Tuesday, April 30, 2019

Fly model related to Fragile X syndrome used to explore cellular mechanism

Sears JC, Choi WJ, Broadie K. Fragile X Mental Retardation Protein positively regulates PKA anchor Rugose and PKA activity to control actin assembly in learning/memory circuitry. Neurobiol Dis. 2019 Feb 13;127:53-64. PMID: 30771457.

From the abstract: "Recent work shows Fragile X Mental Retardation Protein (FMRP) drives the translation of very large proteins (>2000 aa) mediating neurodevelopment. Loss of function results in Fragile X syndrome (FXS), the leading heritable cause of intellectual disability (ID) and autism spectrum disorder (ASD). Using the Drosophila FXS disease model, we discover FMRP positively regulates the translation of the very large A-Kinase Anchor Protein (AKAP) Rugose (>3000 aa), homolog of ASD-associated human Neurobeachin (NBEA). ... Here, we link two ASD states; Neurobeachin (NBEA) associated ASD and Fragile X syndrome (FXS), the most common inherited ASD. Using established Drosophila disease models, we find Fragile X Mental Retardation Protein (FMRP) positively regulates translation of NBEA homolog Rugose, consistent with a recent advance showing FMRP promotes translation of very large proteins associated with ASD."

Cross-species approach to "mining druggable modifiers of disease-associated proteins"

Huichalaf CH, Al-Ramahi I, Park KW, Grunke SD, Lu N, Haro M, El-Zein K, Gallego-Flores T, Perez AM, Jung SY, Botas J, Zoghbi HY, Jankowsky JL. Cross-species genetic screens to identify kinase targets for APP reduction in Alzheimer's disease. Hum Mol Genet. 2019 Feb 12. PMID: 30753434.

Abstract: "An early hallmark of Alzheimer's disease is the accumulation of amyloid-β, inspiring numerous therapeutic strategies targeting this peptide. An alternative approach is to destabilize the amyloid precursor protein (APP) from which Aβ is derived. We interrogated innate pathways governing APP stability using a siRNA screen for modifiers whose own reduction diminished APP in human cell lines and transgenic Drosophila. As proof-of-principle, we validated PKCβ - a known modifier identified by the screen - in an APP transgenic mouse model. PKCβ was genetically targeted using a novel AAV shuttle vector to deliver microRNA-adapted shRNA via intracranial injection. In vivo reduction of PKCβ initially diminished APP and delayed plaque formation. Despite persistent PKCβ suppression, the effect on APP and amyloid diminished over time. Our study advances this approach for mining druggable modifiers of disease-associated proteins, while cautioning that prolonged in vivo validation may be needed to reveal emergent limitations on efficacy."

Flies used as living test tube to study genes implicated in long lifespan

Xiao FH, Chen XQ, Yu Q, Ye Y, Liu YW, Yan D, Yang LQ, Chen G, Lin R, Yang L, Liao X, Zhang W, Zhang W, Tang NL, Wang XF, Zhou J, Cai WW, He YH, Kong QP. Transcriptome evidence reveals enhanced autophagy-lysosomal function in centenarians. Genome Res. 2018 Nov;28(11):1601-1610. PMID: 30352807; PMCID: PMC6211641.

From the abstract: "... Here, we analyzed the transcriptomes of 76 centenarians, 54 centenarian-children, and 41 spouses of centenarian-children by RNA sequencing and found that ... the autophagy-lysosomal pathway is significantly up-regulated. Overexpression of several genes from this pathway, CTSB, ATP6V0C, ATG4D, and WIPI1, could promote autophagy and delay senescence in cultured IMR-90 cells, while overexpression of the Drosophila homolog of WIPI1, Atg18a, extended the life span in transgenic flies. ... these findings ... suggest that the enhanced waste-cleaning activity via autophagy may serve as a conserved mechanism to prolong the life span from Drosophila to humans."

New fly model for understanding cognitive deficits associated with disruption of FOXP genes

Castells-Nobau A, Eidhof I, Fenckova M, Brenman-Suttner DB, Scheffer-de Gooyert JM, Christine S, Schellevis RL, van der Laan K, Quentin C, van Ninhuijs L, Hofmann F, Ejsmont R, Fisher SE, Kramer JM, Sigrist SJ, Simon AF, Schenck A. Conserved regulation of neurodevelopmental processes and behavior by FoxP in Drosophila. PLoS One. 2019 Feb 12;14(2):e0211652. PMID: 30753188; PMCID: PMC6372147.

From the abstract: "FOXP proteins form a subfamily of evolutionarily conserved transcription factors ... In humans, mutations in FOXP1 and FOXP2 have been implicated in cognitive deficits including intellectual disability and speech disorders. Drosophila exhibits a single ortholog, called FoxP ... Here we show that the dimerization property required for mammalian FOXP function is conserved in Drosophila. In flies, FoxP is enriched in the adult brain, showing strong expression in ~1000 neurons of cholinergic, glutamatergic and GABAergic nature. ... At the cellular level, ... Drosophila FoxP is required in larvae for synaptic morphogenesis at axonal terminals of the neuromuscular junction and for dendrite development of dorsal multidendritic sensory neurons. In the developing brain, we find that FoxP plays important roles in α-lobe mushroom body formation. Finally, at a behavioral level, we show that Drosophila FoxP is important for locomotion, habituation learning and social space behavior of adult flies. Our work shows that Drosophila FoxP is important for regulating several neurodevelopmental processes and behaviors that are related to human disease or vertebrate disease model phenotypes. This suggests a high degree of functional conservation with vertebrate FOXP orthologues and established flies as a model system for understanding FOXP related pathologies."

Article addresses the importance of standardized diets in fly studies related to nutrition research

Lüersen K, Röder T, Rimbach G. Drosophila melanogaster in nutrition research-the importance of standardizing experimental diets. Genes Nutr. 2019 Feb 1;14:3. doi: 10.1186/s12263-019-0627-9. eCollection 2019. Review. PubMed PMID: 30766617; PubMed Central PMCID: PMC6359822.

Abstract: "The fruit fly Drosophila melanogaster has been increasingly recognized as an important model organism in nutrition research. In order to conduct nutritional studies in fruit flies, special attention should be given to the composition of the experimental diets. Besides complex diets, which are often based on maize, yeast, sucrose, and agar, Drosophila can be also fed chemically defined diets. These so-called holidic diets are standardized in terms of their macro- and micronutrient composition although the quantitative nutrient requirements of flies have yet not been fully established and warrant further investigations. For instance, only few studies address the fatty acid, vitamin, mineral, and trace element requirements of fruit flies. D. melanogaster may be also of interest in the field of nutritional medicine. Diet-induced diabetes and obesity models have been established, and in this context, often, the so-called high-fat and high-sugar diets are fed. However, the composition of these diets is not sufficiently defined and varies between studies. A consensus within the scientific community needs to be reached to standardize the exact composition of experimental complex and holidic diets for D. melanogaster in nutrition research. Since D. melanogaster is an established valuable model system for numerous human diseases, standardized diets are also a prerequisite to conduct diet-disease interaction studies. We suggest that a comprehensive approach, which combines deep phenotyping with disease-related Drosophila models under defined dietary conditions, might lead to the foundation of a so-called fly clinic."

Autophagy, lysosomes, and ALS/FTD

Şentürk M, Mao D, Bellen HJ. Loss of proteins associated with amyotrophic lateral sclerosis affect lysosomal acidification via different routes. Autophagy. 2019 Apr 28:1-3. PubMed
PMID: 31032688.

Abstract: "Abnormal accumulation of proteins is a hallmark of a variety of neurological diseases including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Maintenance of protein homeostasis (proteostasis) in neurons via proteasomal and macroautophagy/autophagy-lysosomal degradation is thought to be central for proper neuronal function and survival. We recently reported evolutionarily conserved roles for two ALS-linked proteins, UBQLN2 (ubiquilin 2) and VAPB, in regulation of lysosomal degradation. Ubiquilins are required for v-ATPase-mediated lysosomal acidification, whereas VAPs are required for the PtdIns4P-mediated endo-lysosomal trafficking pathway."

Review -- Drosophila models for the study of motor neuron diseases

Walters R, Manion J, Neely GG. Dissecting Motor Neuron Disease With Drosophila melanogaster. Front Neurosci. 2019 Apr 12;13:331. doi: 10.3389/fnins.2019.00331. PMID: 31031583; PMCID: PMC6473072.

Abstract: "Motor Neuron Disease (MND) typically affects patients during the later stages of life, and thus, MND is having an increasingly devastating impact on diagnosed individuals, their families and society. The umbrella term MND refers to diseases which cause the progressive loss of upper and/or lower motor neurons and a subsequent decrease in motor ability such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA). The study of these diseases is complex and has recently involved the use of genome-wide association studies (GWAS). However, in the case of MND, it has been difficult to identify the complex genetics involved in subtypes, and functional investigation of new candidate disease genes is warranted. Drosophila is a powerful model for addressing these complex diseases. The UAS/Gal4/Gal80 system allows for the upregulation of Drosophila genes, the "knockdown" of genes and the ectopic expression of human genes or mutations in a tissue-specific manner; often resulting in Drosophila models which exhibit typical MND disease pathologies. These can then be further interrogated to identify disease-modifying genes or mutations and disease pathways. This review will discuss two common MNDs and the current Drosophila models which are being used to research their genetic basis and the different pathologies of MND."

Thursday, April 25, 2019

Fly studies contribute to analysis of human gene variants associated with body-fat distribution

Justice AE, Karaderi T, Highland HM, et al. Protein-coding variants implicate novel genes related to lipid homeostasis contributing to body-fat distribution. Nat Genet. 2019 Mar;51(3):452-469. doi: 10.1038/s41588-018-0334-2. PMID: 30778226.

From the abstract: "... We analyzed the association of body-fat distribution ... with 228,985 predicted coding and splice site variants available on exome arrays in up to 344,369 individuals from five major ancestries (discovery) and 132,177 European-ancestry individuals (validation). We identified 15 common ... and nine low-frequency or rare ... novel variants. Pathway/gene set enrichment analyses identified lipid particle, adiponectin, abnormal white adipose tissue physiology and bone development and morphology as important contributors to fat distribution ... In functional follow-up analyses, specifically in Drosophila RNAi-knockdowns, we observed a significant increase in the total body triglyceride levels for two genes (DNAH10 and PLXND1). We implicate novel genes in fat distribution, stressing the importance of interrogating low-frequency and protein-coding variants."

Monday, April 22, 2019

Three reports related to Alzheimer's disease

Feng G, Pang J, Yi X, Song Q, Zhang J, Li C, He G, Ping Y. Down-Regulation of K(V)4 Channel in Drosophila Mushroom Body Neurons Contributes to Aβ42-Induced Courtship Memory Deficits. Neuroscience. 2018 Feb 1;370:236-245. doi: 10.1016/j.neuroscience.2017.06.008. Epub 2017 Jun 13. PubMed PMID: 28627422.

From the abstract: "Accumulation of amyloid-β (Aβ) is widely believed to be an early event in the pathogenesis of Alzheimer's disease (AD). Kv4 is an A-type K+ channel, and our previous report shows the degradation of Kv4, induced by the Aβ42 accumulation, may be a critical contributor to the hyperexcitability of neurons in a Drosophila AD model. Here, we used well-established courtship memory assay to investigate the contribution of the Kv4 channel to short-term memory (STM) deficits in the Aβ42-expressing AD model. ... These data highlight causal relationship between region-specific Kv4 degradation and age-dependent learning decline in the AD model, and provide a mechanism for the disturbed cognitive function in AD."

Lee BI, Suh YS, Chung YJ, Yu K, Park CB. Shedding Light on Alzheimer's β-Amyloidosis: Photosensitized Methylene Blue Inhibits Self-Assembly of β-Amyloid Peptides and Disintegrates Their Aggregates. Sci Rep. 2017 Aug 8;7(1):7523. doi: 10.1038/s41598-017-07581-2. PubMed PMID: 28790398; PubMed Central PMCID: PMC5548810.

From the abstract: "Abnormal aggregation of β-amyloid (Aβ) peptides is a major hallmark of Alzheimer's disease (AD). ... Among many candidate chemicals, methylene blue (MB) has proved its therapeutic potential for AD in a number of in vitro and in vivo studies; but the result of recent clinical trials performed with MB and its derivative was negative. Here ... we first report that photoexcited MB molecules can block Aβ42 aggregation in vitro. Furthermore, our in vivo study using Drosophila AD model demonstrates that photoexcited MB is highly effective in suppressing synaptic toxicity ... Our work suggests that light illumination can provide an opportunity to boost the efficacies of MB toward photodynamic therapy of AD in future."

Papanikolopoulou K, Grammenoudi S, Samiotaki M, Skoulakis EMC. Differential effects of 14-3-3 dimers on Tau phosphorylation, stability and toxicity in vivo. Hum Mol Genet. 2018 Jul 1;27(13):2244-2261. doi: 10.1093/hmg/ddy129. PubMed PMID: 29659825.

From the abstract: "Neurodegenerative dementias collectively known as Tauopathies involve aberrant phosphorylation and aggregation of the neuronal protein Tau. ... We use .. the Drosophila system to investigate in vivo whether 14-3-3s are causal or synergistic with Tau accumulation in precipitating pathogenesis. ... Our collective data demonstrate the complexity of 14-3-3/Tau interactions in vivo and suggest that 14-3-3 attenuation is not appropriate ameliorative treatment of Tauopathies. ..."

From the blog author

Dear Readers,

Wow, it's hard to keep up with the literature! It's exciting to see so many reports using the fly to study human diseases. I have been trying to catch up recently. But it's quite a task. Gaps will remain.

Please feel free to reach out if you have a suggestion of a paper from your lab or another lab that should be added. I've already added papers based on suggestions like this. Don't be shy.

Kind regards,
Your blog author, S.E. Mohr

High-sugar diet, diabetes and the innate immune system

Yu S, Zhang G, Jin LH. A high-sugar diet affects cellular and humoral immune responses in Drosophila. Exp Cell Res. 2018 Jul 15;368(2):215-224. PMID: 29727694.

From the abstract: "A high-sugar diet (HSD) induces Type 2 diabetes (T2D) and obesity, which severely threaten human health. ... few studies have focused on the relationship between a HSD and the innate immune response in Drosophila. In this study, we fed flies a high-sucrose diet and observed defects in the phagocytosis of latex beads and B. bassiana spores. The actin cytoskeleton was also disrupted in hemocytes from HSD-fed larvae. Furthermore, HSD induced the differentiation of lamellocytes in the lymph gland and circulating hemolymph, which rarely occurs in healthy bodies, via JNK signaling. In addition, the Toll and JNK pathways were excessively activated in the fat bodies of HSD-fed larvae, and a large number of dead cells were observed. Finally, HSD induced the aberrant activation of the innate immune system, including inflammation. Our results have established a connection between T2D and the innate immune response."

Fly models of mitochondrial disorders "with different levels of severity" useful for screening for new therapeutics

Foriel S, Renkema GH, Lasarzewski Y, Berkhout J, Rodenburg RJ, Smeitink JAM, Beyrath J, Schenck A. A Drosophila Mitochondrial Complex I Deficiency Phenotype Array. Front Genet. 2019 Mar 27;10:245. PMID: 30972103; PMCID: PMC6445954.

Abstract: "Mitochondrial diseases are a group of rare life-threatening diseases often caused by defects in the oxidative phosphorylation system. No effective treatment is available for these disorders. Therapeutic development is hampered by the high heterogeneity in genetic, biochemical, and clinical spectra of mitochondrial diseases and by limited preclinical resources to screen and identify effective treatment candidates. Alternative models of the pathology are essential to better understand mitochondrial diseases and to accelerate the development of new therapeutics. The fruit fly Drosophila melanogaster is a cost- and time-efficient model that can recapitulate a wide range of phenotypes observed in patients suffering from mitochondrial disorders. We targeted three important subunits of complex I of the mitochondrial oxidative phosphorylation system with the flexible UAS-Gal4 system and RNA interference (RNAi): NDUFS4 (ND-18), NDUFS7 (ND-20), and NDUFV1 (ND-51). Using two ubiquitous driver lines at two temperatures, we established a collection of phenotypes relevant to complex I deficiencies. Our data offer models and phenotypes with different levels of severity that can be used for future therapeutic screenings. These include qualitative phenotypes that are amenable to high-throughput drug screening and quantitative phenotypes that require more resources but are likely to have increased potential and sensitivity to show modulation by drug treatment."

Models and methods -- fly research related to heart disease

Bell KM, Kronert WA, Huang A, Bernstein SI, Swank DM. The R249Q hypertrophic cardiomyopathy myosin mutation decreases contractility in Drosophila by impeding force production. J Physiol. 2019 Apr 4. doi: 10.1113/JP277333. PMID: 30950055.

From the abstract: "... Hypertrophic cardiomyopathy (HCM) is a genetic disease that causes thickening of the heart's ventricular walls and is a leading cause of sudden cardiac death. HCM is caused by missense mutations in muscle proteins including myosin ... We investigated the disease mechanism for HCM myosin mutation R249Q by expressing it in the indirect flight muscle of Drosophila melanogaster and measuring alterations to muscle and flight performance. Muscle mechanical analysis revealed R249Q decreased muscle power production due to slower muscle kinetics and decreased force production; force production was reduced because fewer mutant myosin cross-bridges were bound simultaneously to actin. ... Overall, our results do not support the increased contractility hypothesis. Instead, our results suggest the ventricular hypertrophy for human R249Q mutation is a compensatory response to decreases in heart muscle power output."

Lee CY, Wang HJ, Jhang JD, Cho IC. Automated drosophila heartbeat counting based on image segmentation technique on optical coherence tomography. Sci Rep. 2019 Apr 3;9(1):5557. PMID: 30944361; PMCID: PMC6447591.

From the abstract: "... studies on drosophila cardiac have improved our understanding of human cardiovascular disease. Drosophila cardiac consist of five circular chambers: a conical chamber (CC) and four ostia sections (O1-O4). ... An automated drosophila heartbeat counting algorithm based on the chamber segmentation is developed for OCT in this study. This algorithm has two parts: automated chamber segmentation and heartbeat counting. In addition, this study proposes a principal components analysis (PCA)-based supervised learning method for training the chamber contours to make chamber segmentation more accurate. ... The average calculated heart rates of two-week and six-week drosophila were about 4.77 beats/s and 4.73 beats/s, respectively, which was consistent with the results of manual counting."

Saturday, April 20, 2019

Reports related to neurodegenerative and neuromuscular disorders

Bondar VV, Adamski CJ, Onur TS, Tan Q, Wang L, Diaz-Garcia J, Park J, Orr HT, Botas J, Zoghbi HY. PAK1 regulates ATXN1 levels providing an opportunity to modify its toxicity in spinocerebellar ataxia type 1. Hum Mol Genet. 2018 Aug 15;27(16):2863-2873.PMID: 29860311; PMCID: PMC6077814.

From the abstract: "... Loss-of-function of fly Pak3 or Pak1, whose mammalian homologs belong to Group I of PAK proteins, reduces ATXN1 levels, and accordingly, improves disease pathology in a Drosophila model of SCA1. Knockdown of PAK1 potently reduces ATXN1 levels in mammalian cells ... this study identifies PAK signaling as a distinct molecular pathway that regulates ATXN1 levels and presents a promising opportunity to pursue for developing potential therapeutics for SCA1."

Adusumalli S, Ngian ZK, Lin WQ, Benoukraf T, Ong CT. Increased intron retention is a post-transcriptional signature associated with progressive aging and Alzheimer's disease. Aging Cell. 2019 Mar 13:e12928. PMID: 30868713.

From the abstract: "Intron retention (IR) by alternative splicing is a conserved regulatory mechanism that can affect gene expression and protein function during adult development and age-onset diseases. ... By profiling the transcriptome of Drosophila head cells at different ages, we observed a significant increase in IR events for many genes during aging. ... our results suggest that an increased IR is an conserved signature that is associated with aging. ... changes of IR pattern during aging may regulate the transition from healthy to pathological state in late-onset sporadic AD."

Wang TH, Wang SY, Wang XD, Jiang HQ, Yang YQ, Wang Y, Cheng JL, Zhang CT, Liang WW, Feng HL. Fisetin Exerts Antioxidant and Neuroprotective Effects in Multiple Mutant hSOD1 Models of Amyotrophic Lateral Sclerosis by Activating ERK. Neuroscience. 2018 May 21;379:152-166. PMID: 29559385.

From the abstract: "Oxidative stress exhibits a central role in the course of amyotrophic lateral sclerosis (ALS) ... Fisetin, a natural antioxidant, has shown benefits in varied neurodegenerative diseases. ... Three different hSOD1-related mutant models were used ... results indicate that fisetin protects cells from ROS damage and improves the pathological behaviors caused by oxidative stress in disease models related to SOD1 gene mutations ... providing a potential treatment for ALS."

Lakkappa N, Krishnamurthy PT, M D P, Hammock BD, Hwang SH. Soluble epoxide hydrolase inhibitor, APAU, protects dopaminergic neurons against rotenone induced neurotoxicity: Implications for Parkinson's disease. Neurotoxicology. 2019 Jan;70:135-145. PMID: 30472438.

From the abstract: "Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid, play a crucial role in cytoprotection by attenuating oxidative stress, inflammation and apoptosis. EETs are rapidly metabolised in vivo by the soluble epoxide hydrolase (sEH). Increasing the half life of EETs by inhibiting the sEH enzyme is a novel strategy for neuroprotection. ... sEH inhibitors APAU was screened in silico and further evaluated for their antiparkinson activity against rotenone (ROT) induced neurodegeneration in N27 dopaminergic cell line and Drosophila melanogaster model of Parkinson disease (PD). ..."

 Krench M, Cho RW, Littleton JT. A Drosophila model of Huntington disease-like 2 exhibits nuclear toxicity and distinct pathogenic mechanisms from Huntington disease. Hum Mol Genet. 2016 Aug 1;25(15):3164-3177. PMID: 27288455; PMCID: PMC5179919.

From the abstract: "Huntington disease-like 2 (HDL2) and Huntington disease (HD) are adult-onset neurodegenerative diseases ... we generated and characterized a Drosophila HDL2 model and compared it with a previously established HD model. We find that neuronal expression of HDL2-Q15 is not toxic, while the expression of an expanded HDL2-Q138 protein is lethal. HDL2-Q138 forms large nuclear aggregates, with only smaller puncta observed in the cytoplasm. This is in contrast to what is observed in a Drosophila model of HD, where polyQ aggregates localize exclusively to the cytoplasm. ... We conclude that while HD and HDL2 have similar clinical profiles, distinct pathogenic mechanisms are likely to drive toxicity in Drosophila models of these disorders."

Poska H, Haslbeck M, Kurudenkandy FR, Hermansson E, Chen G, Kostallas G, Abelein A, Biverstål H, Crux S, Fisahn A, Presto J, Johansson J. Dementia-related Bri2 BRICHOS is a versatile molecular chaperone that efficiently inhibits Aβ42 toxicity in Drosophila. Biochem J. 2016 Oct 15;473(20):3683-3704. PMID: 27514716.

From the abstract: "Formation of fibrils of the amyloid-β peptide (Aβ) is suggested to play a central role in neurodegeneration in Alzheimer's disease (AD), for which no effective treatment exists. The BRICHOS domain is a part of several disease-related proproteins ... we find that transgenic expression of the Bri2 BRICHOS domain in the Drosophila central nervous system (CNS) or eyes efficiently inhibits Aβ42 toxicity. ... These findings suggest that Bri2 BRICHOS can be a physiologically relevant chaperone for Aβ in the CNS ..."

Rapamycin and succinate dehydrogenase deficiency

Fan F, Sam R, Ryan E, Alvarado K, Villa-Cuesta E. Rapamycin as a potential treatment for succinate dehydrogenase deficiency. Heliyon. 2019 Feb 11;5(2):e01217. PMID: 30805566; PMCID: PMC6374580.

Abstract: "Drosophila melanogaster is a powerful model to study mitochondrial respiratory chain defects, particularly succinate dehydrogenase (SDH) deficiency. Mutations in sdh genes cause degenerative disorders and often lead to death. Therapies for such pathologies are based on a combination of vitamins and dietary supplements, and are rarely effective. In Drosophila, mutations in several of the genes encoding SDH resemble the pathology of SDH deficiency in humans, enabling the Drosophila model to be used in finding treatments for this condition. Here we show that exposure to the drug rapamycin improves the survival of sdh mutant strains, the activity of SDH and the impaired climbing associated with sdh mutations. However, the production of reactive oxygen species, the oxygen consumption of isolated mitochondria and the resistance to hyperoxia were minimally affected. Our results contribute to the current research seeking a treatment for mitochondrial disease."

Arginine and a Drosophila model of von Hippel-Lindau syndrome

Shmueli MD, Levy-Kanfo L, Haj E, Schoenfeld AR, Gazit E, Segal D. Arginine refolds, stabilizes, and restores function of mutant pVHL proteins in animal model of the VHL cancer syndrome. Oncogene. 2019 Feb;38(7):1038-1049. PMID: 30194449.

Abstract: "The von Hippel-Lindau (VHL) syndrome is a rare inherited cancer, caused by mutations in the VHL gene, many of which render the VHL protein (pVHL) unstable. pVHL is a tumor-suppressor protein implicated in a variety of cellular processes, most notably in response to changes in oxygen availability, due to its role as part of an E3-ligase complex which targets the hypoxia-inducible factor (HIF) for degradation. Previously we reported, using in silico and in vitro analyses, that common oncogenic VHL mutations render pVHL less stable than the wild-type protein, distort its core domain and as a result reduce the ability of the protein to bind its target HIF-1α. Among various chemical chaperones tested, arginine was the most effective in refolding mutant of pVHL. Here we examined the consequences of administering L- or D-arginine to a Drosophila VHL model and to human renal carcinoma cells, both expressing misfolded versions of human pVHL. Arginine treatment increased pVHL solubility in both models and increased the half-life of the mutant pVHL proteins in the cell culture. In both models, L- as well as D-arginine enhanced the ability of wild-type pVHL and certain misfolded mutant versions of pVHL to bind ODD, the HIF-derived target peptide, reflecting restoration of pVHL function. Moreover, continuous feeding of Drosophila expressing misfolded versions of pVHL either L- or D-arginine rich diet rescued their lethal phenotype. Collectively, these in vivo results suggest that arginine supplementation should be examined as a potential novel treatment for VHL cancer syndrome."

Don't sleep now, sleep later. Fly study related to insomnia.

Belfer SJ, Bashaw AG, Perlis ML, Kayser MS. A Drosophila model of sleep restriction therapy for insomnia. Mol Psychiatry. 2019 Mar 1. PMID: 30824866.

From the abstract: "Insomnia is the most common sleep disorder among adults ... Cognitive behavioral therapy for insomnia (CBT-I) is the first-line treatment for insomnia; a key component of this intervention is restriction of sleep opportunity, which optimizes matching of sleep ability and opportunity, leading to enhanced sleep drive. Despite the well-documented efficacy of CBT-I, little is known regarding how CBT-I works at a cellular and molecular level to improve sleep ... Here, guided by human behavioral sleep therapies, we developed a Drosophila model for sleep restriction therapy (SRT) of insomnia. We demonstrate that restriction of sleep opportunity through manipulation of environmental cues improves sleep efficiency in multiple short-sleeping Drosophila mutants. The response to sleep opportunity restriction requires ongoing environmental inputs, but is independent of the molecular circadian clock. We apply this sleep opportunity restriction paradigm to aging and Alzheimer's disease fly models, and find that sleep impairments in these models are reversible with sleep restriction, with associated improvement in reproductive fitness and extended lifespan. This work establishes a model to investigate the neurobiological basis of CBT-I, and provides a platform that can be exploited toward novel treatment targets for insomnia."

A review and a research study: Fly models of Fragile X syndrome

Specchia V, Puricella A, D'Attis S, Massari S, Giangrande A, Bozzetti MP. Drosophila melanogaster as a Model to Study the Multiple Phenotypes, Related to Genome Stability of the Fragile-X Syndrome. Front Genet. 2019 Feb 13;10:10. PMID: 30815010; PMCID: PMC6381874.

Abstract: "Fragile-X syndrome is one of the most common forms of inherited mental retardation and autistic behaviors. The reduction/absence of the functional FMRP protein, coded by the X-linked Fmr1 gene in humans, is responsible for the syndrome. Patients exhibit a variety of symptoms predominantly linked to the function of FMRP protein in the nervous system like autistic behavior and mild-to-severe intellectual disability. Fragile-X (FraX) individuals also display cellular and morphological traits including branched dendritic spines, large ears, and macroorchidism. The dFmr1 gene is the Drosophila ortholog of the human Fmr1 gene. dFmr1 mutant flies exhibit synaptic abnormalities, behavioral defects as well as an altered germline development, resembling the phenotypes observed in FraX patients. Therefore, Drosophila melanogaster is considered a good model to study the physiopathological mechanisms underlying the Fragile-X syndrome. In this review, we explore how the multifaceted roles of the FMRP protein have been addressed in the Drosophila model and how the gained knowledge may open novel perspectives for understanding the molecular defects causing the disease and for identifying novel therapeutical targets."

Wang C, Ge L, Wu J, Wang X, Yuan L. MiR-219 represses expression of dFMR1 in Drosophila melanogaster. Life Sci. 2019 Feb 1;218:31-37. PMID: 30528775.

From the abstract: "... Here, we aimed to identify miRNAs regulating FMRP levels in Drosophila. ...
Using online software, we predicted and selected 11 miRNAs potentially acting on the Drosophila fragile X mental retardation 1 (dFMR1) transcript. ... Among the 11 miRNAs screened, miR-219 and miR-960 reduced luciferase gene activity by binding to the 3'-UTR of the dFMR1 transcript. Mutation of the miR-219 or miR-960 binding sites on the transcript resulted in complete or partial elimination of the miRNA-induced repression. Western blots revealed that dFMRP expression was decreased in the miR-219 overexpression model (Elav>miR-219). Drosophila larvae overexpressing miR-219 showed morphological abnormalities at the neuromuscular junction (increased synaptic boutons and synaptic branches). This finding is consistent with some phenotypes observed in dfmr1 mutants. ... Our results suggest that miR-219 regulates dFMR1 expression in Drosophila and is involved in fragile X syndrome pathogenesis. Collectively, these findings expand the current understanding of miRNA-mediated regulation of target molecule-related functions."

Exploration of sleep in a fly model related to schizophrenia

Nagy S, Maurer GW, Hentze JL, Rose M, Werge TM, Rewitz K. AMPK signaling linked to the schizophrenia-associated 1q21.1 deletion is required for neuronal and sleep maintenance. PLoS Genet. 2018 Dec 19;14(12):e1007623. PMID: 30566533; PMCID: PMC6317821.

Abstract: "The human 1q21.1 deletion of ten genes is associated with increased risk of schizophrenia. This deletion involves the β-subunit of the AMP-activated protein kinase (AMPK) complex, a key energy sensor in the cell. Although neurons have a high demand for energy and low capacity to store nutrients, the role of AMPK in neuronal physiology is poorly defined. Here we show that AMPK is important in the nervous system for maintaining neuronal integrity and for stress survival and longevity in Drosophila. To understand the impact of this signaling system on behavior and its potential contribution to the 1q21.1 deletion syndrome, we focused on sleep, an important role of which is proposed to be the reestablishment of neuronal energy levels that are diminished during energy-demanding wakefulness. Sleep disturbances are one of the most common problems affecting individuals with psychiatric disorders. We show that AMPK is required for maintenance of proper sleep architecture and for sleep recovery following sleep deprivation. Neuronal AMPKβ loss specifically leads to sleep fragmentation and causes dysregulation of genes believed to play a role in sleep homeostasis. Our data also suggest that AMPKβ loss may contribute to the increased risk of developing mental disorders and sleep disturbances associated with the human 1q21.1 deletion."

Functional studies in flies help contributed to identification of novel disease-causing variant of GED associated with static encephalopathy

Assia Batzir N, Bhagwat P, Eble T, Liu P, Eng C, Elsea SH, Robak LA, Scaglia F, Goldman A, Dhar SU, Wangler MF. De novo missense variant in the GTPase effector domain (GED) of DNM1L leads to static encephalopathy and seizures. Cold Spring Harb Mol Case Stud. 2019 Mar 8. pii: mcs.a003673. PMID: 30850373.

Abstract: "DNM1L encodes a GTPase of the dynamin superfamily which plays a crucial role in mitochondrial and peroxisomal fission. Pathogenic variants affecting the middle domain and the GTPase domain of DNM1L have been implicated in encephalopathy due to defective mitochondrial and peroxisomal fission 1 (EMPF1, MIM #614388). Patients show variable phenotypes ranging from severe hypotonia leading to death in the neonatal period to developmental delay/regression, with or without seizures. Familial pathogenic variants in the GTPase domain have also been associated with isolated optic atrophy. We present a 27 year old woman with static encephalopathy, a history of seizures and nystagmus, in whom a novel de novo heterozygous variant was detected in the GTPase Effector Domain (GED) of DNM1L (c.2072A>G, p.Tyr691Cys). Functional studies in Drosophila demonstrate large, abnormally-distributed peroxisomes and mitochondria, an effect very similar to that of middle domain missense alleles observed in pediatric subjects with EMPF1. To our knowledge, not only is this the first report of a disease-causing variant in the GED domain in humans, but this is also the oldest living individual reported with EMPF1. Longitudinal data of this kind helps to expand our knowledge of the natural history of a growing list of DNM1L-related disorders."

Two fly studies related to obesity -- effects of high-fat diet

Stobdan T, Sahoo D, Azad P, Hartley I, Heinrichsen E, Zhou D, Haddad GG. High fat diet induces sex-specific differential gene expression in Drosophila melanogaster. PLoS One. 2019 Mar 2;14(3):e0213474. PMID: 30861021; PMCID: PMC6413938.

From the abstract: "Currently about 2 billion adults globally are estimated to be overweight and ~13% of them are obese. High fat diet (HFD) is one of the major contributing factor to obesity ... studies involving Drosophila melanogaster as a model organism can provide insight into the molecular mechanisms involving human disease. ... we investigated the effect of HFD on the transcriptome in the heads and bodies of male and female flies kept on either HFD or regular diet (RD). ... we found that HFD induces a number of responses that are sexually dimorphic in nature. There was a robust transcriptional response consisting of a downregulation of stress-related genes in the heads and glycoside hydrolase activity genes in the bodies of males. In the females, the HFD led to an increased transcriptional change in lipid metabolism. A strong correlation also existed between the takeout gene and hyperphagic behavior in both males and females. ... our data on the transcriptional responses in flies to HFD provides potentially relevant information to human conditions including obesity."

Rivera O, McHan L, Konadu B, Patel S, Sint Jago S, Talbert ME. A high-fat diet impacts memory and gene expression of the head in mated female Drosophila melanogaster. J Comp Physiol B. 2019 Apr;189(2):179-198. PMID: 30810797.

From the abstract: "Obesity predisposes humans to a range of life-threatening comorbidities ... Obesity also aggravates neural pathologies, such as Alzheimer's disease, but this class of comorbidity is less understood. When Drosophila melanogaster (flies) are exposed to high-fat diet (HFD) by supplementing a standard medium with coconut oil, they adopt an obese phenotype of decreased lifespan, increased triglyceride storage, and hindered climbing ability. ... Our objective was to establish the obesity phenotype in Drosophila and identify a potential correlation, if any, between obesity and neurological decline ... We found that mated female w1118 flies exposed to HFD maintained an obese phenotype throughout adult life starting at 7 days, evidenced by increased triglyceride stores, diminished life span, and impeded climbing ability. While climbing ability worsened cumulatively between 7 and 14 days of exposure to HFD, there was no corresponding alteration in triglyceride content. Microarray analysis of the mated female w1118 fly head revealed HFD-induced changes in expression of genes with functions in memory, metabolism, olfaction, mitosis, cell signaling, and motor function. Meanwhile, an Aversive Phototaxis Suppression assay in mated female flies indicated reduced ability to recall an entrained memory 6 h after training. Overall, our results support the suitability of mated female flies for examining connections between diet-induced obesity and nervous or neurobehavioral pathology, and provide many directions for further investigation."

Results of studies in human cells, zebrafish and fly suggest that variants in ZBTB11 can result in intellectual disability

Fattahi Z, Sheikh TI, Musante L, Rasheed M, Taskiran II, Harripaul R, Hu H, Kazeminasab S, Alam MR, Hosseini M, Larti F, Ghaderi Z, Celik A, Ayub M, Ansar M, Haddadi M, Wienker TF, Ropers HH, Kahrizi K, Vincent JB, Najmabadi H. Biallelic missense variants in ZBTB11 can cause intellectual disability in humans. Hum Mol Genet. 2018 Sep 15;27(18):3177-3188. PMID: 29893856.

From the abstract: "Exploring genes and pathways underlying intellectual disability (ID) provides insight into brain development and function ... linkage analysis and next-generation sequencing revealed Zinc Finger and BTB Domain Containing 11 (ZBTB11) as a novel candidate ID gene. ZBTB11 encodes a little-studied transcription regulator, and the two identified missense variants in this study are predicted to disrupt canonical Zn2+-binding residues of its C2H2 zinc finger domain, leading to possible altered DNA binding. Using HEK293T cells transfected with wild-type and mutant GFP-ZBTB11 constructs, we found the ZBTB11 mutants being excluded from the nucleolus, where the wild-type recombinant protein is predominantly localized. Pathway analysis applied to ChIP-seq data deposited in the ENCODE database supports the localization of ZBTB11 in nucleoli, highlighting associated pathways such as ribosomal RNA synthesis, ribosomal assembly, RNA modification and stress sensing, and provides a direct link between subcellular ZBTB11 location and its function. Furthermore, given the report of prominent brain and spinal cord degeneration in a zebrafish Zbtb11 mutant, we investigated ZBTB11-ortholog knockdown in Drosophila melanogaster brain by targeting RNAi using the UAS/Gal4 system. The observed approximate reduction to a third of the mushroom body size-possibly through neuronal reduction or degeneration-may affect neuronal circuits in the brain that are required for adaptive behavior, specifying the role of this gene in the nervous system. In conclusion, we report two ID families segregating ZBTB11 biallelic mutations disrupting Zn2+-binding motifs and provide functional evidence linking ZBTB11 dysfunction to this phenotype."

Fly study explores effects of PolyQ on glia and the blood-brain barrier -- relevance to neurodegenerative diseases

Yeh PA, Liu YH, Chu WC, Liu JY, Sun YH. Glial expression of disease-associated poly-glutamine proteins impairs the blood-brain barrier in Drosophila. Hum Mol Genet. 2018 Jul 15;27(14):2546-2562. PMID: 29726932.

From the abstract: "Expansion of poly-glutamine (polyQ) stretches in several proteins has been linked to neurodegenerative diseases. The effects of polyQ-expanded proteins on neurons have been extensively studied, but their effects on glia remain unclear. We found that expression of distinct polyQ proteins exclusively in all glia or specifically in the blood-brain barrier (BBB) and blood-retina barrier (BRB) glia caused cell-autonomous impairment of BBB/BRB integrity ... Furthermore, we also found that BBB/BRB leakage in Drosophila is reflected in reversed waveform polarity on the basis of electroretinography (ERG) ... The polyQ-expanded protein Atxn3-84Q forms aggregates, induces BBB/BRB leakage, restricts Drosophila lifespan and reduces the level of Repo ... Expression of Repo in BBB/BRB glia can rescue BBB/BRB leakage ... Coexpression of the chaperon HSP40 and HSP70 effectively rescues the effects of Atxn3-84Q ... coexpression of wild-type Atxn3-27Q can also rescue BBB/BRB impairment, suggesting that normal polyQ protein may have a protective function."

Drosophila studies help elucidate the genetic nature of disease-associated variants--study from the NIH Undiagnosed Diseases Network

Liu N, Schoch K, Luo X, Pena LDM, Bhavana VH, Kukolich MK, Stringer S, Powis Z, Radtke K, Mroske C, Deak KL, McDonald MT, McConkie-Rosell A, Markert ML, Kranz PG, Stong N, Need AC, Bick D, Amaral MD, Worthey EA, Levy S; Undiagnosed Diseases Network (UDN), Wangler MF, Bellen HJ, Shashi V, Yamamoto S. Functional variants in TBX2 are associated with a syndromic cardiovascular and skeletal developmental disorder. Hum Mol Genet. 2018 Jul 15;27(14):2454-2465. PMID: 29726930; PMCID: PMC6030957.

Abstract: "The 17 genes of the T-box family are transcriptional regulators that are involved in all stages of embryonic development, including craniofacial, brain, heart, skeleton and immune system. Malformation syndromes have been linked to many of the T-box genes. For example, haploinsufficiency of TBX1 is responsible for many structural malformations in DiGeorge syndrome caused by a chromosome 22q11.2 deletion. We report four individuals with an overlapping spectrum of craniofacial dysmorphisms, cardiac anomalies, skeletal malformations, immune deficiency, endocrine abnormalities and developmental impairments, reminiscent of DiGeorge syndrome, who are heterozygotes for TBX2 variants. The p.R20Q variant is shared by three affected family members in an autosomal dominant manner; the fourth unrelated individual has a de novo p.R305H mutation. Bioinformatics analyses indicate that these variants are rare and predict them to be damaging. In vitro transcriptional assays in cultured cells show that both variants result in reduced transcriptional repressor activity of TBX2. We also show that the variants result in reduced protein levels of TBX2. Heterologous over-expression studies in Drosophila demonstrate that both p.R20Q and p.R305H function as partial loss-of-function alleles. Hence, these and other data suggest that TBX2 is a novel candidate gene for a new multisystem malformation disorder."

Review: Drosophila as a model to study cancer

Mirzoyan Z, Sollazzo M, Allocca M, Valenza AM, Grifoni D, Bellosta P. Drosophila melanogaster: A Model Organism to Study Cancer. Front Genet. 2019 Mar 1;10:51. PMID: 30881374; PMCID: PMC6405444.

Abstract: "Cancer is a multistep disease driven by the activation of specific oncogenic pathways concomitantly with the loss of function of tumor suppressor genes that act as sentinels to control physiological growth. The conservation of most of these signaling pathways in Drosophila, and the ability to easily manipulate them genetically, has made the fruit fly a useful model organism to study cancer biology. In this review we outline the basic mechanisms and signaling pathways conserved between humans and flies responsible of inducing uncontrolled growth and cancer development. Second, we describe classic and novel Drosophila models used to study different cancers, with the objective to discuss their strengths and limitations on their use to identify signals driving growth cell autonomously and within organs, drug discovery and for therapeutic approaches."

Drosophila studies help elucidate metabolic processes relevant to combined D- and L-2HG aciduria (D-/L-2HGA)

Li H, Hurlburt AJ, Tennessen JM. A Drosophila model of combined D-2- and L-2-hydroxyglutaric aciduria reveals a mechanism linking mitochondrial citrate export with oncometabolite accumulation. Dis Model Mech. 2018 Sep 21;11(9). PMID: 30108060; PMCID: PMC6177012.

From the abstract: "The enantiomers of 2-hydroxyglutarate (2HG) are potent regulators of metabolism, chromatin modifications and cell fate decisions. ... The metabolic mechanisms that control 2HG metabolism in vivo are poorly understood. One clue towards how cells regulate 2HG levels has emerged from an inborn error of metabolism known as combined D- and L-2HG aciduria (D-/L-2HGA), which results in elevated D- and L-2HG accumulation. Because this disorder is caused by mutations in the mitochondrial citrate transporter (CIC), citrate must somehow govern 2HG metabolism in healthy cells. ... Here, we use the fruit fly Drosophila melanogaster to elucidate a metabolic link between citrate transport and L-2HG accumulation. Our study reveals that the Drosophila gene scheggia (sea), which encodes the fly CIC homolog, dampens glycolytic flux and restricts L-2HG accumulation. Moreover, we find that sea mutants accumulate excess L-2HG ... which inhibits L-2HG degradation by interfering with L-2HG dehydrogenase activity. This unexpected result demonstrates that citrate indirectly regulates L-2HG stability and reveals a feedback mechanism that coordinates L-2HG metabolism with glycolysis and the tricarboxylic acid cycle. Finally, our study also suggests a potential strategy for preventing L-2HG accumulation in human patients with CIC deficiency."

Sleep defects observed in fly model of mitochondrial encephalomyopathies

Fogle KJ, Mobini CL, Paseos AS, Palladino MJ. Sleep and circadian defects in a Drosophila model of mitochondrial encephalomyopathy. Neurobiol Sleep Circadian Rhythms. 2019 Jan;6:44-52. PMID: 30868108; PMCID: PMC6411073.

From the abstract: "Mitochondrial encephalomyopathies (ME) are complex, incurable diseases characterized by severe bioenergetic distress that can affect the function of all major organ systems but is especially taxing to neuromuscular tissues. Animal models of MEs are rare, but the Drosophila ATP61 mutant ... accurately models progressive human mitochondrial diseases such as Maternally-Inherited Leigh Syndrome (MILS), Neuropathy, Ataxia, and Retinitis Pigmentosa (NARP), and Familial Bilateral Striatal Necrosis (FBSN). While it is established that this model exhibits important hallmarks of ME ... it is unknown whether it exhibits defects in sleep or circadian function. This is a clinically relevant question, as many neurological and neurodegenerative diseases are characterized by such disturbances, which can exacerbate other symptoms and worsen quality of life. ... we found that day-time and night-time activity and sleep are altered through disease progression, and that circadian patterns are disrupted at both the behavioral and neuronal levels. These results establish ATP61 as an important model of sleep and circadian disruption in ME that can be studied mechanistically at the molecular, cellular, and behavioral level to uncover underlying pathophysiology and test novel therapies."

Friday, April 12, 2019

Methods review with workflow diagram, lists of relevant genes and molecular genetic fly stocks, and more, on the use of Drosophila to study retinitis pigmentosa

Lehmann M, Knust E, Hebbar S. Drosophila melanogaster: A Valuable Genetic Model Organism to Elucidate the Biology of Retinitis Pigmentosa. Methods Mol Biol. 2019;1834:221-249. PMID: 30324448.

Abstract: "Retinitis pigmentosa (RP) is a complex inherited disease. It is associated with mutations in a wide variety of genes with many different functions. These mutations impact the integrity of rod photoreceptors and ultimately result in the progressive degeneration of rods and cone photoreceptors in the retina, leading to complete blindness. A hallmark of this disease is the variable degree to which symptoms are manifest in patients. This is indicative of the influence of the environment, and/or of the distinct genetic makeup of the individual.The fruit fly, Drosophila melanogaster, has effectively proven to be a great model system to better understand interconnected genetic networks. Unraveling genetic interactions and thereby different cellular processes is relatively easy because more than a century of research on flies has enabled the creation of sophisticated genetic tools to perturb gene function. A remarkable conservation of disease genes across evolution and the similarity of the general organization of the fly and vertebrate photoreceptor cell had prompted research on fly retinal degeneration. To date six fly models for RP, including RP4, RP11, RP12, RP14, RP25, and RP26, have been established, and have provided useful information on RP disease biology. In this chapter, an outline of approaches and experimental specifications are described to enable utilizing or developing new fly models of RP."

Wednesday, April 10, 2019

FRET-based assay related to Huntington's disease tested in mouse and fly models

Ast A, Buntru A, Schindler F, Hasenkopf R, Schulz A, Brusendorf L, Klockmeier K, Grelle G, McMahon B, Niederlechner H, Jansen I, Diez L, Edel J, Boeddrich A, Franklin SA, Baldo B, Schnoegl S, Kunz S, Purfürst B, Gaertner A, Kampinga HH, Morton AJ, Petersén Å, Kirstein J, Bates GP, Wanker EE. mHTT Seeding Activity: A Marker of Disease Progression and Neurotoxicity in Models of Huntington's Disease. Mol Cell. 2018 Sep 6;71(5):675-688.e6. PMID: 30193095.

From the abstract: "Self-propagating, amyloidogenic mutant huntingtin (mHTT) aggregates may drive progression of Huntington's disease (HD). Here, we report the development of a FRET-based mHTT aggregate seeding (FRASE) assay that enables the quantification of mHTT seeding activity (HSA) ... Application of the FRASE assay revealed HSA in brain homogenates of presymptomatic HD transgenic and knockin mice and its progressive increase with phenotypic changes, suggesting that HSA quantitatively tracks disease progression. Biochemical investigations ... demonstrated that small, rather than large, mHTT structures are responsible for the HSA measured in FRASE assays. Finally, we assessed the neurotoxicity of mHTT seeds in an inducible Drosophila model transgenic for HTTex1. We found a strong correlation between the HSA measured in adult neurons and the increased mortality of transgenic HD flies, indicating that FRASE assays detect disease-relevant, neurotoxic, mHTT structures with severe phenotypic consequences in vivo."

Studies in fly nephrocytes (renal cells) help implicate NUP160 in steroid-resistant nephrotic syndrome (SRNS)

Zhao F, Zhu JY, Richman A, Fu Y, Huang W, Chen N, Pan X, Yi C, Ding X, Wang S, Wang P, Nie X, Huang J, Yang Y, Yu Z, Han Z. Mutations in NUP160 Are Implicated in Steroid-Resistant Nephrotic Syndrome. J Am Soc Nephrol. 2019 Mar 25. PMID: 30910934.

From the abstract: "Studies have identified mutations in >50 genes that can lead to monogenic steroid-resistant nephrotic syndrome (SRNS). The NUP160 gene, which encodes one of the protein components of the nuclear pore complex nucleoporin 160 kD (Nup160), is expressed in both human and mouse kidney cells. Knockdown of NUP160 impairs mouse podocytes in cell culture. Recently, siblings with SRNS and proteinuria in a nonconsanguineous family were found to carry compound-heterozygous mutations in NUP160. ... We identified NUP160 mutations by whole-exome and Sanger sequencing of genomic DNA from a young girl with familial SRNS and FSGS who did not carry mutations in other genes known to be associated with SRNS. We performed in vivo functional validation studies on the NUP160 mutations using a Drosophila model. ... We identified two compound-heterozygous NUP160 mutations, NUP160R1173× and NUP160E803K . We showed that silencing of Drosophila NUP160 specifically in nephrocytes (fly renal cells) led to functional abnormalities, reduced cell size and nuclear volume, and disorganized nuclear membrane structure. These defects were completely rescued by expression of the wild-type human NUP160 gene in nephrocytes. By contrast, expression of the NUP160 mutant allele NUP160R1173× completely failed to rescue nephrocyte phenotypes, and mutant allele NUP160E803K rescued only nuclear pore complex and nuclear lamin localization defects. ... Our findings indicate that NUP160 should be included in the SRNS diagnostic gene panel to identify additional patients with SRNS and homozygous or compound-heterozygous NUP160 mutations and further strengthen the evidence that NUP160 mutations can cause SRNS."

Flies, folic acid, CMT2A -- and a video abstract

Garrido-Maraver J, Celardo I, Costa AC, Lehmann S, Loh SHY, Martins LM. Enhancing folic acid metabolism suppresses defects associated with loss of Drosophila mitofusin. Cell Death Dis. 2019 Mar 25;10(4):288. PMID: 30911005; PMCID: PMC6433915.

Exploration of sensitivity to iron in fly models related to Parkinson's disease

Zhu ZJ, Wu KC, Yung WH, Qian ZM, Ke Y. Differential interaction between iron and mutant alpha-synuclein causes distinctive Parkinsonian phenotypes in Drosophila. Biochim Biophys Acta. 2016 pr;1862(4):518-525. PMID: 26769358.

From the abstract: "Alpha-synuclein aggregation is the central hallmark of both sporadic and familial Parkinson's disease (PD). ... Iron accumulation is invariably found in PD. Recent studies showed that mutant and wild-type alpha-synuclein may have differential interaction with iron and mutant alpha-synuclein toxicity could be preferentially exacerbated by iron. We hence hypothesized that iron overload could selectively influence mutant alpha-synuclein toxicity and disease phenotypes. To test the hypothesis, we investigated if Drosophila melanogaster over-expressing A53T, A30P, and wild-type (WT) alpha-synuclein have different responses to iron treatment. We showed that iron treatment induced similar reduction of survival rate in all flies but induced a more severe motor decline in A53T and A30P mutant alpha-synuclein expressing flies, suggesting interaction between mutant alpha-synuclein and iron. Although no significant difference in total head iron content was found among these flies, we demonstrated that iron treatment induced selective DA neuron loss in motor-related PPM3 cluster only in the flies that express A53T and A30P mutant alpha-synuclein. We provided the first in vivo evidence that iron overload could induce distinctive neuropathology and disease phenotypes in mutant but not WT alpha-synuclein expressing flies ..."

Fly and yeast models help identify genetic variants of WARS2 as disease-causing for a mitochondrial-related leukoencephalopathy

Maffezzini C, Laine I, Dallabona C, Clemente P, Calvo-Garrido J, Wibom R, Naess K, Barbaro M, Falk A, Donnini C, Freyer C, Wredenberg A, Wedell A. Mutations in the mitochondrial tryptophanyl-tRNA synthetase cause growth retardation and progressive leukoencephalopathy. Mol Genet Genomic Med. 2019 Mar 28:e654. PMID: 30920170.

From the abstract: "Mutations in mitochondrial aminoacyl tRNA synthetases form a subgroup of mitochondrial disorders often only perturbing brain function by affecting mitochondrial translation. Here we report two siblings with mitochondrial disease, due to compound heterozygous mutations in the mitochondrial tryptophanyl-tRNA synthetase (WARS2) gene, presenting with severe neurological symptoms but normal mitochondrial function in skeletal muscle biopsies and cultured skin fibroblasts. ... Whole exome sequencing on genomic DNA samples from both subjects and their parents identified two compound heterozygous variants c.833T>G (p.Val278Gly) and c.938A>T (p.Lys313Met) in the WARS2 gene as potential disease-causing variants. We generated patient-derived neuroepithelial stem cells and modeled the disease in yeast and Drosophila melanogaster to confirm pathogenicity. ... Biochemical analysis of patient-derived neuroepithelial stem cells revealed a mild combined complex I and IV defect, while modeling the disease in yeast demonstrated that the reported aminoacylation defect severely affects respiration and viability. Furthermore, silencing of wild type WARS2 in Drosophila melanogaster showed that a partial defect in aminoacylation is enough to cause lethality. ... Our results establish the identified WARS2 variants as disease-causing and highlight the benefit of including human neuronal models, when investigating mutations specifically affecting the nervous system."

Review -- fly models of Myotonic Dustrophy Type 1

Souidi A, Zmojdzian M, Jagla K. Dissecting Pathogenetic Mechanisms and Therapeutic Strategies in Drosophila Models of Myotonic Dystrophy Type 1. Int J Mol Sci. 2018 Dec 18;19(12). PMID: 30567354; PMCID: PMC6321436.

From the abstract: "... Here we provide an overview of insights gained from fruit fly DM1 models, either: (i) fundamental with particular focus on newly identified gene deregulations and their link with DM1 symptoms; or (ii) applied via genetic modifiers and drug screens to identify promising therapeutic targets. ..."

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Fig. 1 from Souidi et al. 2018

Potential new drug repurposing strategy identified using fly model of ALS

Xu W, Bao P, Jiang X, Wang H, Qin M, Wang R, Wang T, Yang Y, Lorenzini I, Liao L, Sattler R, Xu J. Reactivation of nonsense-mediated mRNA decay protects against C9orf72 dipeptide-repeat neurotoxicity. Brain. 2019 Apr 1. pii: awz070. PMID: 30938419.

From the abstract: "Amyotrophic lateral sclerosis is a deleterious neurodegenerative disease without effective treatment options. Recent studies have indicated the involvement of the dysregulation of RNA metabolism in the pathogenesis of amyotrophic lateral sclerosis. Among the various RNA regulatory machineries, nonsense-mediated mRNA decay (NMD) is a stress responsive cellular surveillance system that degrades selected mRNA substrates to prevent the translation of defective or harmful proteins. ... Here we report the inhibition of NMD by arginine-rich dipeptide repeats derived from C9orf72 hexanucleotide repeat expansion, the most common cause of familial amyotrophic lateral sclerosis. Bioinformatic analysis of multiple transcriptome profiles revealed significant overlap of upregulated genes in NMD-defective cells with those in the brain tissues, micro-dissected motor neurons, or induced pluripotent stem cell-derived motor neurons specifically from amyotrophic lateral sclerosis patients carrying C9orf72 hexanucleotide repeat expansion, suggesting the suppression of NMD pathway in these patients. Using Drosophila as a model, we have validated that the C9orf72 hexanucleotide repeat expansion products could lead to the accumulation of the NMD substrates and identified arginine-rich dipeptide repeats ... Remarkably, expression of UPF1, a core gene in the NMD pathway, efficiently blocked neurotoxicity caused by arginine-rich dipeptide repeats in both cellular and Drosophila models. ... UPF2 also ameliorated the degenerative phenotypes in dipeptide repeat-expressing flies ... Finally, after validating tranilast as an NMD-activating drug, we demonstrated the therapeutic potential of this asthma drug in cellular and Drosophila models of C9orf72 dipeptide repeat neurotoxicity. Therefore, our study has revealed a cellular mechanism whereby arginine-rich C9orf72 dipeptide repeats could inhibit NMD activities by reducing the abundance of processing bodies. Furthermore, our results suggested that activation of the NMD pathway could be a potential therapeutic strategy for amyotrophic lateral sclerosis with defective RNA metabolism."

Domain-sensory processing approach--new fly model for study of genetic basis of autism

Vilidaite G, Norcia AM, West RJH, Elliott CJH, Pei F, Wade AR, Baker DH. Autism sensory dysfunction in an evolutionarily conserved system. Proc Biol Sci. 2018 Dec 19;285(1893):20182255. PMID: 30963913.

Abstract: "There is increasing evidence for a strong genetic basis for autism, with many genetic models being developed in an attempt to replicate autistic symptoms in animals. However, current animal behaviour paradigms rarely match the social and cognitive behaviours exhibited by autistic individuals. Here, we instead assay another functional domain-sensory processing-known to be affected in autism to test a novel genetic autism model in Drosophila melanogaster. We show similar visual response alterations and a similar development trajectory in Nhe3 mutant flies (total n = 72) and in autistic human participants (total n = 154). We report a dissociation between first- and second-order electrophysiological visual responses to steady-state stimulation in adult mutant fruit flies that is strikingly similar to the response pattern in human adults with ASD as well as that of a large sample of neurotypical individuals with high numbers of autistic traits. We explain this as a genetically driven, selective signalling alteration in transient visual dynamics. In contrast to adults, autistic children show a decrease in the first-order response that is matched by the fruit fly model, suggesting that a compensatory change in processing occurs during development. Our results provide the first animal model of autism comprising a differential developmental phenotype in visual processing."

Monday, April 8, 2019

Fly studies relevant to myopathies

Jean S, Cox S, Schmidt EJ, Robinson FL, Kiger A. Sbf/MTMR13 coordinates PI(3)P and Rab21 regulation in endocytic control of cellular remodeling. Mol Biol Cell. 2012 Jul;23(14):2723-40. doi: 10.1091/mbc.E12-05-0375. PubMed PMID: 22648168; PubMed Central PMCID: PMC3395661.

Updated in 2019 to include this 2011 study from the same group:

Ribeiro I, Yuan L, Tanentzapf G, Dowling JJ, Kiger A. Phosphoinositide regulation of integrin trafficking required for muscle attachment and maintenance. PLoS Genet. 2011 Feb 10;7(2):e1001295. PMID: 21347281; PMCID: PMC3037412.

From the abstract: "Muscles must maintain cell compartmentalization when remodeled during development and use. How spatially restricted adhesions are regulated with muscle remodeling is largely unexplored. We show that the myotubularin (mtm) phosphoinositide phosphatase is required for integrin-mediated myofiber attachments in Drosophila melanogaster, and that mtm-depleted myofibers exhibit hallmarks of human XLMTM myopathy. ... Importantly, similar integrin localization defects found in human XLMTM myofibers signify conserved MTM1 function in muscle membrane trafficking. Our results indicate that regulation of distinct phosphoinositide pools plays a central role in maintaining cell compartmentalization and attachments during muscle remodeling, and they suggest involvement of Class II PI3-kinase in MTM-related disease.

Preprint: Fly model of neurodegeneration -- exploring LRRK2 and Rab's

New on the BioRxiv preprint server (not yet peer reviewed):

Neurodegeneration caused by LRRK2-G2019S requires Rab10 in select dopaminergic neurons

Stavroula Petridi, C. Adam Middleton, Alison Fellgett, Laura Covill, Amy Stewart, Jack Munns, Friederike Elisabeth Kohrs, P. Robin Hiesinger, Laurence Wilson, Sangeeta Chawla, Christopher J. H. Elliott


From the Significance Statement: "A key question in Parkinson’s is why dopamine neurons die particularly fast in some parts of the substantia nigra. We focused on the commonest Parkinson’s-related mutation, LRRK2-G2019S. ... Rab10 knock-out rescues G2019S-induced visual and movement degeneration, leaving sleep dysfunction unaffected. ... Rab3 is found in a different subset of dopaminergic neurons and shows no synergy with LRRK2-G2019S. We propose that variations in Rab expression contribute to differences in neurodegeneration ..."