Recombinant adeno-associated virus vector-based gene transfer for defects in oxidative metabolism

R. Owen IV, A. P. Lewin, A. Peel, J. Wang, J. Guy, W. W. Hauswirth, P. W. Stacpoole, T. R. Flotte

Research output: Contribution to journalArticlepeer-review

33 Scopus citations


Defects in oxidative metabolism may be caused by mutations either in nuclear genes or in mitochondrial DNA (mtDNA). We tested the hypothesis that recombinant adeno-associated virus (rAAV) could be used to complement mtDNA mutations. AAV vector constructs were designed to express the reporter gene encoding green fluorescent protein (GFP), fused to a targeting presequence that directed GFP to be translocated into mitochondria. These vectors mediated expression of mitochondrial-localized GFP, as indicated by fluorescence microscopy and electron microscopy, in respiring human embryonic kidney 293 cells and nonrespiring mtDNA-deficient (ρ0) cells. However, when sequences encoding hydrophobic segments of proteins normally encoded by mtDNA were inserted between the presequence and GFP, mitochondrial import failed to occur. In similar experiments, a fusion was created between pyruvate dehydrogenase (PDH) E1α subunit, a nuclear-encoded mitochondrial gene with its own targeting presequence, and GFP. With this construct, expression of GFP was observed in mitochondria in vitro and in vivo. We conclude that the hydrophobicity of mtDNA-encoded proteins limits their ability to be transported from the cytoplasm. However, rAAV-based gene therapy may hold promise for gene therapy of PDH deficiency, the most common biochemically proven cause of congenital lactic acidosis.

Original languageEnglish (US)
Pages (from-to)2067-2078
Number of pages12
JournalHuman gene therapy
Issue number15
StatePublished - Oct 10 2000
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics


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