserve using Drosophila melanogaster (Drosophila) models of AD. Specifically, a theoretical approach using experimental evolution to drive a population of AD-like Drosophila carrying a tau mutation to develop cognitive reserve is proposed. To accomplish this, a population of AD-like Drosophila will be placed in a single population cage along with wild-type flies and forced to compete for food and water. The first generation of AD-like Drosophila will be generated using random mutagenesis of the initially isogenic AD-like fly. The selected tau mutant displays a rough eye condition which allows for easy distinction between tau mutant and wild-type flies. It is hypothesised that AD-like flies with cognitive decline will be unable to survive because their limited cognitive abilities will prevent them from effectively competing for food and water. In contrast, AD-like flies with mutations that promote cognitive reserve will be better capable of survival. After 90-99% of mutant flies have died, the surviving mutant flies will be back-crossed to the P1 mutant to maintain tau mutation stability. It is expected that artificial selection will result in the creation of a generation of tau mutant flies that demonstrate cognitive abilities comparable to those of wild-type flies despite maintaining an AD-like tau mutation. This approach will monitor the successful trajectory of the evolution of increased cognitive reserve through survival curve analysis and measures of cognition. A limitation of the method is that only a dominant mutation or series of dominant mutations would be identified using this approach.

The circadian clock is required for rhythmic lipid transport in Drosophila in interaction with diet and photic condition

Modern lifestyle is often at odds with endogenously driven rhythmicity, which can lead to circadian disruption and metabolic syndrome. One signature for circadian disruption is a reduced or altered metabolite cycling in the circulating tissue reflecting the current metabolic status. Drosophila is a well-established model in chronobiology, but day-time dependent variations of transport metabolites in the fly circulation are poorly characterized. Here, we sampled fly hemolymph throughout the day and analyzed diacylglycerols (DGs), phosphoethanolamines (PEs) and phosphocholines (PCs) using LC-MS. In wild-type flies kept on sugar-only medium under a light-dark cycle, all transport lipid species showed a synchronized bimodal oscillation pattern with maxima at the beginning and end of the light phase which were impaired in period01 clock mutants. In wild-type flies under constant dark conditions, the oscillation became monophasic with a maximum in the middle of the subjective day. In strong support of clock-driven oscillations, levels of the targeted lipids peaked once in the middle of the light phase under time-restricted feeding independent of the time of food intake. When wild-type flies were reared on full standard medium, the rhythmic alterations of hemolymph lipid levels were greatly attenuated. Our data suggest that the circadian clock aligns daily oscillations of DGs, PEs, and PCs in the hemolymph to the anabolic siesta phase, with a strong influence of light on phase and modality.

Drosophila melanogaster as a model of systemic dermatophytosis.

Dermatophytosis is one of the most common fungal infections worldwide. The distribution of dermatophytes varies across continents, but the genera Trichophyton and Microsporum have emerged as the main isolated agents in humans and animals. To validate Drosophila melanogaster flies as a fast and feasible model to study dermatophytic infections. Wild-type (WT) and Toll-deficient D. melanogaster flies were infected by Trichophyton rubrum, T. mentagrophytes, Microsporum canis and Nannizzia gypsea by pricking with a needle previously dipped in inoculum concentrations ranging from 103 to 108 colony-forming units/mL. Establishment of infection was confirmed by survival curves, histopathological analysis and fungal burden. Thereafter, flies were treated with terbinafine, itraconazole and clioquinol. WT flies were predominantly resistant to the infection, whereas Toll-deficient flies succumbed to the four dermatophyte genera tested. The antifungal drugs protected flies from the infection, except for N. gypsea whose survival curves did not differ from the untreated group. This pilot study confirms that D. melanogaster is a suitable model to study the virulence and antifungal drug efficacy in dermatophyte species.

Measuring O2 Consumption in Drosophila Melanogaster using Coulometric Microrespirometry.

Coulometric microrespirometry is a straightforward, inexpensive method for measuring the O2 consumption of small organisms while maintaining a stable environment. A coulometric microrespirometer consists of an airtight chamber in which O2 is consumed and the CO2 produced by the organism is removed by an absorbent medium. The resulting pressure decrease triggers electrolytic O2 production, and the amount of O2 produced is measured by recording the amount of charge used to generate it. In the present study, the method has been adapted to Drosophila melanogaster tested in small groups, with the sensitivity of the apparatus and the environmental conditions optimized for high stability. The amount of O2 consumed by wildtype flies in this apparatus is consistent with that measured by previous studies. Mass-specific O2 consumption by CASK mutants, which are smaller and known to be less active, was not different from congenic controls. However, the small size of CASK mutants resulted in a significant reduction in O2 consumption on a per-fly basis. Therefore, the microrespirometer is capable of measuring O2 consumption in D. melanogaster, can distinguish modest differences between genotypes, and adds a versatile tool for measuring metabolic rates.

The NF-κB/Relish Activates miR-308 to Negatively Regulate Imd Pathway Immune Signaling in Drosophila.

The strength and duration of the NF-κB signaling response must be tightly modulated to avoid an inadequate or excessive immune response. Relish, a core NF-κB transcription factor of the Drosophila Imd pathway, can control the expression of antimicrobial peptides, including Dpt and AttA, to defend against Gram-negative bacterial infections, but whether Relish may regulate miRNA expression to participate in the immune response remains unclear. In this study, taking advantage of Drosophila S2 cells and different overexpression/knockout/knockdown flies, we first found that Relish could directly activate the expression of miR-308 to negatively regulate the immune response and promote the survival of Drosophila during Enterobacter cloacae infection. Second, our results demonstrated that Relish-mediated expression of miR-308 could suppress target gene Tab2 to attenuate the Drosophila Imd pathway signal during the middle and late stages of the immune response. Third, we detected the dynamic expression patterns of Dpt, AttA, Relish, miR-308, and Tab2 in wild-type flies after E. coli infection, which further revealed that the feedback regulatory loop of Relish-miR-308-Tab2 plays a crucial role in the immune response and homeostasis maintenance of the Drosophila Imd pathway. Overall, our present study not only illustrates an important mechanism by which this Relish-miR-308-Tab2 regulatory axis can negatively control the Drosophila immune response and participate in homeostasis maintenance but also provides new insights into the dynamic regulation of the NF-κB/miRNA expression network of animal innate immunity.

A new Drosophila melanogaster fly that expresses GFP-tagged Orco

Insects perceive countless odors with the help of odorant receptors (ORs) expressed by olfactory sensory neurons (OSNs). OR complexes are composed of odor-specific proteins OrX and olfactory co-receptor proteins Orco, with both extracellular C-terminus and intracellular N-terminus. Here, we generated a genetically modified Drosophila melanogaster fly line that expresses Orco proteins with an N-terminally fused green fluorescent protein (GFP). We generated four genotypes (homozygous and heterozygous) for which we performed antennal cryosectioning and immunolabeling to determine which genotype would show high GFP labeling intensity in the dendritic areas of OSNs. We found that fluorescent intensity was the highest in homozygous flies. As Orco is known to guide OrX proteins to the OSN dendrites, we checked whether the fusion of GFP to the Orco N-terminus may affect such trafficking and thereby OR function. Using single sensillum recording (SSR), we found no difference in the olfactory capability of homozygous flies compared with wild-type flies. As future application, we want to use this transgenic fly line to resolve the structure of OR complexes using Cryo electron microscopy (cryo-EM).

A role of cryptochrome for magnetic field-dependent improvement of sleep quality, lifespan, and motor function in Drosophila.

Understanding the molecular genetic basis of animal magnet reception has been one of the big challenges in molecular biology. Recently it was discovered that the magnetic sense of Drosophila melanogaster is mediated by the ultraviolet (UV)-A/blue light photoreceptor cryptochrome (Cry). Here, using the fruit fly as a magnet-receptive model organism, we show that the magnetic field exposure (0.4-0.6 mT) extended lifespan under starvation, but not in cryptochrome mutant flies (cryb ). The magnetic field exposure increases motor function in wild type and neurodegenerative disease model flies. Furthermore, the magnetic field exposure improved sleep quality at night-time specific manner, but not in cryb . We also showed that repeated AC magnetic field exposure increased climbing activity in wild-type Drosophila, but not in cryb . The data suggests that magnetic field-dependent improvement of lifespan, sleep quality, and motor function is mediated through a cry-dependent pathway in Drosophila.