Drosophila used in study related to fragile X-associated primary ovariant insufficiency

Fertil Steril. 2021 Sep;116(3):843-854. doi: 10.1016/j.fertnstert.2021.04.021 

Identifying susceptibility genes for primary ovarian insufficiency on the high-risk genetic background of a fragile X premutation. 

Trevino CE, Rounds JC, Charen K, Shubeck L, Hipp HS, Spencer JB, Johnston HR, Cutler DJ, Zwick ME, Epstein MP, Murray A, Macpherson JN, Mila M, Rodriguez-Revenga L, Berry-Kravis E, Hall DA, Leehey MA, Liu Y, Welt C, Warren ST, Sherman SL. Jin P, Allen EG

Abstract:

OBJECTIVE: To identify modifying genes that explains the risk of fragile X-associated primary ovarian insufficiency (FXPOI).

DESIGN: Gene-based, case/control association study, followed by a functional screen of highly ranked genes using a Drosophila model.

SETTING: Participants were recruited from academic and clinical settings.

PATIENT(S): Women with a premutation (PM) who experienced FXPOI at the age of 35 years or younger (n = 63) and women with a PM who experienced menopause at the age of 50 years or older (n = 51) provided clinical information and a deoxyribonucleic acid sample for whole genome sequencing. The functional screen was on the basis of Drosophila TRiP lines.

INTERVENTION(S): Clinical information and a DNA sample were collected for whole genome sequencing.

MAIN OUTCOME MEASURES: A polygenic risk score derived from common variants associated with natural age at menopause was calculated and associated with the risk of FXPOI. Genes associated with the risk of FXPOI were identified on the basis of the P-value from gene-based association test and an altered level of fecundity when knocked down in the Drosophila PM model.

RESULTS: The polygenic risk score on the basis of common variants associated with natural age at menopause explained approximately 8% of the variance in the risk of FXPOI. Further, SUMO1 and KRR1 were identified as possible modifying genes associated with the risk of FXPOI on the basis of an untargeted gene analysis of rare variants.

CONCLUSIONS: In addition to the large genetic effect of a PM on ovarian function, the additive effects of common variants associated with natural age at menopause and the effect of rare modifying variants appear to play a role in FXPOI risk.

DOI: 10.1016/j.fertnstert.2021.04.021
PMID: 34016428

Identification of protein interactors with fly GOLPH3 points to potential targets for cancer and other therapeutics

Cells. 2021 Sep 6;10(9):2336. doi: 10.3390/cells10092336.

Identification of GOLPH3 Partners in Drosophila Unveils Potential Novel Roles in Tumorigenesis and Neural Disorders.

Sechi S, Karimpour-Ghahnavieh A, Frappaolo A, Di Francesco L, Piergentili R, Schininà E, D'Avino PP, Giansanti MG

Author information:
(1)Istituto di Biologia e Patologia Molecolari del CNR, c/o Dipartimento di
Biologia e Biotecnologie, Sapienza Università di Roma, Piazzale A. Moro 5, 00185
Roma, Italy.
(2)Dipartimento di Scienze Biochimiche A. Rossi Fanelli, Sapienza Università di
Roma, Piazzale A. Moro 5, 00185 Roma, Italy.
(3)Department of Pathology, University of Cambridge, Tennis Court Road,
Cambridge CB2 1QP, UK.

Abstract: "Golgi phosphoprotein 3 (GOLPH3) is a highly conserved peripheral membrane protein localized to the Golgi apparatus and the cytosol. GOLPH3 binding to Golgi membranes depends on phosphatidylinositol 4-phosphate [PI(4)P] and regulates Golgi architecture and vesicle trafficking. GOLPH3 overexpression has been correlated with poor prognosis in several cancers, but the molecular mechanisms that link GOLPH3 to malignant transformation are poorly understood. We recently showed that PI(4)P-GOLPH3 couples membrane trafficking with contractile ring assembly during cytokinesis in dividing Drosophila spermatocytes. Here, we use affinity purification coupled with mass spectrometry (AP-MS) to identify the protein-protein interaction network (interactome) of Drosophila GOLPH3 in testes. Analysis of the GOLPH3 interactome revealed enrichment for proteins involved in vesicle-mediated trafficking, cell proliferation and cytoskeleton dynamics. In particular, we found that dGOLPH3 interacts with the Drosophila orthologs of Fragile X mental retardation protein and Ataxin-2, suggesting a potential role in the pathophysiology of disorders of the nervous system. Our findings suggest novel molecular targets associated with GOLPH3 that might be relevant for therapeutic intervention in cancers and other human diseases."

DOI: 10.3390/cells10092336
PMID: 34571985

Study looks at molecular mechsnisms underlying circadian rhythm disruption in fly model of Fragile X syndrome

Neurosci Bull. 2021 Apr 15. doi: 10.1007/s12264-021-00682-z.

Dysregulated CRMP Mediates Circadian Deficits in a Drosophila Model of Fragile X Syndrome.

Zhao J, Xue J, Zhu T, He H, Kang H, Jiang X, Huang W, Duan R

Abstract:

Fragile X syndrome (FXS) is the leading inherited cause of intellectual disability, resulting from the lack of functional fragile X mental retardation protein (FMRP), an mRNA binding protein mainly serving as a translational regulator. Loss of FMRP leads to dysregulation of target mRNAs. The Drosophila model of FXS show an abnormal circadian rhythm with disruption of the output pathway downstream of the clock network. Yet the FMRP targets involved in circadian regulation have not been identified. Here, we identified collapsing response mediator protein (CRMP) mRNA as a target of FMRP. Knockdown of pan-neuronal CRMP expression ameliorated the circadian defects and abnormal axonal structures of clock neurons (ventral lateral neurons) in dfmr1 mutant flies. Furthermore, specific reduction of CRMP in the downstream output insulin-producing cells attenuated the aberrant circadian behaviors. Molecular analyses revealed that FMRP binds with CRMP mRNA and negatively regulates its translation. Our results indicate that CRMP is an FMRP target and establish an essential role for CRMP in the circadian output in FXS Drosophila.

DOI: 10.1007/s12264-021-00682-z
PMID: 33856646

Detailed genetic characterization of a Drosophila fragile X model

Kennedy T, Rinker D, Broadie K. Genetic background mutations drive neural circuit hyperconnectivity in a fragile X syndrome model. BMC Biol. 2020 Jul 30;18(1):94. doi: 10.1186/s12915-020-00817-0. PMID: 32731855.

Abstract:

Background: Neural circuits are initially assembled during development when neurons synapse with potential partners and later refined as appropriate connections stabilize into mature synapses while inappropriate contacts are eliminated. Disruptions to this synaptogenic process impair connectivity optimization and can cause neurodevelopmental disorders. Intellectual disability (ID) and autism spectrum disorder (ASD) are often characterized by synaptic overgrowth, with the maintenance of immature or inappropriate synapses. Such synaptogenic defects can occur through mutation of a single gene, such as fragile X mental retardation protein (FMRP) loss causing the neurodevelopmental disorder fragile X syndrome (FXS). FXS represents the leading heritable cause of ID and ASD, but many other genes that play roles in ID and ASD have yet to be identified.

Results: In a Drosophila FXS disease model, one dfmr150M null mutant stock exhibits previously unreported axonal overgrowths at developmental and mature stages in the giant fiber (GF) escape circuit. These excess axon projections contain both chemical and electrical synapse markers, indicating mixed synaptic connections. Extensive analyses show these supernumerary synapses connect known GF circuit neurons, rather than new, inappropriate partners, indicating hyperconnectivity within the circuit. Despite the striking similarities to well-characterized FXS synaptic defects, this new GF circuit hyperconnectivity phenotype is driven by genetic background mutations in this dfmr150M stock. Similar GF circuit synaptic overgrowth is not observed in independent dfmr1 null alleles. Bulked segregant analysis (BSA) was combined with whole genome sequencing (WGS) to identify the quantitative trait loci (QTL) linked to neural circuit hyperconnectivity. The results reveal 8 QTL associated with inappropriate synapse formation and maintenance in the dfmr150M mutant background.

Conclusions: Synaptogenesis is a complex, precisely orchestrated neurodevelopmental process with a large cohort of gene products coordinating the connectivity, synaptic strength, and excitatory/inhibitory balance between neuronal partners. This work identifies a number of genetic regions that contain mutations disrupting proper synaptogenesis within a particularly well-mapped neural circuit. These QTL regions contain potential new genes involved in synapse formation and refinement. Given the similarity of the synaptic overgrowth phenotype to known ID and ASD inherited conditions, identifying these genes should increase our understanding of these devastating neurodevelopmental disease states.

Methods report -- Drosophila and the study of Fragile X Syndrome

Kong HE, Lim J, Jin P. Application of Drosophila Model Toward Understanding the Molecular Basis of Fragile X Syndrome. Methods Mol Biol. 2019;1942:141-153. PubMed PMID: 30900182.

Flies used to test drug activity predicted using dynamic combinatorial chemistry approach

Canal-Martín A, Sastre J, Sánchez-Barrena MJ, Canales A, Baldominos S, Pascual N, Martínez-González L, Molero D, Fernández-Valle ME, Sáez E, Blanco-Gabella P, Gómez-Rubio E, Martín-Santamaría S, Sáiz A, Mansilla A, Cañada FJ, Jiménez-Barbero J, Martínez A, Pérez-Fernández R. Insights into real-time chemical processes in a calcium sensor protein-directed dynamic library. Nat Commun. 2019 Jun 26;10(1):2798. PMID: 31243268; PMCID: PMC6595003.

Abstract: "Dynamic combinatorial chemistry (DCC) has proven its potential in drug discovery speeding the identification of modulators of biological targets. However, the exchange chemistries typically take place under specific reaction conditions, with limited tools capable of operating under physiological parameters. Here we report a catalyzed protein-directed DCC working at low temperatures that allows the calcium sensor NCS-1 to find the best ligands in situ. Ultrafast NMR identifies the reaction intermediates of the acylhydrazone exchange, tracing the molecular assemblies and getting a real-time insight into the essence of DCC processes at physiological pH. Additionally, NMR, X-ray crystallography and computational methods are employed to elucidate structural and mechanistic aspects of the molecular recognition event. The DCC approach leads us to the identification of a compound stabilizing the NCS-1/Ric8a complex and whose therapeutic potential is proven in a Drosophila model of disease with synaptic alterations."

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."

Study of a fly model of fragile X-syndrome suggests "a potentially general mechanism for intellectual disability"

Franco LM, Okray Z, Linneweber GA, Hassan BA, Yaksi E. Reduced Lateral Inhibition Impairs Olfactory Computations and Behaviors in a Drosophila Model of Fragile X Syndrome. Curr Biol. 2017 Apr 24;27(8):1111-1123. PMID: 28366741; PMCID: PMC5405172.

From the abstract: "Fragile X syndrome (FXS) patients present neuronal alterations that lead to severe intellectual disability, but the underlying neuronal circuit mechanisms are poorly understood. An emerging hypothesis postulates that reduced GABAergic inhibition of excitatory neurons is a key component in the pathophysiology of FXS. Here, we directly test this idea in a FXS Drosophila model. ... We provide direct evidence that deficient inhibition impairs sensory computations and behavior in an in vivo model of FXS. Together with evidence of impaired inhibition in autism and Rett syndrome, these findings suggest a potentially general mechanism for intellectual disability."

Also appearing recently, this review:

Drozd M, Bardoni B, Capovilla M. Modeling Fragile X Syndrome in Drosophila. Front Mol Neurosci. 2018 Apr 16;11:124. doi: 10.3389/fnmol.2018.00124. PMID: 29713264; PMCID: PMC5911982.

Using flies to learn information related to Fragile X syndrome

Doll CA, Broadie K. Neuron class-specific requirements for Fragile X Mental Retardation Protein in critical period development of calcium signaling in learning and memory circuitry. Neurobiol Dis. 2016 May;89:76-87. PMID: 26851502; PMCID: PMC4785039.

From the abstract: "... Fragile X Mental Retardation Protein (FMRP), the gene product lost in Fragile X syndrome (FXS), acts as an activity sensor during critical period development, both as an RNA-binding translation regulator and channel-binding excitability regulator. Here, we employ a Drosophila FXS disease model to assay calcium signaling dynamics with a targeted transgenic GCaMP reporter during critical period development of the mushroom body (MB) learning/memory circuit. We find FMRP regulates depolarization-induced calcium signaling in a neuron-specific manner within this circuit, suppressing activity-dependent calcium transients in excitatory cholinergic MB input projection neurons and enhancing calcium signals in inhibitory GABAergic MB output neurons. ... These results show FMRP shapes neuron class-specific calcium signaling in excitatory vs. inhibitory neurons in developing learning/memory circuitry, and that FMRP mediates activity-dependent regulation of calcium signaling specifically during the early-use critical period."

Doll CA, Broadie K. Activity-dependent FMRP requirements in development of the neural circuitry of learning and memory. Development. 2015 Apr 1;142(7):1346-56. PMID: 25804740; PMCID: PMC4378248.

From the abstract: "... Here, we use optogenetic tools in the Drosophila FXS disease model to test activity-dependent dendritogenesis in two extrinsic neurons of the mushroom body (MB) learning and memory brain center: (1) the input projection neuron (PN) innervating Kenyon cells (KCs) in the MB calyx microglomeruli and (2) the output MVP2 neuron innervated by KCs in the MB peduncle. Both input and output neuron classes exhibit distinctive activity-dependent critical period dendritic remodeling. MVP2 arbors expand in Drosophila mutants null for fragile X mental retardation 1 (dfmr1), as well as following channelrhodopsin-driven depolarization during critical period development, but are reduced by halorhodopsin-driven hyperpolarization. ... results show that dfmr1 acts in a neuron type-specific activity-dependent manner for sculpting dendritic arbors during early-use, critical period development of learning and memory circuitry in the Drosophila brain."

Evidence from mammalian cells and Drosophila model contributes to a study related to Down syndrome and Fragile X syndrome

Sterne GR, Kim JH, Ye B. Dysregulated Dscam levels act through Abelson tyrosine kinase to enlarge presynaptic arbors. Elife. 2015 May 19;4. PMID: 25988807; PMCID: PMC4434255.

The abstract: "Increased expression of Down Syndrome Cell Adhesion Molecule (Dscam) is implicated in the pathogenesis of brain disorders such as Down syndrome (DS) and fragile X syndrome (FXS). Here, we show that the cellular defects caused by dysregulated Dscam levels can be ameliorated by genetic and pharmacological inhibition of Abelson kinase (Abl) both in Dscam-overexpressing neurons and in a Drosophila model of fragile X syndrome. This study offers Abl as a potential therapeutic target for treating brain disorders associated with dysregulated Dscam expression."

Review of Drosophila a model for neuropsychopharmacology-related research

Narayanan AS, Rothenfluh A. I Believe I Can Fly!: Use of Drosophila as a Model Organism in Neuropsychopharmacology Research. Neuropsychopharmacology. 2015 Oct 30. PMID: 26576740.

From the abstract:  "... Here, we outline why we study an invertebrate organism in the context of neuropsychiatric disorders, and we discuss how we can gain insight from studies in Drosophila. ... Highlighting some translational examples, we underline the fact that their brains works more like ours than one would have anticipated."

Their Fig. 1 provides a nice graphical summary of translational approaches.

Clinical trial to follow study of GABAAergic system in Fragile X syndrome using mice and flies

Braat S, Kooy RF. Insights into GABAAergic system deficits in fragile X syndrome lead to clinical trials. Neuropharmacology. 2015 Jan;88:48-54. PMID: 25016041.

From the abstract: "An increasing number of studies implicate the GABAAergic system in the pathophysiology of the fragile X syndrome, a frequent cause of intellectual disability and autism. Animal models have proven invaluable ... Aberrations compatible with those described in the mouse model were detected in dfmr1 deficient Drosophila melanogaster, a validated fly model for the fragile X syndrome. Treatment with drugs that ameliorate the GABAAergic deficiency in both animal models have demonstrated that the GABAA receptor is a promising target for the treatment of fragile X patients. Based on these preclinical studies, clinical trials in patients have been initiated."

Review looks at contributions of Drosophila research related to Fragile X syndrome and discusses discordance

Weisz ED, Monyak RE, Jongens TA. Deciphering Discord: How Drosophila research has enhanced our understanding of the importance of FMRP in different spatial and temporal contexts. Exp Neurol. 2015 May 27. pii: S0014-4886(15)30001-7. PMID: 26026973.

From the abstract: "Fragile X Syndrome (FXS) is the most common heritable form of intellectual impairment as well as the leading monogenetic cause of autism. ... Research efforts in Drosophila melanogaster have revealed key insights into the mechanistic underpinnings of FXS. While much remains unknown, it is increasingly apparent that FXS involves a myriad of spatial and temporally specific alterations in cellular function. Consequently, the literature is filled with numerous discordant findings. Researchers and clinicians alike must be cognizant of this dissonance, as it will likely be important for the design of preclinical studies to assess the efficacy of therapeutic strategies to improve the lives of FXS patients."

Review looks at fly and mouse models of Fragile X syndrome

Santos AR, Kanellopoulos AK, Bagni C. Learning and behavioral deficits associated with the absence of the fragile X mental retardation protein: what a fly and mouse model can teach us. Learn Mem. 2014 Sep 16;21(10):543-55. PMID: 25227249; PMCID: PMC4175497.

From the abstract: "... Here, we review to which extent these biological models are affected by the absence of FMRP, pointing out the similarities with the observed human dysfunction. Additionally, we discuss several potential treatments under study in animal models that are able to partially revert some of the FXS abnormalities. "

Drosophila used to help characterize a newly identified mutation in FMRP associated with Fragile X syndrome

Okray Z, de Esch CE, Van Esch H, Devriendt K, Claeys A, Yan J, Verbeeck J, Froyen G, Willemsen R, de Vrij FM, Hassan BA. A novel fragile X syndrome mutation reveals a conserved role for the carboxy-terminus in FMRP localization and function. EMBO Mol Med. 2015 Feb 17. pii: e201404576. PMID: 25693964.

From the abstract: "... In vivo analyses in Drosophila demonstrate that a patient-mimetic mutation alters the localization and function of Dfmrp in neurons, leading to neomorphic neuronal phenotypes."

Review--insights from fly and mouse models of Fragile X

Santos AR, Kanellopoulos AK, Bagni C. Learning and behavioral deficits associated with the absence of the fragile X mental retardation protein: what a fly and mouse model can teach us. Learn Mem. 2014 Sep 16;21(10):543-555. PMID: 25227249.

Impaired GABAergic circuit structure and function reported for fly model of Fragile-X syndrome

Gatto CL, Pereira D, Broadie K. GABAergic circuit dysfunction in the Drosophila Fragile X syndrome model. Neurobiol Dis. 2014 May;65:142-59. PMID: 24423648; PMCID: PMC3988906.

Catching up on neurodegenerative disease-related reports--Parkinson's disease, Fragile X syndrome, ALS

Parkinson's disease

Chuang CL, Lu YN, Wang HC, Chang HY. Genetic dissection reveals that Akt is the critical kinase downstream of LRRK2 to phosphorylate and inhibit FOXO1, and promotes neuron survival. Hum Mol Genet. 2014 Jun 10. PMID: 24916379. [fly and rat studies]

Hwang RD, Wiemerslage L, LaBreck CJ, Khan M, Kannan K, Wang X, Zhu X, Lee D, Fridell YW. The neuroprotective effect of human uncoupling protein 2 (hUCP2) requires cAMP-dependent protein kinase in a toxin model of Parkinson's disease. Neurobiol Dis. 2014 Sep;69:180-91. PMID: 24965893.

Ivatt RM, Whitworth AJ. SREBF1 links lipogenesis to mitophagy and sporadic Parkinson disease. Autophagy. 2014 Jun 27;10(8). Review. PMID: 24991824.

Bingol B, Tea JS, Phu L, Reichelt M, Bakalarski CE, Song Q, Foreman O, Kirkpatrick DS, Sheng M. The mitochondrial deubiquitinase USP30 opposes parkin-mediated mitophagy. Nature. 2014 Jun 19;510(7505):370-5. PMID: 24896179.

Fragile X syndrome

He F, Krans A, Freibaum BD, Taylor JP, Todd PK. TDP-43 suppresses CGG repeat-induced neurotoxicity through interactions with HnRNP A2/B1. Hum Mol Genet. 2014 May 8. PMID: 24920338. [From the abstract: "... suggest a convergence of pathogenic cascades between repeat expansion disorders and RNA-binding proteins implicated in neurodegenerative disease."]

Friedman SH, Dani N, Rushton E, Broadie K. Fragile X mental retardation protein regulates trans-synaptic signaling in Drosophila. Dis Model Mech. 2013 Nov;6(6):1400-13. PMID: 24046358; PMCID: PMC3820263.

ALS

Miskiewicz K, Jose LE, Yeshaw WM, Valadas JS, Swerts J, Munck S, Feiguin F, Dermaut B, Verstreken P. HDAC6 Is a Bruchpilot Deacetylase that Facilitates Neurotransmitter Release. Cell Rep. 2014 Jul 10;8(1):94-102. PMID: 24981865.

Fly study reports uncovering a nuclear role for FMRP.

Zhang W, Cheng Y, Li Y, Chen Z, Jin P, Chen D. A feed-forward mechanism involving Drosophila fragile X mental retardation protein triggers a replication stress-induced DNA damage response. Hum Mol Genet. 2014 May 15. pii: ddu241. PMID: 24833720.

From the abstract: "Fragile X syndrome, a common form of inherited mental retardation, is caused by loss of the fragile X mental retardation protein (FMRP). ... Here we show that Drosophila dFMR1 in nucleus is required for replication stress-induced H2Av phosphorylation in the DNA damage response (DDR). ..."

Fragile X model and GABAergic dysfunction

Gatto CL, Pereira D, Broadie K. GABAergic circuit dysfunction in the Drosophila Fragile X syndrome model. Neurobiol Dis. 2014 Jan 12. pii: S0969-9961(14)00015-1. PMID: 24423648.