title: Zebrafish Tg(hb9:MTS-Kaede): a new in vivo tool for studying the axonal movement of mitochondria.
abstract: OBJECTIVE
  Deregulation of axonal transport in neurons is emerging as the major cause of many neurodegenerative diseases in human, such as Charcot-Marie-Tooth (CMT) neuropathy. However, little is known about how mitochondria move in vivo and whether cell culture systems truly represent what happens in living animals. Here we describe the generation of a new zebrafish transgenic line that specifically allows to study mitochondrial dynamics in motor neurons and its application to analyse mitochondrial movement in zebrafish models expressing CMT2A causing mutations.
  
  
  METHODS
  The Tol2 transposon system was used to generate a transgenic zebrafish line expressing the photoconvertible fluorescent protein Kaede in mitochondria of motor neurons. Mitochondrial shape and movement were monitored by time-lapse confocal live imaging and measured by kymograph analysis. The effects of two well-known CMT causing mutations, L76P and R94Q substitutions in MFN2, were then investigated with the same methods.
  
  
  RESULTS
  We generated the transgenic zebrafish Tg(hb9:MTS-Kaede) line with genetically labelled mitochondria in motor neurons. Kaede protein was correctly and stably targeted to mitochondrial matrix while retaining its photoconvertibility, thus qualifying this model for in vivo studies. Expression of the L76P and R94Q mutations reduced mitochondrial movement in axons and altered mitochondrial distribution in distinct ways.
  
  
  CONCLUSIONS
  These findings confirm previously published data obtained in cell cultures and strengthen the hypothesis of different mechanism of action of the two MFN2 mutations. Considering the number of neurodegenerative diseases associated to mitochondrial dynamics, the Tg(hb9:MTS-Kaede) zebrafish line is a promising model to study in vivo alterations of mitochondrial transport underlying human diseases.
authors.affiliation: Dipartimento di Biologia, Università degli Studi di Padova, Via Ugo Bassi 58/B, 35121 Padova (PD), Italy. Electronic address: giorgia.bergamin@kcl.ac.uk.
authors.forename: Giorgia
authors.initials: G
authors.lastname: Bergamin
authors.affiliation: Dipartimento di Biologia, Università degli Studi di Padova, Via Ugo Bassi 58/B, 35121 Padova (PD), Italy.
authors.forename: Domenico
authors.initials: D
authors.lastname: Cieri
authors.affiliation: Dipartimento di Biologia, Università degli Studi di Padova, Via Ugo Bassi 58/B, 35121 Padova (PD), Italy. Electronic address: giovanni.vazza@unipd.it.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Vazza
authors.affiliation: Dipartimento di Biologia, Università degli Studi di Padova, Via Ugo Bassi 58/B, 35121 Padova (PD), Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Argenton
authors.affiliation: Dipartimento di Biologia, Università degli Studi di Padova, Via Ugo Bassi 58/B, 35121 Padova (PD), Italy.
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Mostacciuolo
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001370:Axonal Transport / Q000502:physiology*; D002607:Charcot-Marie-Tooth Disease / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000502:physiology; D008928:Mitochondria / Q000502:physiology*; D063154:Mitochondrial Dynamics; D024101:Mitochondrial Proteins / Q000502:physiology; D009154:Mutation; D015027:Zebrafish
chemical_list: D024101:Mitochondrial Proteins; D020558:GTP Phosphohydrolases; C470018:MFN2 protein, human
keywords: Charcot-Marie-Tooth disease; Mitochondrial dynamics; Motor neuron; Zebrafish