DNA REPAIR PROTEINS TARGETED TO THE MITOCHONDRIAL MATRIX

Project: Research project

Project Details

Description

Metabolism of nutrients in mitochondria is the major source of energy for eukaryotic cells. Consequently, mitochondrial dysfunction can impair cellular and organismal homeostasis. The effects of mitochondrial dysfunction are exemplified by heritable genetic diseases and perhaps, normal aging. Mitochondria require the expression and translocation of nuclear proteins, as well as proteins encoded by mitochondrial DNA. Therefore, mutations that lead to dysfunction can occur in nuclear or mitochondrial DNA. The rate of mutation in primate mitochondrial DNA has been shown to be 5 to 10 times higher than in nuclear DNA. There are several factors that may contribute to the increased mutation rate detected in mitochondrial DNA including: 1) mitochondria utilize 90% of the cells 02 and generate many reactive oxygen species that can lead to mutations, 2) mitochondrial DNA is not associated with histones, and 3) mitochondrial DNA repair mechanisms are more limited in scope than nuclear DNA repair pathways. The primary objective of research described in this proposal is to determine if DNA repair in mitochondria can be increased by targeting DNA repair proteins to the mitochondrial matrix. This is the first step in positioning our laboratory to directly test the hypothesis that accumulation of mitochondrial mutations contribute to the aging process. The specific aims of the project are: 1) to generate and characterize stably transfected mammalian neuronal cell lines containing coding sequences for the luciferase reporter gene with or without mitochondrial matrix presequences, 2) to generate and characterize stably transfected mammalian neuronal cell lines containing DNA repair genes with or without mitochondrial matrix presequences, and 3) to generate and characterize transgenic mice containing transgenes that code for neuron-specific expression of DNA repair genes with or without mitochondrial matrix presequences. The proposed studies will determine whether mitochondrial matrix presequences can be fused to direct translocation of a non-mitochondrial protein to the mitochondrial matrix in mammalian cells. If successful, the next step will be to demonstrate that DNA repair proteins can be targeted to the mitochondrial matrix and result in elevated levels of repair for the specific lesions they recognize. Finally, transgenic mice will be generated using transgenes that contain mitochondrial matrix presequences to demonstrate that functional DNA repair proteins can be targeted to the matrix and effect increased DNA repair. Completion of this study will effectively position our laboratory to test the hypothesis that mitochondrial mutations contribute to aging.
StatusFinished
Effective start/end date7/20/956/30/01

Funding

  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health: $178,715.00
  • National Institutes of Health
  • National Institutes of Health

ASJC

  • Medicine(all)

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