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Drosophila models of motor neuron diseases

Research Center
Research Area

Supervisor

Dott. Andrea Daga
PhD, Research Unit coordinator
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Team

Dott.ssa Vantaggiato Chiara
Biologist, PhD
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Dott. Aldo Montagna
Biotechnologist, Research Fellow, PhD
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Dott.ssa Tatiana Trevisan
Biotechnologist, Research Fellow, PhD
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Dott.ssa Panzeri Elena
Biotechnologist, Research Fellow, PhD
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Description

Our laboratory uses Drosophila melanogaster as a model for human motor neuron disease. An approach toward disease insight is to model disease mechanisms, establish disease gene function and identify disease-modifying pathways in less complex organisms. The fruit fly Drosophila melanogaster has produced crucial advances into many neurological and neurodegenerative diseases, providing both an understanding of the biological pathways impaired in disease and clues for therapeutic approaches in mammalian systems. A combination of several important factors makes Drosophila a uniquely powerful animal model for neuroscience research. Flies have a short life cycle and are inexpensively to maintain. Gene manipulation in Drosophila is fast and relatively simple. For the study of the nervous system, the fly also offers unique advantages. In fact, the fly has a complex, yet les complicated than humans, central nervous system containing both neurons and glia, its brain is protected by a blood-brain barrier, and shares striking organizational, functional, developmental and molecular similarities with the vertebrate brain. Finally, the smaller number of genes in flies compared to the human genome implies that many genes that in humans are part of gene families composed of paralogues with redundant and/or overlapping functions are represented by a single member allowing much easier interpretation of loss-of-function studies.

We have  used Drosophila to model SPG3 and SPG4 hereditary spastic paraplegias caused by mutation of the atlastin and spastin encoding genes, respectively. We found that loss of spastin in Drosophila causes an aberrantly stabilized microtubule cytoskeleton in neurons and defects in synaptic growth and neurotransmission. We generated a spastin-linked HSP Drosophila model and showed that neural knockdown of Dspastin and, conversely, neural overexpression of Dspastin containing a conserved pathogenic mutation recapitulate some phenotypic aspects of the human disease, including adult onset, locomotor impairment, and neurodegeneration. At the subcellular level, neuronal expression of both spastin RNA interference and mutant spastin cause an excessive stabilization of microtubules in the neuromuscular junction synapse, however, administration of the microtubule targeting drug vinblastine significantly attenuates these phenotypes in vivo. Thus, loss of spastin function elicits HSP-like phenotypes in Drosophila providing novel insights into the molecular mechanism of spastin mutations, and raising the possibility that therapy with Vinca alkaloids may be efficacious in spastin-associated HSP.

Using Drosophila we were able to uncover the function of atlastin, the causative protein of SPG3a, one of the most common form of hereditary spastic paraplegia. Our studies demonstrated that Atlastin is the GTPase required for the homotypic fusion of ER membranes and suggested that pathological mutations in human atlastin-1 are likely to perturb membrane fusion. Thus, the ensuing loss of ER integrity is the likely defect underlying progressive axonal degeneration in atlastin-1-linked hereditary spastic paraplegia.

We are currently investigating the role of reticulon proteins in the ER and how this function may be linked to reticulon-dependent hereditary spastic paraplegia. Furthermore, we are now developing Drosophila models for SPG12 and SPG15 exploiting the most recent in vivo CRISPR/Cas9 mediated gene editing technology.

Recently, we entered the arena of mitochondria research focusing on mitochondria dynamics and the diseases caused by mutation of mitochondria dynamics genes. We have studied how perturbation of the fusion/fission balance affects mitochondria distribution in Drosophila axons. We have generated an in vivo system in which mitochondria fragmentation is not accompanied by encumbering bioenergetic dysfunctions. Mitochondrial fragmentation is observed in a variety of pathological conditions and our results indicate that in diseases characterized by altered mitochondrial morphology and altered axonal distribution, these defects may not be the primary cause of pathology since in our Drosophila model restoration of morphology and distribution without restoration of respiratory capacity does not permit survival of the fly.

Selected Publications

  • Trotta, N., Orso, G., Rossetto M.G., Daga, A., Broadie, K. (2004). The Hereditary Spastic Paraplegia Gene, spastin, regulates microtubule stability to modulate synaptic structure and function. Curr. Biol. 14(13):1135-47.
  • Orso, G., Martinuzzi, A., Rossetto, MG., Sartori, E., Feany, M., Daga, A. (2005). Disease-related phenotypes in a Drosophila model of Hereditary Spastic Paraplegia are ameliorated by treatment with the microtubule destabilizing agent vinblastine. J. Clin. Invest., 115(11):3026-3034.
  • Crippa F., Panzeri C., Martinuzzi A., Airoldi A., Redaelli F., Tonelli A., Baschirotto C., Vazza G., Mostacciuolo M.L., Daga A., Orso G., Profice P., Trabacca A., D’Angelo M.G., Comi G.P., Galbiati S., Lamperti C., Bonato S., Pandolfo M., Meola G., Musumeci O., Toscano A., Trevisan C.P., Bresolin N., Bassi MT. (2005). Eight novel mutations in SPG4 in a large sample of patients with Hereditary Spastic Paraplegia. Arch Neurol. 2006 May;63(5):750-5.
  • Panzeri C, De Palma C, Martinuzzi A, Daga A, De Polo G, Bresolin N, Miller CC, Tudor EL, Clementi E, Bassi MT. The first ALS2 missense mutation associated with JPLS reveals new aspects of alsin biological function. Brain. 2006 Jul;129(Pt 7):1710-9.
  • Tauber E, Zordan M, Sandrelli F, Pegoraro M, Osterwalder N, Breda C, Daga A, Selmin A, Monger K, Benna C, Rosato E, Kyriacou CP, Costa R. Natural selection favors a newly derived timeless allele in Drosophila melanogaster. Science. 2007 Jun 29;316(5833):1895-8
  • Lissandron V, Rossetto MG, Erbguth K, Fiala A, Daga A, Zaccolo M. Transgenic fruit-flies expressing a FRET-based sensor for in vivo imaging of cAMP dynamics. Cell Signal. 2007 Nov;19(11):2296-303.
  • Crimella C, Arnoldi A, Crippa F, Mostacciuolo ML, Boaretto F, Sironi M, D'Angelo MG, Manzoni S, Piccinini L, Turconi AC, Toscano A, Musumeci O, Benedetti S, Fazio R, Bresolin N, Daga A, Martinuzzi A, Bassi MT. Point mutations and a large intragenic deletion in SPG11 in complicated spastic paraplegia without thin corpus callosum. J Med Genet. 2009 May;46(5):345-51.
  • Orso G, Pendin D, Liu S, Tosetto J, Moss TJ, Faust JE, Micaroni M, Egorova A, Martinuzzi A, McNew JA, Daga A. Homotypic fusion of endoplasmic reticulum membranes requires the Dynamin-like GTPase atlastin. Nature. 2009 Aug 20;460(7258):978-83.
  • Moss TJ, Daga A, McNew JA. Fusing a lasting relationship between ER tubules. Trends Cell Biol. 2011 Jul;21(7):416-23.
  • Pendin D, McNew JA, Daga A. Balancing ER dynamics: shaping, bending, severing, and mending membranes. Curr Opin Cell Biol. 2011 Aug;23(4):435-42.
  • Moss TJ, Andreazza C, Verma A, Daga A, McNew JA. Membrane fusion by the GTPase atlastin requires a conserved C-terminal cytoplasmic tail and dimerization through the middle domain. Proc Natl Acad Sci U S A. 2011 Jul 5;108(27):11133-8.
  • Rossetto MG, Zanarella E, Orso G, Kumar V, Delgado-Escueta AV, Daga A. The juvenile myoclonic epilepsy gene EFHC1 modulates neurite architecture and basal synaptic activity. Hum Mol Genet. 2011 Nov 1;20(21):4248-57.
  • Pendin D, Tosetto J, Moss TJ, Andreazza C, Moro S, McNew JA, Daga A. GTP-dependent packing of a 3-helix bundle is required for atlastin-mediated fusion. Proc Natl Acad Sci U S A. 2011 Sep 27;108(39):16283-8.
  • Grisar T, Lakaye B, de Nijs L, LoTurco J, Daga A, Delgado-Escueta AV. Myoclonin1/EFHC1 in cell division, neuroblast migration, synapse/dendrite formation in juvenile myoclonic epilepsy. In: Noebels JL, Avoli M, Rogawski MA, Olsen RW, Delgado-Escueta AV, editors. Jasper's Basic Mechanisms of the Epilepsies. 4th edition. Bethesda (MD): National Center for Biotechnology Information (US); 2012.
  • Debattisti V, Pendin D, Ziviani E, Daga A, Scorrano L. Reduction of endoplasmic reticulum stress attenuates the defects caused by Drosophila mitofusin depletion. J Cell Biol. 2014 Feb 3;204(3):303-12.
  • Papadopoulos C, Orso G, Mancuso G, Herholz M, Gumeni S, Jüngst C, Tadepalle N, Tzschichholz A, Schauss A, Höning S, Trifunovic A, Daga A, Rugarli EI. Spastin binds to lipid droplets and affects lipid metabolism in D. melanogaster and C. elegans. PLoS Genet. 2015 Apr 13;11(4):e1005149. doi: 10.1371/journal.pgen.1005149.
  • Summerville JB, Faust JF, Fan E, Pendin D, Daga A, Formella J, Stern M, McNew JA. The effects of ER morphology on synaptic structure and function in Drosophila melanogaster. J Cell Sci. 2016 Apr 15;129(8):1635-48. doi: 10.1242/jcs.184929.
  • Bailey JN, Patterson C, de Nijs L, Durón RM, Nguyen VH, Tanaka M, Medina MT, Jara-Prado A, Martínez-Juárez IE, Ochoa A, Molina Y, Suzuki T, Alonso ME, Wight JE, Lin YC, Guilhoto L, Targas Yacubian EM, Machado-Salas J, Daga A, Yamakawa K, Grisar TM, Lakaye B, Delgado-Escueta AV. EFHC1 variants in juvenile myoclonic epilepsy: reanalysis according to NHGRI and ACMG guidelines for assigning disease causality. Genet Med. 2017 Jul 28. doi: 10.1038/gim.2016.86.
  • Trevisan T, Pendin D, Montagna A, Bova S, Ghelli AM, Daga A. Manipulation of Mitochondria Dynamics Reveals Separate Roles for Form and Function in Mitochondria Distribution. Cell Rep. 2018 May 8;23(6):1742-1753. doi: 10.1016/j.celrep.2018.04.017.

Partnerships

  • Vadim Frolov, Unidad de Biofísica (UPV/EHU, CSIC), Leioa, Spain
  • Antonio Delgado-Escueta, Department of Neurology, Ronald Reagan UCLA Medical Center, University of California Los Angeles, USA
  • Masahiro Shirakawa, Department of Molecular Engineering, Kyoto University Katsura, Japan
  • James McNew, Department of Biosciences, Rice University, Houston, USA

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