How mtDNA Mutations Cause Mitochondrial Disease
Abstract
The work proposed here directly relates to the topic area of Mitochondrial Disease. Mitochondria are the “powerhouse” of the cell. The food we eat and the oxygen we breathe are combined inside the mitochondria to produce energy that sustains life. Mitochondria are unique among all the organelles because they have their own DNA, the mitochondrial DNA. Because all genes in the mitochondrial DNA are essential, mutations in these genes can cause mitochondrial diseases. These diseases can manifest at any point during the life of an individual. However, developmental disorders in children and degenerative diseases in older individuals are the most common. Likewise, mutations in the mitochondrial DNA can affect any organ in the body but the brain, muscles, and sperm, which have large energetic needs, are particularly vulnerable to mitochondrial DNA mutations. Finally, lifestyle, diet, and environmental insults such as exposure to toxins, trauma, and stress can modulate the susceptibility to mitochondrial DNA disorders. Here, I propose to use a basic sciences approach in a simple animal model system to gain insights into mitochondrial DNA diseases. A central challenge in studying mitochondrial DNA diseases arises from there being hundreds to thousands of copies of the mitochondrial DNA in each cell. This is in contrast to the presence of only two copies of the nuclear chromosomal DNA. While all copies of the mitochondrial DNA are typically normal, some can become mutated. When the number of mutant copies of the mitochondrial DNA becomes predominant and there are insufficient copies of normal mitochondrial DNA to support cellular function, disease state ensues. In most cases, mutant mitochondrial DNA rises to pathogenically high levels in only some tissues and cell types. Hence, a fundamental challenge is to understand how mutant mitochondrial DNA comes to predominate in certain tissues during development and during normal course of aging. Solving this challenge will allow us to better predict the risks of developing mitochondrial DNA diseases. It is also crucial for the success of mitochondrial replacement therapy, which aims to eliminate inheritance of mutant copies of mitochondrial DNA by children. In this procedure, mutant copies of mitochondrial DNA in eggs from one female are swapped with normal copies from eggs of another female. However, it is nearly impossible to obtain a complete swap and, inevitably, embryos generated via this procedure still harbor a few copies of the mutant mitochondrial DNA. Recent studies suggest that these few copies can proliferate in the developing embryo, eventually rising to high enough levels to cause disease. Clearly, we need a better understanding of how mutant mitochondrial DNA copies proliferate during development and aging. Traditionally, studies of mitochondrial DNA mutant levels in tissues have been performed in mice. However, mechanistic studies in mice are limited due to time and cost constraints. Instead, we propose to use cutting-edge technologies in a simple nematode C. elegans, which is easy and cheap to cultivate in large numbers in the laboratory. Given its small size, it is challenging to dissect out individual tissues from C. elegans. Instead, we will dissolve animals into their constituent cells, and then use a cell-sorting method to isolate cell types of choice. Not only does this method overcome the limitations of performing dissections, but it also allows us to isolate almost any cell type of choice with exquisite specificity. We will then measure mutant mitochondrial DNA levels in the isolated cells with a highly sensitive methodology called droplet digital PCR. With these technological innovations, we will test the hypothesis that mitochondrial DNA mutant levels change in a stereotypical manner across cell types during development and aging. We will also test the role of mitochondrial physiology to determine the cellular and molecular mechani
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Oct 29, 2018
- Source ID
- W81XWH1810181
Entities
People
- Maulik R. Patel
Organizations
- United States Army
- Vanderbilt University