title: Severe impairment of complex I-driven adenosine triphosphate synthesis in leber hereditary optic neuropathy cybrids.
abstract: BACKGROUND
  Leber hereditary optic neuropathy (LHON) is a maternally inherited form of central vision loss associated with mitochondrial DNA point mutations that affect the ND subunits of complex I.
  
  
  OBJECTIVE
  To elucidate the bioenergetic consequences of complex I dysfunction in LHON.
  
  
  METHODS
  The biochemical phenotypes of LHON mutations have been investigated using the transmitochondrial cytoplasmic hybrid (cybrid) cell model derived from the osteocarcoma parental cell line 143B.TK-.
  
  
  METHODS
  Research laboratories at neuroscience and biochemistry departments at the University of Bologna, Scientific Institute "E. Medea," and University of College Medical School.
  
  
  METHODS
  Fibroblast cell lines were obtained from patients affected with LHON, as defined by the presence of 1 pathogenic mutation, and from healthy volunteers as controls to construct cybrid cell lines.
  
  
  METHODS
  Complex I (glutamate-malate)- and complex II (succinate)-dependent adenosine triphosphate (ATP) synthesis, their respective respiratory rates, and total cellular ATP content were investigated using digitonin permeabilized cybrid cells. Multiple cybrid cell lines were constructed, introducing into osteosarcoma-derived rho(0) cells either wild-type or LHON mutant mitochondria carrying each of the 3 common mutations at positions 11778/ND4, 3460/ND1, and 14484/ND6.
  
  
  RESULTS
  All 3 LHON mutations impaired ATP synthesis and the respiratory control ratio driven by complex I substrates. In contrast, succinate-driven ATP synthesis, respiration rates, and respiratory control ratios were not affected. However, the defective ATP synthesis with complex I substrates did not result in reduced ATP cellular content, indicating a compensatory mechanism.
  
  
  CONCLUSIONS
  The LHON pathogenic mutations profoundly impair complex I-dependent synthesis of ATP, providing a common biochemical feature that may play a major role in LHON pathogenesis. Stratification of the results by mutation suggests that the 11778/ND4 mutation may induce an uncoupling of cybrid respiration, whereas the other 2 mutations impair the oxygen consumption rate.
authors.affiliation: Dipartimento di Biochimica, University of Bologna, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Baracca
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Solaini
authors.forename: Gianluca
authors.initials: G
authors.lastname: Sgarbi
authors.forename: Giorgio
authors.initials: G
authors.lastname: Lenaz
authors.forename: Agostino
authors.initials: A
authors.lastname: Baruzzi
authors.forename: Anthony H V
authors.initials: AH
authors.lastname: Schapira
authors.forename: Andrea
authors.initials: A
authors.lastname: Martinuzzi
authors.forename: Valerio
authors.initials: V
authors.lastname: Carelli
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D002478:Cells, Cultured; D019343:Citric Acid / Q000378:metabolism; D004272:DNA, Mitochondrial / Q000235:genetics*; D042967:Electron Transport Complex I / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology; D005971:Glutamates / Q000378:metabolism; D006801:Humans; D006822:Hybrid Cells; D009154:Mutation; D029242:Optic Atrophy, Hereditary, Leber / Q000201:enzymology / Q000235:genetics*; D010101:Oxygen Consumption / Q000235:genetics; D019802:Succinic Acid / Q000378:metabolism
chemical_list: D004272:DNA, Mitochondrial; D005971:Glutamates; D019343:Citric Acid; D000255:Adenosine Triphosphate; D019802:Succinic Acid; D042967:Electron Transport Complex I