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The Other Genome
Since the discovery that Leber hereditary optic neuropathy (LHON) results from mutations in mitochondrial
DNA (mtDNA), considerable attention has been focused
on this alternative genome and on development of the
scientific tools needed to study this remarkable genetic
pathway (1, 2 ). In this issue, Chen et al. (3 ) describe the
application of temporal temperature gradient gel electrophoresis to the detection of mtDNA mutations and show
that this technique offers great promise in this application.
The study of mitochondrial gene mutations presents
investigators with new technical problems not inherent to
the study of nuclear gene mutations. This genome is
thought to be derived from an evolutionarily ancient
organism that parasitized primitive cells, conferring on
them enhanced oxidative capacity and the capability of
making profitable use of atmospheric oxygen, a fairly
toxic substance. The structure of the present day human
mitochondrial genome reflects its unusual origin. The
mitochondrial genome is a small (16.5 kb) circular DNA
encoding only 13 proteins, 2 rRNAs, and a set of tRNAs.
All proteins encoded by the mitochondrial genome are
components of the mitochondrial electron transport chain,
the energy-transducing, oxidative apparatus of the cell.
Unlike nuclear genes, which exist in pairs by virtue of
their location on paired chromosomes, mitochondrial
genes exist in numerous copies per cell, with each mitochondrion containing several copies of the genome and
each cell containing many mitochondria. In the case of a
nuclear gene, only a limited number of combinations of
mutated genes are possible: a situation in which both are
wild type, a situation in which one is mutated and one is
wild type, or a situation in which both are mutated. This
binary type of arithmetic gives rise to the well-known
patterns of Mendelian inheritance with recessive and
dominant traits. In contrast, mitochondrial gene mutations can exist in a continuous spectrum ranging from
none of the copies of a given mitochondrial gene being
mutated to all of the copies being mutated. The existence
of more than one genotype is termed heteroplasmy. The
degree of heteroplasmy correlates with phenotype, and
there appears to be a threshold of mutational burden
below which the cell retains enough mitochondrial capacity to function normally and no abnormality of phenotype
results. Further complicating the situation is the fact that
an individual’s mitochondrial genotype may change over
time. This occurs because mitochondria replicate (through
fission), and there is some evidence that defective mitochondria may replicate more quickly than healthy (mutation-free) mitochondria (4 ). Consequently, the mitochondrial genetics of disease may represent a kind of
Darwinian population genetics within the cell. Thus,
quick, accurate detection and quantification of mitochondrial gene mutations is critical. This need points up the
importance of the findings of Chen et al. (3 ) who present
a simple, rapid, and sensitive method for the detection of
heteroplasmic mutations.
Traditional arguments have held that hereditary disorders involving mitochondrial gene mutations should be
readily identifiable through analysis of pedigree of information because mtDNA is inherited exclusively maternally with no paternal contribution. A disorder derived
from mtDNA should be transmitted vertically through a
family in a situation resembling autosomal dominant
inheritance with both genders expressing the phenotype,
but there should be no instances of father-to-child transmission. Indeed, David C. Wallace (5, 6 ) recognized the
genetically unusual situation in LHON long before it was
understood that this disorder is mitochondrially inherited
(7 ). This type of pedigree analysis has led to the correct
identification of other mtDNA-derived disorders such as
myoclonus-epilepsy, ragged red fiber disease, and the
syndrome of myopathy, encephalopathy, lactic acidosis,
and stroke (8 ).
The emerging challenge in the field is the identification
of a role for mitochondrial genes in disorders that occur
sporadically and without a clear pattern of maternal
inheritance. A theoretical argument was advanced a decade ago that some disorders resulting from mtDNA
mutations might appear sporadically in a population and
that mitochondrial genetics might offer a general approach to the problem of sporadic disease (9 ). In support
of this argument, a specific mitochondrial electron transport chain defect (complex I, NADH:ubiquinone oxidoreductase) was identified in platelets of patients with
sporadic Parkinson disease (PD), suggesting that PD is in
fact a systemic disorder and not confined to a small
portion of the brain (9 ). This same defect has been
identified in PD brain and other tissues (10 ). This is not
unlike the situation with LHON in which there is a
widespread biochemical and genetic lesion but pathology
is typically confined to the optic nerve. Further importance is lent to the finding of complex I deficiency in PD
by the fact that complex I is the target enzyme of the
PD-causing neurotoxin, methylphenyltetrahydropyridine
(11 ).
The origin of the complex I defect in PD has been
investigated through cybrid technology (12 ). This technique involves creation of culturable human cell lines that
have been depleted of their own endogenous mtDNA by
prolonged culture in the presence of ethidium bromide,
which concentrates in mitochondria and binds to mtDNA.
This ultimately leads to formation of a cell line that lacks
mtDNA and which is designated r0. These cells can be
repopulated with mtDNA of the investigator’s choosing
by fusion of these cells with enucleated cytoplasts or with
platelets (which lack a nucleus but contain mitochondria
and mtDNA). The resulting cytoplasmic hybrid (cybrid)
can be passed in culture. Cell lines repopulated with PD
mtDNA can be compared to lines derived from the same
r0 cells but repopulated with mtDNA from controls. Any
differences likely arise from mtDNA. PD cybrid lines
express complex I deficiency, indicating that this bio-
Clinical Chemistry 45, No. 8, 1999
Parker: Editorial
chemical lesion is probably genetic in origin and arises
from mtDNA (12 ). In addition to a loss of complex I
activity, these cells also demonstrate other pathogenic
features typical of PD, including increased production of
oxygen radicals, up-regulation of oxygen radical defense
enzymes, tendency toward apoptotic cell death, and perturbed calcium metabolism (13 ). These results suggest
that mtDNA is pathogenically important in PD and that
studies are needed of abnormalities of mtDNA sequences
in PD.
Inheritance of mtDNA mutation is clearly one route to
human disease. Another approach has been suggested by
Wallace (14 ), who has stressed the role of acquired
mtDNA mutation. The intramitochondrial location of
mtDNA and its lack of a protective histone coating make
mtDNA susceptible to oxidative damage and mutation. It
is possible that damage acquired over a lifetime could
ultimately lead to loss of adequate mitochondrial function
and cell death. This is a particularly appealing mechanism
in the case of age-associated neurodegenerative disorders
such as PD. Careful studies of mtDNA sequence may
distinguish between these two possibilities or may show
that both are important.
There is now abundant evidence that defective mitochondria can play a critical role in the initiation of
apoptosis through release of cytochrome c and perhaps
other factors as well (15, 16 ). This link between mitochondrial dysfunction and programmed cell death again emphasizes the need to understand the origin of diseaseassociated mitochondrial dysfunction. If it is indeed
primary, pharmacologic interventions aimed either at
repairing mitochondrial dysfunction or at breaking the
signaling link between mitochondria and apoptosis
should be useful.
Mitochondrial genes, acting either alone or in concert
with nuclear genes and/or environmental factors, may be
of great importance in the pathogenesis of relatively
common disorders. Thorough investigations of sequence
changes in mtDNA will be critical in proving or rejecting
this hypothesis and may bring about a new appreciation
of this emerging field of human genetic disease.
1. Wallace DC, Singh G, Lott MT, Hodge JA, Schurr TG, Lezza AMS, et al.
Mitochondrial DNA mutation associated with Leber’s hereditary optic neuropathy. Science 1988;242:1427–30.
2. Parker WD, Oley CA, Parks JK. Deficient NADH:coenzyme Q oxidoreductase
in Leber’s hereditary optic neuropathy. N Engl J Med 1989;320:1331–3.
3. Chen T-J, Boles RG, Wong L-JC. Detection of mitochondrial DNA mutations by
temporal temperature gradient gel electrophoresis. Clin Chem 1999;45:
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for the agency responsible for its unusual pattern of inheritance. Brain
6. Wallace DC. Leber’s optic atrophy: a possible example of vertical transmission of a slow virus in man. Aust Ann Med 1970;3:259 – 62.
7. Howell N, McCullough D. An example of Leber hereditary optic neuropathy
not involving a mutation in the mitochondrial ND4 gene. Am J Hum Genet
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8. Shoffner JM, Lott MT, Lezza AMS, Seibel P, Ballinger SW, Wallace DC.
Myoclonic epilepsy and ragged-red fiber disease (MERRF) is associated with
a mitochondrial DNA tRNA{1lys} mutation. Cell 1990;61:931–7.
9. Parker WD, Boyson SJ, Parks JK. Abnormalities of the electron transport
chain in idiopathic Parkinson’s disease. Ann Neurol 1989;26:719 –23.
10. Schapira AHV, Cooper JM, Dexter D, Clark JB, Jenner P, Marsden CD.
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11. Vyas I, Heikkila RE, Nicklas WJ. Studies on the neurotoxicity of 1-methyl-4phenyl-1,2,5,6-tetrahydropyridine: inhibition of NAD-linked substrate oxidation by its metabolite, 1-methyl-4-pyridinium. J Neurochem 1986; 46:
12. Swerdlow RH, Parks JK, Miller SW, Tuttle JB, Trimmer PA, Sheehan JP, et al.
The origin and functional consequences of the complex I defect in Parkinson’s disease. Ann Neurol 1996;40:663–71.
13. Parker WD, Swerdlow RH. Mitochondrial dysfunction in idiopathic Parkinson’s disease. Am J Hum Genet 1998;66:758 – 62.
14. Wallace DC. Mitochondrial genetics: a paradigm for aging and degenerative
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15. Liu X, Kim CN, Yang J, Jemmerson R, Wang X. Induction of apoptotic
program in cell-free extracts: requirement for dATP and cytochrome c. Cell
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W. Davis Parker, Jr.
Departments of Neurology and Pediatrics
and the Center for the
Study of Neurodegenerative Disorders
University of Virginia School of Medicine
Charlottesville, VA 22901
E-mail [email protected]