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Optimization of hybridization chain reaction for imaging single RNA molecules in Drosophila larvae. 优化用于果蝇幼虫单个 RNA 分子成像的杂交链反应。
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-10-01 DOI: 10.1080/19336934.2024.2409968
Julia Olivares-Abril, Jana Joha, Jeffrey Y Lee, Ilan Davis
{"title":"Optimization of hybridization chain reaction for imaging single RNA molecules in <i>Drosophila</i> larvae.","authors":"Julia Olivares-Abril, Jana Joha, Jeffrey Y Lee, Ilan Davis","doi":"10.1080/19336934.2024.2409968","DOIUrl":"10.1080/19336934.2024.2409968","url":null,"abstract":"<p><p><i>In situ</i> hybridization techniques are powerful methods for exploring gene expression in a wide range of biological contexts, providing spatial information that is most often lost in traditional biochemical techniques. However, many <i>in situ</i> hybridization methods are costly and time-inefficient, particularly for screening-based projects that follow on from single-cell RNA sequencing data, which rely on of tens of custom-synthetized probes against each specific RNA of interest. Here we provide an optimized pipeline for Hybridization Chain Reaction (HCR)-based RNA visualization, including an open-source code for optimized probe design. Our method achieves high specificity and sensitivity with the option of multiplexing using only five pairs of probes, which greatly lowers the cost and time of the experiment. These features of our HCR protocol are particularly useful and convenient for projects involving screening several genes at medium throughput, especially as the method include an amplification step, which makes the signal readily visible at low magnification imaging.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11446410/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142344320","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
An unusual Toll/MyD88-mediated Drosophila host defence against Talaromyces marneffei. 一种不寻常的 Toll/MyD88 介导的果蝇宿主防御马拉尼菲氏菌(Talaromyces marneffei)的方法。
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-09-06 DOI: 10.1080/19336934.2024.2398300
Xiaoyue Wang, Qinglin Qu, Zi Li, Sha Lu, Dominique Ferrandon, Liyan Xi
{"title":"An unusual Toll/MyD88-mediated <i>Drosophila</i> host defence against <i>Talaromyces marneffei</i>.","authors":"Xiaoyue Wang, Qinglin Qu, Zi Li, Sha Lu, Dominique Ferrandon, Liyan Xi","doi":"10.1080/19336934.2024.2398300","DOIUrl":"10.1080/19336934.2024.2398300","url":null,"abstract":"<p><p>Talaromycosis, caused by <i>Talaromyces marneffei</i> (<i>T. marneffei</i>, formerly known as <i>Penicillium marneffei</i>), is an opportunistic invasive mycosis endemic in tropical and subtropical areas of Asia with high mortality rate. Despite various infection models established to study the immunological interaction between <i>T. marneffei</i> and the host, the pathogenicity of this fungus is not yet fully understood. So far, <i>Drosophila melanogaster</i>, a well-established genetic model organism to study innate immunity, has not been used in related research on <i>T. marneffei</i>. In this study, we provide the initial characterization of a systemic infection model of <i>T. marneffei</i> in the <i>D. melanogaster</i> host. Survival curves and fungal loads were tested as well as Toll pathway activation was quantified by RT-qPCR of several antimicrobial peptide (AMP) genes including <i>Drosomycin</i>, <i>Metchnikowin</i>, and <i>Bomanin Short 1</i>. We discovered that whereas most wild-type flies were able to overcome the infection, <i>MyD88</i> or <i>Toll</i> mutant flies failed to prevent fungal dissemination and proliferation and ultimately succumbed to this challenge. Unexpectedly, the induction of classical Toll pathway activation readouts, <i>Drosomycin</i> and <i>Bomanin Short 1</i>, by live or killed <i>T. marneffei</i> was quite limited in wild-type flies, suggesting that the fungus largely escapes detection by the systemic immune system. This unusual situation of a poor systemic activation of the Toll pathway and a strong susceptibility phenotype of <i>MyD88</i>/<i>Toll</i> might be accounted for by a requirement for this host defence in only specific tissues, a hypothesis that remains to be rigorously tested.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11382710/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142139738","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Mifepristone and rapamycin have non-additive benefits for life span in mated female Drosophila. 米非司酮和雷帕霉素对交配雌果蝇的寿命具有非叠加效益。
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-10-23 DOI: 10.1080/19336934.2024.2419151
Gary N Landis, Britta Baybutt, Shoham Das, Yijie Fan, Kate Olsen, Karissa Yan, John Tower
{"title":"Mifepristone and rapamycin have non-additive benefits for life span in mated female <i>Drosophila</i>.","authors":"Gary N Landis, Britta Baybutt, Shoham Das, Yijie Fan, Kate Olsen, Karissa Yan, John Tower","doi":"10.1080/19336934.2024.2419151","DOIUrl":"10.1080/19336934.2024.2419151","url":null,"abstract":"<p><p>The drugs mifepristone and rapamycin were compared for their relative ability to increase the life span of mated female <i>Drosophila melanogaster</i>. Titration of rapamycin indicated an optimal concentration of approximately 50 μM, which increased median life span here by average +81%. Meta-analysis of previous mifepristone titrations indicated an optimal concentration of approximately 466 μM, which increased median life span here by average +114%. Combining mifepristone with various concentrations of rapamycin did not produce further increases in life span, and instead reduced life span relative to either drug alone. Assay of maximum midgut diameter indicated that rapamycin was equally efficacious as mifepristone in reducing mating-induced midgut hypertrophy. The mito-QC mitophagy reporter is a previously described green fluorescent protein (GFP)-mCherry fusion protein targeted to the outer mitochondrial membrane. Inhibition of GFP fluorescence by the acidic environment of the autophagolysosome yields an increased red/green fluorescence ratio indicative of increased mitophagy. Creation of a multi-copy mito-QC reporter strain facilitated assay in live adult flies, as well as in dissected midgut tissue. Mifepristone was equally efficacious as rapamycin in activating the mito-QC mitophagy reporter in the adult female fat-body and midgut. The data suggest that mifepristone and rapamycin act through a common pathway to increase mated female <i>Drosophila</i> life span, and implicate increased mitophagy and decreased midgut hypertrophy in that pathway.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11514543/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142497591","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
A novel adipose loss-of-function mutant in Drosophila. 果蝇的一种新型脂肪功能缺失突变体
IF 1.2 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-05-13 DOI: 10.1080/19336934.2024.2352938
Nicole A Losurdo, Adriana Bibo, Jacob Bedke, Nichole Link
{"title":"A novel <i>adipose</i> loss-of-function mutant in <i>Drosophila</i>.","authors":"Nicole A Losurdo, Adriana Bibo, Jacob Bedke, Nichole Link","doi":"10.1080/19336934.2024.2352938","DOIUrl":"10.1080/19336934.2024.2352938","url":null,"abstract":"<p><p>To identify genes required for brain growth, we took an RNAi knockdown reverse genetic approach in <i>Drosophila</i>. One potential candidate isolated from this effort is the anti-lipogenic gene <i>adipose</i> (<i>adp</i>). Adp has an established role in the negative regulation of lipogenesis in the fat body of the fly and adipose tissue in mammals. While fat is key to proper development in general, <i>adp</i> has not been investigated during brain development. Here, we found that RNAi knockdown of <i>adp</i> in neuronal stem cells and neurons results in reduced brain lobe volume and sought to replicate this with a mutant fly. We generated a novel <i>adp</i> mutant that acts as a loss-of-function mutant based on buoyancy assay results. We found that despite a change in fat content in the body overall and a decrease in the number of larger (>5 µm) brain lipid droplets, there was no change in the brain lobe volume of mutant larvae. Overall, our work describes a novel <i>adp</i> mutant that can functionally replace the long-standing <i>adp</i><sup><i>60</i></sup> mutant and shows that the <i>adp</i> gene has no obvious involvement in brain growth.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11095658/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140915803","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Conserved A-to-I RNA editing with non-conserved recoding expands the candidates of functional editing sites. 保守的 A 到 I RNA 编辑与非保守的重新编码扩大了功能性编辑位点的候选范围。
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-06-18 DOI: 10.1080/19336934.2024.2367359
Yuange Duan, Ling Ma, Tianyou Zhao, Jiyao Liu, Caiqing Zheng, Fan Song, Li Tian, Wanzhi Cai, Hu Li
{"title":"Conserved A-to-I RNA editing with non-conserved recoding expands the candidates of functional editing sites.","authors":"Yuange Duan, Ling Ma, Tianyou Zhao, Jiyao Liu, Caiqing Zheng, Fan Song, Li Tian, Wanzhi Cai, Hu Li","doi":"10.1080/19336934.2024.2367359","DOIUrl":"10.1080/19336934.2024.2367359","url":null,"abstract":"<p><p>Adenosine-to-inosine (A-to-I) RNA editing recodes the genome and confers flexibility for the organisms to adapt to the environment. It is believed that RNA recoding sites are well suited for facilitating adaptive evolution by increasing the proteomic diversity in a temporal-spatial manner. The function and essentiality of a few conserved recoding sites are recognized. However, the experimentally discovered functional sites only make up a small corner of the total sites, and there is still the need to expand the repertoire of such functional sites with bioinformatic approaches. In this study, we define a new category of RNA editing sites termed 'conserved editing with non-conserved recoding' and systematically identify such sites in <i>Drosophila</i> editomes, figuring out their selection pressure and signals of adaptation at inter-species and intra-species levels. Surprisingly, conserved editing sites with non-conserved recoding are not suppressed and are even slightly overrepresented in <i>Drosophila</i>. DNA mutations leading to such cases are also favoured during evolution, suggesting that the function of those recoding events in different species might be diverged, specialized, and maintained. Finally, structural prediction suggests that such recoding in potassium channel Shab might increase ion permeability and compensate the effect of low temperature. In conclusion, conserved editing with non-conserved recoding might be functional as well. Our study provides novel aspects in considering the adaptive evolution of RNA editing sites and meanwhile expands the candidates of functional recoding sites for future validation.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11188811/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141418492","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
The astrocyte-enriched gene deathstar plays a crucial role in the development, locomotion, and lifespan of D. melanogaster. 富含星形胶质细胞的死亡之星基因在黑腹蝇蛆的发育、运动和寿命中起着至关重要的作用。
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-06-17 DOI: 10.1080/19336934.2024.2368336
Xiaoli Zhang, Dongyu Sun, Kyle Wong, Ammar Salkini, Hadi Najafi, Woo Jae Kim
{"title":"The astrocyte-enriched gene <i>deathstar</i> plays a crucial role in the development, locomotion, and lifespan of <i>D. melanogaster</i>.","authors":"Xiaoli Zhang, Dongyu Sun, Kyle Wong, Ammar Salkini, Hadi Najafi, Woo Jae Kim","doi":"10.1080/19336934.2024.2368336","DOIUrl":"10.1080/19336934.2024.2368336","url":null,"abstract":"<p><p>The <i>Drosophila melanogaster</i> brain is a complex organ with various cell types, orchestrating the development, physiology, and behaviors of the fly. While each cell type in <i>Drosophila</i> brain is known to express a unique gene set, their complete genetic profile is still unknown. Advances in the RNA sequencing techniques at single-cell resolution facilitate identifying novel cell type markers and/or re-examining the specificity of the available ones. In this study, exploiting a single-cell RNA sequencing data of <i>Drosophila</i> optic lobe, we categorized the cells based on their expression pattern for known markers, then the genes with enriched expression in astrocytes were identified. <i>CG11000</i> was identified as a gene with a comparable expression profile to the <i>Eaat1</i> gene, an astrocyte marker, in every individual cell inside the <i>Drosophila</i> optic lobe and midbrain, as well as in the entire <i>Drosophila</i> brain throughout its development. Consistent with our bioinformatics data, immunostaining of the brains dissected from transgenic adult flies showed co-expression of <i>CG11000</i> with <i>Eaat1</i> in a set of single cells corresponding to the astrocytes in the <i>Drosophila</i> brain. Physiologically, inhibiting <i>CG11000</i> through RNA interference disrupted the normal development of male <i>D. melanogaster</i>, while having no impact on females. Expression suppression of <i>CG11000</i> in adult flies led to decreased locomotion activity and also shortened lifespan specifically in astrocytes, indicating the gene's significance in astrocytes. We designated this gene as '<i>deathstar</i>' due to its crucial role in maintaining the star-like shape of glial cells, astrocytes, throughout their development into adult stage.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11185185/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141330680","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Ribose-cysteine and levodopa abrogate Parkinsonism via the regulation of neurochemical and redox activities in alpha-synuclein transgenic Drosophila melanogaster models. 核糖-半胱氨酸和左旋多巴通过调节α-突触核蛋白转基因黑腹果蝇模型的神经化学和氧化还原活动来缓解帕金森症
IF 2.4 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-01-29 DOI: 10.1080/19336934.2024.2306687
Olumayowa K Idowu, Ademola A Oremosu, Olufunke O Dosumu, Abdullahi A Mohammed
{"title":"Ribose-cysteine and levodopa abrogate Parkinsonism via the regulation of neurochemical and redox activities in alpha-synuclein transgenic <i>Drosophila melanogaster</i> models.","authors":"Olumayowa K Idowu, Ademola A Oremosu, Olufunke O Dosumu, Abdullahi A Mohammed","doi":"10.1080/19336934.2024.2306687","DOIUrl":"10.1080/19336934.2024.2306687","url":null,"abstract":"<p><p>Parkinson's disease (PD), the most prevalent type of parkinsonism, is a progressive neurodegenerative condition marked by several non-motor and motor symptoms. PD is thought to have a complex aetiology that includes a combination of age, genetic predisposition, and environmental factors. Increased expression of α-synuclein (α-Syn) protein is central to the evolvement of neuropathology in this devastating disorder, but the potential of ribose-cysteine and levodopa in abating pathophysiologic changes in PD model is unknown. Crosses were set up between flies conditionally expressing a pathological variant of human α-Syn (UAS-α-Syn) and those expressing GAL4 in neurons (elav-GAL4) to generate offspring referred to as PD flies. Flies were randomly assigned to five groups (<i>n</i> = 40) from the total population of flies, with each group having five replicates. Groups of PD flies were treated with either 500 mg/kg ribose-cysteine diet, 250 mg/kg levodopa diet, or a combination of the two compounds for 21 days, whereas the control group (w<sup>1118</sup>) and the PD group were exposed to a diet without ribose-cysteine or levodopa. In addition to various biochemical and neurochemical assays, longevity, larval motility, and gravitaxis assays were carried out. Locomotive capability, lifespan, fecundity, antioxidant state, and neurotransmitter systems were all significantly (<i>p</i> < 0.05) compromised by overexpression of α-Syn. However, flies treated both ribose cysteine and levodopa showed an overall marked improvement in motor functions, lifespan, fecundity, antioxidant status, and neurotransmitter system functions. In conclusion, ribose-cysteine and levodopa, both singly and in combination, potentiated a therapeutic effect on alpha-synuclein transgenic <i>Drosophila melanogaster</i> models of Parkinsonism.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10826630/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139574667","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Effects of unstable β-PheRS on food avoidance, growth, and development are suppressed by the appetite hormone CCHa2. 不稳定的β-PheRS对食物回避、生长和发育的影响受到食欲激素CCHa2的抑制。
IF 1.2 4区 生物学
Fly Pub Date : 2024-12-01 Epub Date: 2024-02-19 DOI: 10.1080/19336934.2024.2308737
Dominique Brunßen, Beat Suter
{"title":"Effects of unstable β-PheRS on food avoidance, growth, and development are suppressed by the appetite hormone CCHa2.","authors":"Dominique Brunßen, Beat Suter","doi":"10.1080/19336934.2024.2308737","DOIUrl":"10.1080/19336934.2024.2308737","url":null,"abstract":"<p><p>Amino acyl-tRNA synthetases perform diverse non-canonical functions aside from their essential role in charging tRNAs with their cognate amino acid. The phenylalanyl-tRNA synthetase (PheRS/FARS) is an α<sub>2</sub>β<sub>2</sub> tetramer that is needed for charging the tRNA<sup>Phe</sup> for its translation activity. Fragments of the α-subunit have been shown to display an additional, translation-independent, function that activates growth and proliferation and counteracts Notch signalling. Here we show in <i>Drosophila</i> that overexpressing the β-subunit in the context of the complete PheRS leads to larval roaming, food avoidance, slow growth, and a developmental delay that can last several days and even prevents pupation. These behavioural and developmental phenotypes are induced by PheRS expression in CCHa2<sup>+</sup> and Pros<sup>+</sup> cells. Simultaneous expression of β-PheRS, α-PheRS, and the appetite-inducing CCHa2 peptide rescued these phenotypes, linking this <i>β-PheRS</i> activity to the appetite-controlling pathway. The fragmentation dynamic of the excessive β-PheRS points to β-PheRS fragments as possible candidate inducers of these phenotypes. Because fragmentation of human FARS has also been observed in human cells and mutations in human <i>β-PheRS (FARSB)</i> can lead to problems in gaining weight, Drosophila <i>β-PheRS</i> can also serve as a model for the human phenotype and possibly also for obesity.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10880493/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139905439","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
The utility and caveat of split-GAL4s in the study of neurodegeneration. 分裂- gal4s在神经退行性疾病研究中的应用和警告。
IF 1.2 4区 生物学
Fly Pub Date : 2023-12-01 DOI: 10.1080/19336934.2023.2192847
Luca Stickley, Rafael Koch, Emi Nagoshi
{"title":"The utility and caveat of split-GAL4s in the study of neurodegeneration.","authors":"Luca Stickley, Rafael Koch, Emi Nagoshi","doi":"10.1080/19336934.2023.2192847","DOIUrl":"10.1080/19336934.2023.2192847","url":null,"abstract":"<p><p>Parkinson's disease (PD) is the second most common neurodegenerative disorder, afflicting over 1% of the population of age 60 y and above. The loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) is the primary cause of its characteristic motor symptoms. Studies using <i>Drosophila melanogaster</i> and other model systems have provided much insight into the pathogenesis of PD. However, little is known why certain cell types are selectively susceptible to degeneration in PD. Here, we describe an approach to identify vulnerable subpopulations of neurons in the genetic background linked to PD in <i>Drosophila</i>, using the split-GAL4 drivers that enable genetic manipulation of a small number of defined cell populations. We identify split-GAL4 lines that target neurons selectively vulnerable in a model of <i>leucine-rich repeat kinase 2</i> (<i>LRRK2</i>)-linked familial PD, demonstrating the utility of this approach. We also show an unexpected caveat of the split-GAL4 system in ageing-related research: an age-dependent increase in the number of GAL4-labelled cells.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2023-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10038051/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9953689","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 2
A tissue dissociation method for ATAC-seq and CUT&RUN in Drosophila pupal tissues. 果蝇蛹组织中ATAC-seq和CUT&RUN的组织分离方法。
IF 1.2 4区 生物学
Fly Pub Date : 2023-12-01 DOI: 10.1080/19336934.2023.2209481
Elli M Buchert, Elizabeth A Fogarty, Christopher M Uyehara, Daniel J McKay, Laura A Buttitta
{"title":"A tissue dissociation method for ATAC-seq and CUT&RUN in <i>Drosophila</i> pupal tissues.","authors":"Elli M Buchert, Elizabeth A Fogarty, Christopher M Uyehara, Daniel J McKay, Laura A Buttitta","doi":"10.1080/19336934.2023.2209481","DOIUrl":"10.1080/19336934.2023.2209481","url":null,"abstract":"<p><p>Chromatin accessibility, histone modifications, and transcription factor binding are highly dynamic during <i>Drosophila</i> metamorphosis and drive global changes in gene expression as larval tissues differentiate into adult structures. Unfortunately, the presence of pupa cuticle on many <i>Drosophila</i> tissues during metamorphosis prevents enzyme access to cells and has limited the use of enzymatic in situ methods for assessing chromatin accessibility and histone modifications. Here, we present a dissociation method for cuticle-bound pupal tissues that is compatible for use with ATAC-Seq and CUT&RUN to interrogate chromatin accessibility and histone modifications. We show this method provides comparable chromatin accessibility data to the non-enzymatic approach FAIRE-seq, with only a fraction of the amount of input tissue required. This approach is also compatible with CUT&RUN, which allows genome-wide mapping of histone modifications with less than 1/10th of the tissue input required for more conventional approaches such as Chromatin Immunoprecipitation Sequencing (ChIP-seq). Our protocol makes it possible to use newer, more sensitive enzymatic in situ approaches to interrogate gene regulatory networks during <i>Drosophila</i> metamorphosis.</p>","PeriodicalId":12128,"journal":{"name":"Fly","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2023-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10208176/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9609652","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
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