title: Expression of the Opa1 mitochondrial protein in retinal ganglion cells: its downregulation causes aggregation of the mitochondrial network.
abstract: OBJECTIVE
  Mutations in the mitochondrial dynamin-related GTPase OPA1 cause autosomal dominant optic atrophy (ADOA), but the pathophysiology of this disease is unknown. As a first step in functional studies, this study was conducted to evaluate the expression of Opa1 in whole retina and in isolated retinal ganglion cells (RGCs) and to test the effects of Opa1 downregulation in cultured RGCs.
  
  
  METHODS
  Opa1 mRNA isoforms from total retina and from RGCs freshly isolated by immunopanning were determined by RT-PCR. Protein expression was examined by immunohistochemistry and Western blot with antibodies against Opa1 and cytochrome c, and the mitochondrial network was visualized with a mitochondrial marker. Short interfering (si)RNA targeting OPA1 mRNAs were transfected to cultured RGCs and mitochondrial network phenotypes were followed for 15 days, in comparison with those of cerebellar granule cells (CGCs).
  
  
  RESULTS
  Opa1 expression did not predominate in rat postnatal RGCs as found by immunohistochemistry and Western blot analysis. The pattern of mRNA isoforms was similar in whole retina and RGCs. After a few days in culture, isolated RGCs showed fine mitochondrial punctiform structures in the soma and neurites that colocalized with cytochrome c and Opa1. Opa1 knockdown in RGCs induced mitochondrial network aggregation at a higher rate than in CGCs.
  
  
  CONCLUSIONS
  Results suggest that the level of expression and the mRNA isoforms do not underlie the vulnerability of RGCs to OPA1 mutations. However, aggregation of the mitochondrial network induced by the downregulation of Opa1 appears more frequent in RGCs than in control CGCs.
authors.affiliation: Institut National de la Santé et de la Recherche Médicale (INSERM) U583, Physiopathologie et Thérapie des Déficits Sensoriels et Moteurs, Institut des Neurosciences de Montpellier, Hôpital Saint Eloi, France.
authors.forename: Satomi
authors.initials: S
authors.lastname: Kamei
authors.forename: Murielle
authors.initials: M
authors.lastname: Chen-Kuo-Chang
authors.forename: Chantal
authors.initials: C
authors.lastname: Cazevieille
authors.forename: Guy
authors.initials: G
authors.lastname: Lenaers
authors.forename: Aurélien
authors.initials: A
authors.lastname: Olichon
authors.forename: Pascale
authors.initials: P
authors.lastname: Bélenguer
authors.forename: Gautier
authors.initials: G
authors.lastname: Roussignol
authors.forename: Nicole
authors.initials: N
authors.lastname: Renard
authors.forename: Michel
authors.initials: M
authors.lastname: Eybalin
authors.forename: Adeline
authors.initials: A
authors.lastname: Michelin
authors.forename: Cécile
authors.initials: C
authors.lastname: Delettre
authors.forename: Philippe
authors.initials: P
authors.lastname: Brabet
authors.forename: Christian P
authors.initials: CP
authors.lastname: Hamel
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D015536:Down-Regulation; D020558:GTP Phosphohydrolases / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D020868:Gene Silencing / Q000502:physiology; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism* / Q000473:pathology; D028361:Mitochondrial Diseases / Q000378:metabolism* / Q000473:pathology; D024101:Mitochondrial Proteins / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D012164:Retinal Diseases / Q000378:metabolism* / Q000473:pathology; D012165:Retinal Ganglion Cells / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
chemical_list: D007527:Isoenzymes; D024101:Mitochondrial Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D020558:GTP Phosphohydrolases; C498811:Opa1 protein, rat