Molecular Remedy of Mitochondrial Defects

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DESCRIPTION (provided by applicant): The overall goal of this grant application is to identify potential area of treatment and/or remedies for the diseases that result from dysfunctional oxidative phosphorylation system. It has been reported that many human diseases are associated with defects in the mitochondrial proton-translocating NADH-quinone oxidoreductase (complex I). Defects in complex I, which render it dysfunctional, result in the following three problems: (1) impaired ability of the respiratory chain to oxidize NADH; (2) impaired ability of this enzyme to pump protons; (3) production of reactive oxygen species (ROS). Of these 3 problems, impairment of proton pumping at any one of the three proton translocation sites does not appear to present a severe health hazard when compared to the inability of mitochondria to oxidize NADH and/or damage caused by ROS. Baker's yeast mitochondria lack complex I but contain instead a single-subunit NADH dehydrogenase (Ndi1). We have constructed a recombinant adeno-associated virus carrying the NDI1 gene (rAAV-NDI1) and transduced mammalian cell lines. In all cells we have tested so far, the NDI1 gene can be expressed. The expressed Ndi1 was correctly imported into mitochondria guided by its own leading sequence and acted as a member of the respiratory chain in host cells. In addition, functional expression of the Ndi1 was observed in growth-arrested human cells as well as in differentiated rodent dopaminergic nerve cells. These results indicate that the NDI1 gene provides a potentially useful tool for gene therapy of complex I associated diseases. The studies planned for the grant period are as follows. (1) Functional expression of the yeast NDI1 gene in various tissues of rats and mice using rAAV-NDI1. (2) Construction of animal models for human diseases caused by complex I defects. (3) Protection by the Ndi1 enzyme against tissue degradation caused by complex I deficiencies.