Friday, June 28, 2019

Automated tracking of leg movement in free-moving flies contributes to characterization of fly models of neurodegenerative diseases

Wu S, Tan KJ, Govindarajan LN, Stewart JC, Gu L, Ho JWH, Katarya M, Wong BH, Tan EK, Li D, Claridge-Chang A, Libedinsky C, Cheng L, Aw SS. Fully automated leg tracking of Drosophila neurodegeneration models reveals distinct conserved movement signatures. PLoS Biol. 2019 Jun 27;17(6):e3000346. PubMed PMID: 31246996.

Abstract: "Some neurodegenerative diseases, like Parkinsons Disease (PD) and Spinocerebellar ataxia 3 (SCA3), are associated with distinct, altered gait and tremor movements that are reflective of the underlying disease etiology. Drosophila melanogaster models of neurodegeneration have illuminated our understanding of the molecular mechanisms of disease. However, it is unknown whether specific gait and tremor dysfunctions also occur in fly disease mutants. To answer this question, we developed a machine-learning image-analysis program, Feature Learning-based LImb segmentation and Tracking (FLLIT), that automatically tracks leg claw positions of freely moving flies recorded on high-speed video, producing a series of gait measurements. Notably, unlike other machine-learning methods, FLLIT generates its own training sets and does not require user-annotated images for learning. Using FLLIT, we carried out high-throughput and high-resolution analysis of gait and tremor features in Drosophila neurodegeneration mutants for the first time. We found that fly models of PD and SCA3 exhibited markedly different walking gait and tremor signatures, which recapitulated characteristics of the respective human diseases. Selective expression of mutant SCA3 in dopaminergic neurons led to a gait signature that more closely resembled those of PD flies. This suggests that the behavioral phenotype depends on the neurons affected rather than the specific nature of the mutation. Different mutations produced tremors in distinct leg pairs, indicating that different motor circuits were affected. Using this approach, fly models can be used to dissect the neurogenetic mechanisms that underlie movement disorders."

Drosophila studies used to follow up on results of epigenome-wide association studies (EWAS) related to Alzheimer's disease

Higham JP, Malik BR, Buhl E, Dawson JM, Ogier AS, Lunnon K, Hodge JJL. Alzheimer's Disease Associated Genes Ankyrin and Tau Cause Shortened Lifespan and Memory Loss in Drosophila. Front Cell Neurosci. 2019 Jun 11;13:260. PubMed PMID: 31244615; PubMed Central PMCID: PMC6581016.

From the abstract: "Alzheimer's disease (AD) is the most common form of dementia and is characterized by intracellular neurofibrillary tangles of hyperphosphorylated Tau, including the 0N4R isoform and accumulation of extracellular amyloid beta (Aβ) plaques. ... Recent epigenome-wide association studies (EWAS) of AD have identified a number of loci that are differentially methylated in the AD cortex. Indeed, hypermethylation and reduced expression of the Ankyrin 1 (ANK1) gene in AD has been reported in the cortex in numerous different post-mortem brain cohorts. ... We have generated Drosophila models to allow us to functionally characterize Drosophila Ank2, the ortholog of human ANK1 and to determine its interaction with human Tau and Aβ. ... we show that the mis-expression of Ank2 can drive disease relevant processes and phenocopy some features of AD. Therefore, we propose targeting human ANK1 may have therapeutic potential. This represents the first study to characterize an AD-relevant gene nominated from EWAS."

Thursday, June 27, 2019

Fly model provides "first animal models for myosin-based Freeman-Sheldon syndrome (FSS)"

Rao DS, Kronert WA, Guo Y, Hsu KH, Sarsoza F, Bernstein SI. Reductions in ATPase activity, actin sliding velocity, and myofibril stability yield muscle dysfunction in Drosophila models of myosin-based Freeman-Sheldon syndrome. Mol Biol Cell. 2019 Jan 1;30(1):30-41. PubMed PMID: 30379605; PubMed Central PMCID: PMC6337914.

Abstract: "Using Drosophila melanogaster, we created the first animal models for myosin-based Freeman-Sheldon syndrome (FSS), a dominant form of distal arthrogryposis defined by congenital facial and distal skeletal muscle contractures. Electron microscopy of homozygous mutant indirect flight muscles showed normal (Y583S) or altered (T178I, R672C) myofibril assembly followed by progressive disruption of the myofilament lattice. In contrast, all alleles permitted normal myofibril assembly in the heterozygous state but caused myofibrillar disruption during aging. The severity of myofibril defects in heterozygotes correlated with the level of flight impairment. Thus our Drosophila models mimic the human condition in that FSS mutations are dominant and display varied degrees of phenotypic severity. Molecular modeling indicates that the mutations disrupt communication between the nucleotide-binding site of myosin and its lever arm that drives force production. Each mutant myosin showed reduced in vitro actin sliding velocity, with the two more severe alleles significantly decreasing the catalytic efficiency of actin-activated ATP hydrolysis. The observed reductions in actin motility and catalytic efficiency may serve as the mechanistic basis of the progressive myofibrillar disarray observed in the Drosophila models as well as the prolonged contractile activity responsible for skeletal muscle contractures in FSS patients."

Learn about Freeman-Sheldon syndrome online at the US NIH Genetic and Rare Diseases Information Center (GARD).

Parkinson's disease-related study in human cultured cells and Drosophila

Saridaki T, Nippold M, Dinter E, Roos A, Diederichs L, Fensky L, Schulz JB, Falkenburger BH. FYCO1 mediates clearance of α-synuclein aggregates through a Rab7-dependent mechanism. J Neurochem. 2018 Aug;146(4):474-492. PubMed PMID: 29747217.

From the abstract
: "Parkinson's disease can be caused by mutations in the α-synuclein gene and is characterized by aggregates of α-synuclein protein. We have previously shown that over-expression of the small GTPase Rab7 can induce clearance of α-synuclein aggregates. In this study, we investigate which Rab7 effectors mediate this effect. To model Parkinson's disease, we expressed the pathogenic A53T mutant of α-synuclein in HEK293T cells and Drosophila melanogaster. ... Coexpression of FYCO1 in the fly model decreased α-synuclein aggregates as shown by the filter trap assay and rescued the locomotor deficit resulting from neuronal A53T-α-synuclein expression. ..."

Tuesday, June 25, 2019

Fly model of FTD used to identify compound reported to reduce toxic phosphorylated tau

Shim KH, Kim SH, Hur J, Kim DH, Demirev AV, Yoon SY. Small-molecule drug screening identifies drug Ro 31-8220 that reduces toxic phosphorylated tau in Drosophila melanogaster. Neurobiol Dis. 2019 Jun 21:104519. doi: 10.1016/j.nbd.2019.104519. PMID: 31233882.

Abstract: "The intraneuronal aggregates of hyperphosphorylated and misfolded tau (neurofibrillary tangles, NFTs) cause a stereotypical spatiotemporal Alzheimer's disease (AD) progression that correlates with the severity of the associated cognitive decline. Kinase activity contributes to the balance between neuron survival and cell death. Hyperactivation of kinases including the conventional protein kinase C (PKC) is a defective molecular event accompanying associative memory loss, tau phosphorylation, and progression of AD or related neurodegenerative diseases. Here, we investigated the ability of small therapeutic compounds (a custom library) to improve tau-induced rough-eye phenotype in a Drosophila melanogaster model of frontotemporal dementia. We also assessed the tau phosphorylation in vivo and selected hit compounds. Among the potential hits, we investigated Ro 31-8220, described earlier as a potent PKCα inhibitor. Ro 31-8220 robustly improved the rough-eye phenotype, reduced phosphorylated tau species in vitro and in vivo, reversed tau-induced memory impairment, and improved the fly motor functions. In a human neuroblastoma cell line, Ro 31-8220 reduced the PKC activity and the tau phosphorylation pattern, but we also have to acknowledge the compound's wide range of biological activity. Nevertheless, Ro 31-8220 is a novel therapeutic mitigator of tau-induced neurotoxocity."

Monday, June 24, 2019

Drugs, stones, and flies -- high-throughput screen in fly disease model identifies potential therapeutic for prevention of kidney stones

Ali SN, Dayarathna TK, Ali AN, Osumah T, Ahmed M, Cooper TT, Power NE, Zhang D, Kim D, Kim R, St Amant A, Hou J, Tailly T, Yang J, Luyt L, Spagnuolo PA, Burton JP, Razvi H, Leong HS. Drosophila melanogaster as a function-based high-throughput screening model for antinephrolithiasis agents in kidney stone patients. Dis Model Mech. 2018 Nov 16;11(11). pii: dmm035873. doi: 10.1242/dmm.035873. PubMed PMID: 30082495; PubMed Central PMCID: PMC6262805.

Abstract: "Kidney stone disease involves the aggregation of stone-forming salts consequent to solute supersaturation in urine. The development of novel therapeutic agents for this predominantly metabolic and biochemical disorder have been hampered by the lack of a practical pre-clinical model amenable to drug screening. Here, Drosophila melanogaster, an emerging model for kidney stone disease research, was adapted as a high-throughput functional drug screening platform independent of the multifactorial nature of mammalian nephrolithiasis. Through functional screening, the therapeutic potential of a novel compound commonly known as arbutin that specifically binds to oxalate, a key component of kidney calculi, was identified. Through isothermal titration calorimetry, high-performance liquid chromatography and atomic force microscopy, arbutin was determined to interact with calcium and oxalate in both free and bound states, disrupting crystal lattice structure, growth and crystallization. When used to treat patient urine samples, arbutin significantly abrogated calculus formation in vivo and outperformed potassium citrate in low pH urine conditions, owing to its oxalate-centric mode of action. The discovery of this novel antilithogenic compound via D. melanogaster, independent of a mammalian model, brings greater recognition to this platform, for which metabolic features are primary outcomes, underscoring the power of D. melanogaster as a high-throughput drug screening platform in similar disorders. This is the first description of the use of D. melanogaster as the model system for a high-throughput chemical library screen. This article has an associated First Person interview with the first authors of the paper."

Drosophila as "a fundamental genetic model to identify new disease-causing variants" -- review from Bellen, Wangler & Yamamoto

Bellen HJ, Wangler MF, Yamamoto S. The fruit fly at the interface of diagnosis and pathogenic mechanisms of rare and common human diseases. Hum Mol Genet. 2019 Jun 22. pii: ddz135. doi: 10.1093/hmg/ddz135. PubMed PMID: 31227826.

Abstract: "Drosophila melanogaster is a unique, powerful genetic model organism for studying a broad range of biological questions. Human studies that probe the genetic causes of rare and undiagnosed diseases using massive-parallel sequencing often require complementary gene function studies to determine if and how rare variants affect gene function. These studies also provide inroads to disease mechanisms and therapeutic targets. In this review we discuss strategies for functional studies of rare human variants in Drosophila. We focus on our experience in establishing a Drosophila Core for the Model Organisms Screening Center (MOSC) for the Undiagnosed Diseases Network (UDN) and concurrent fly studies with other large genomic rare disease research efforts such as the Centers for Mendelian Genomics (CMG). We outline four major strategies that use the latest technology in fly genetics to understand the impact of human variants on gene function. We also mention general concepts in probing disease mechanisms, therapeutics and using rare disease to understand common diseases. Drosophila is and will continue to be a fundamental genetic model to identify new disease-causing variants, pathogenic mechanisms and drugs that will impact medicine."