Eric A. Schon, PhD

Research Summary
Mitochondrial genetics and the molecular basis of human mitochondrial disease.

Mitochondria are unique among the constituents of the eukaryotic cell in that they are semi-autonomous organelles that contain their own genetic machinery. As such, they operate under the dual genetic controls of nuclear DNA (nDNA) and mitochondrial DNA (mtDNA).

Mitochondrial genetics differs markedly from mendelian genetics, because first, mitochondria are inherited exclusively from the mother, and second, there are hundreds or thousands of mitochondria (and mtDNAs) per cell. In addition, organellar division and mtDNA replication are stochastic processes unrelated to the cell cycle, and mtDNA gene organization, DNA replication, RNA transcription, and protein translation all have a prokaryotic "look" about them. This latter feature is no surprise, given that mitochondria were once bacteria that were taken up by the proto-eukaryotic cell early in evolution. Biochemically, the most relevant aspect of mitochondrial function is the production of oxidative energy via the respiratory chain and oxidative phosphorylation.

Mitochondrial diseases have turned out to be equally unusual. There are maternally-inherited, mendelian-inherited, sporadic, and even environmentally induced mitochondrial disorders, most of which are either severely debilitating or fatal. We are studying the molecular basis of a number of these diseases, most of which are heteroplasmic (i.e. both mutant and wild-type mtDNAs coexist in varying proportions in the same patient), using a novel tissue culture system called "rho cells." These cells contain mitochondria, but the mitochondria are completely devoid of mtDNAs, and are thus respiratorily deficient. We have transferred heteroplasmic mitochondria from patients to these rho cells, thereby creating cytoplasmic hybrids, or "cybrids," that contain known proportions of mutant or wild-type mtDNAs in clonal cell lines that have no contaminating mtDNA background. We are using cybrid technology not only to study genotype phenotype relationships, but also to ask basic questions about mitochondrial biogenesis (e.g. mitochondrial fusion and exchange of genetic information).

We have begun a project on mitochondrial gene therapy, in order to ameliorate the effects of a known pathogenic (and fatal) mtDNA mutation that affects ATP synthesis. In addition, we are trying to make the first "transmitochondrial" mouse model of a human mtDNA disease.

Selected Publications

1. Schon EA. Tales from the crypt. J Clin Invest. 2003 Nov;112(9):1312-6. Full Text

2. Schon EA, DiMauro S. Medicinal and genetic approaches to the treatment of mitochondrial disease. Curr Med Chem. 2003 Dec;10(23):2523-33.

3. DiMauro S, Schon EA. Mitochondrial respiratory-chain diseases. N Engl J Med. 2003 Jun 26;348(26):2656-68. Review. Full Text

4. Gajewski CD, Yang L, Schon EA, Manfredi G. (2003) New insights into the bioenergetics of mitochondrial disorders using intracellular ATP reporters. Mol. Biol. Cell, 14, 3628-3635.

5. Ojaimi J, Pan J, Santra S, Snell WJ, Schon EA. An algal nucleus-encoded subunit of mitochondrial ATP synthase rescues a defect in the analogous human mitochondrial-encoded subunit. Mol Biol Cell. 2002 Nov;13(11):3836-44. Full Text

6. Manfredi G, Fu J, Ojaimi J, Sadlock JE, Kwong JQ, Guy J, Schon EA. (2002) Rescue of a deficiency in ATP synthesis by transfer of MTATP6, a mitochondrial DNA--encoded gene, to the nucleus. Nat Genet, 25, 394-399. Full Text

1. Cytochrome oxidase assembly genes in human disease
The four aims of this proposal are: (1) we will clarify the unexpectedly complex patterns of transcription and protein expression of the two known human SCO genes (hSCO1 and hSCO2); (2) we will study hSCO2 deficiency in two cellular models in which SCO function is compromised, namely, in patient cells harboring hSC02 mutations and in yeast cells harboring SCO1/SCO2 null mutations; (3) we will create mouse models of hSCO2 deficiency (both knock-out and knock-in mice); and (4) we will search for mutations in hSCO1 and hSCO2, and in other COX-assembly genes as well, in a large series of candidate patient tissues available to us and our colleagues here at Columbia.
National Institute of Neurological Disorders and Stroke
4/2000-3/2004

2. Nuclear gene involvement in cytochrome oxidase deficiency
Cytochrome c oxidase (COX), or complex IV of the mitochondrial respiratory chain, is a copper- and heme-containing metalloprotein composed of 13 subunits, 3 encoded by mitochondrial DNA (mtDNA) and 10 by nuclear DNA (nDNA). A number of COX-deficiency disorders are associated with point mutations in mtDNA-encoded COX subunits, but very little is known regarding the molecular basis of mendelian-inherited COX deficiency disorders, which display widely varying phenotypes, including both generalized and tissue-specific clinical presentations. In particular, no mutation in any of the 10 nDNA-encoded COX subunits has yet been found. Mutations have been found in four COX assembly genes - COX10, SCO1, SCO2, and SURF1. We believe that other such genes exist, and propose to identify them by screening our large collection of fibroblasts from patients with COX deficiency. We propose to first, screen for "obvious" candidate genes among the known COX structural and assembly genes. Among those cell lines which survive this first screen, we will use functional complementation by a combination of rodent-human monochromosomal hybrids and microcell-mediated chromosomal transfer to identify complementation groups and candidate chromosomal loci. The culprit genes at these loci will be identified by a combination of microsatelle and deletion mapping, coupled with a judicious sequencing of candidate genes in the region. Any new gene thus identified will be characterized as to its role in COX structure, function, and tissue-specific expression. Uncovering the molecular basis of COX deficiency will be useful in terms of pointing the way to diagnosis and treatment of these generally fatal and untreatable disorders. Moreover, these errors will be extremely useful in understanding fundamental biological phenomena, such as COX holoenzyme assembly, COX function, mitochondrial importation, and energy utilization and production. There are no naturally occurring COX mutants in higher eukaryotes other than those causing the diseases outlined above. Thus, elucidating the molecular basis of the COX deficiencies will afford us new and useful insights from both a clinical and scientific standpoint.
National Institute of Neurological Disorders and Stroke
4/2002-3/2007

3. Therapeutic approaches in models of mitochondrial disease
We will devote our main efforts to improving the efficiency of mitochondrial importation, and to developing stably transfected cell lines expression our constructs. If successful, allotopic expression of the recoded wild-type gene, which has been engineered to be targeted and imported into mitochondria, should ameliorate the effects of the ATPase 6 mutation in mutant cells. Second, we will begin to devise a pharmacological approach to treat these disorders, based on our recent finding that treatment of heteroplasmic cells containing the MILS/NARP mutation with a complex V-specific inhibitor (oligomycin) in medium containing galactose results in a rapid and stable shift in heteroplasmy in favor of wild-type mtDNAs, with a concomitant improvement in mitochondrial function. We will now ask if the same shift can occur in heteroplasmic postmitotic MILS/NARP (myotubes), and also ask if this type of strategy can be generalized to mitochondrial deficiencies in other respiratory complexes. Finally, we will begin to assess the pharmacological ramifications of using oligomycin (and related compounds) in cells and in mice, as a prelude to considering a treatment protocol in MILS patients.
National Institute of Child Health and Human Development
Fiscal Year 2004

Keywords
cytochrome oxidase, genetic disorder, mitochondrial DNA, mitochondrial disease /disorder, molecular assembly /self assembly, congenital disorder, copper, disease /disorder etiology, gene therapy, genetic transcription, membrane protein, myocardium disorder, neuromuscular disorder, pathologic process, phenotype, transport protein
human genetic material tag, human tissue, in situ hybridization, laboratory mouse, northern blotting, tissue /cell culture, transgenic animal, yeast, DNA replication origin, gene rearrangement, mitochondria, molecular pathology, disease /disorder model, gene duplication, model design /development, neurogenetics, clinical research, polymerase chain reaction