Manipulating the Metazoan Mitochondrial Genome with Targeted Restriction Enzymes
Hong Xu,
Steven Z. DeLuca,
Patrick H. O'Farrell*
High copy number and random segregation confound genetic analysis
of the mitochondrial genome. We developed an efficient selection
for heritable mitochondrial genome (mtDNA) mutations in
Drosophila,
thereby enhancing a metazoan model for study of mitochondrial
genetics and mutations causing human mitochondrial disease.
Targeting a restriction enzyme to mitochondria in the germline
compromised fertility, but escaper progeny carried homoplasmic
mtDNA mutations lacking the cleavage site. Among mutations eliminating
a site in the cytochrome c oxidase gene,
mt:CoIA302T was healthy,
mt:CoIR301L was male sterile but otherwise healthy, and
mt:CoIR301S exhibited a wide range of defects, including growth retardation,
neurodegeneration, muscular atrophy, male sterility, and reduced
life span. Thus, germline expression of mitochondrial restriction
enzymes creates a powerful selection and has allowed direct
isolation of mitochondrial mutants in a metazoan.
* To whom correspondence should be addressed. E-mail: ofarrell{at}cgl.ucsf.edu
