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Update on Nuclear Mitochondrial Genesand Neurologic DisordersAmy Goldstein, MD, Poonam Bhatia, MD, and Jodie M. Vento, MGC, CGC

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The majority of primary mitochondrial disorders are due to nuclear gene mutations, notaberrations within the mitochondrial genome. The nervous system is frequently involveddue to its high-energy demands. Many nonspecific neurologic symptoms may be present inmitochondrial disease; however, there are well-recognized red flags that should alert theclinician to the possibility of mitochondrial disease. There is an ever increasing number ofnuclear gene mutations discovered that play a role in primary mitochondrial disease and itsneurologic symptomatology. Neurologists need to be aware of the wide neurologicpresentation, the red-flag symptoms, and the nuclear gene mutations involved in thepathophysiology of mitochondrial disease to diagnose and manage this patient population.Semin Pediatr Neurol 19:181-193 C 2012 Elsevier Inc. All rights reserved.

IntroductionPrimary mitochondrial diseases are a group of clinically and

genetically heterogeneous disorders that result in decreased

energy production of the respiratory chain (RC) in the formof adenosine triphosphate (ATP). The etiology of primary

mitochondrial disease is complex and can include abnorm-

alities in RC function due to structural or assemblyproteins, translation of the mitochondrial deoxyribonucleic

acid (mtDNA), and maintenance of the mtDNA. Primary

mitochondrial disease may be classified clinically bysymptoms or as a defined syndrome or by biochemical or

molecular, or both defects. The same clinical phenotype

may be caused by a variety of genetic etiologies, and thesame mutation can lead to a wide phenotypic spectrum,

even within the same family. For example, a child may have

epilepsy, developmental delay, and hypotonia and undergoa skin biopsy for evaluation. The RC testing may reveal an

isolated complex I deficiency, which may be due to one outof more than 30 known genetic mutations causing complex

I deficiency. In younger children, the RC testing may be

completely normal in tissue, despite harboring a mutationknown to cause complex I deficiency. Current technology

allows for massively parallel sequencing of many nuclear

mitochondrial genes with a genetic mutation discovery inabout 20%-25% of selected patients.1 With the rapid

ront matter & 2012 Elsevier Inc. All rights reserved.

016/j.spen.2012.09.005

Child Neurology, Department of Pediatrics, Chil-

Pittsburgh of UPMC, Pittsburgh, PA.

ests to Amy Goldstein, MD, Children’s Hospital of

Penn Avenue, Pittsburgh, PA 15224. E-mail:

p.edu

evolution of molecular genetic testing, the ability to detect

a pathologic mutation through next-generation sequencingwould dramatically improve our ability to arrive at a

genetic diagnosis. Therefore, classification is complicated

and a combined clinical, biochemical, and geneticapproach is needed. We have organized this paper based

on the neurologic phenotypes as this would be most

helpful to the practicing clinician. However, the betterknown biochemical and molecular defects with heteroge-

neous phenotypes would be highlighted as well, providing

insight into the genetic classification of these disorders.

Neurologic SymptomsIn primary mitochondrial disease, the organ systems

requiring the most ATP to function properly present withevidence of energy failure. Therefore, the energy-

demanding nervous system is frequently affected, especially

in childhood. Brain and muscle, along with vision andhearing, are typically involved, bringing that child to the

attention of the child neurologist.

Mitochondrial disease should be included in the differ-ential diagnosis for almost any neurologic symptom. The

suspicion should be increased when 3 or more organ

systems are involved without a known etiology. Mitochon-drial disease has a wide spectrum of presentations with any

symptom, any organ, any age, and any inheritance pattern.2

However, common presenting symptoms are stroke-likeepisodes, headache, seizures, psychomotor regression,

ataxia, and encephalopathy.3

181

Table 1 Red-Flag Neurologic Symptoms

Neurologic Symptomor Sign

Red-flag Signs

Stroke Nonvascular distribution; MRI/ADC map shows a mixture of hyperintensity andhypointensity

Basal ganglia lesions Bilateral symmetric (characteristic of Leigh syndrome); also with brainstem lesionsEncephalopathy-hepatopathy Precipitated by valproic acid exposure; associated hepatic failureEpilepsy Epilepsia partialis continua (EPC), myoclonus, and status epilepticusCognitive decline Regression with illnessAtaxia Associated with epilepsy or other systemic symptoms; neuroimaging may show cerebellar

atrophy, white matter lesions, and basal ganglia lesionOcular signs Optic nerve atrophy, ophthalmoplegia, and ptosis; retinopathySensorineural hearing loss At early age, accompanied by other systemic symptoms

ADC ¼ apparent diffusion coefficient.

A. Goldstein, P. Bhatia, and J.M. Vento182

Symptoms most suggestive of mitochondrial disease are

typically progressive or recurrent, but may have partial

recovery after a decline. Disease symptoms may be pre-cipitated by a metabolic stressor, such as infection, fasting,

surgery, or any medication that may be toxic to the

mitochondria. The neurologic symptom may be the pre-senting feature of the disease, but they are often a part of a

multisystem clinical picture, and therefore, the clinician

must be aware of other systemic symptoms.Although any neurologic symptom could be the pre-

senting sign of mitochondrial disease, there have been well-

established neurologic red-flag symptoms that should alert theclinician to a possible diagnosis of mitochondrial disease

(Table 1).4,5 Other common neurologic symptoms or

findings include myopathy, neuropathy, and leukoencepha-lopathy. Exercise intolerance that is out of proportion to

weakness is concerning for mitochondrial disease. The

systemic symptoms to be aware of include cardiac conduc-tion disorders such as Wolff-Parkinson-White syndrome,

cardiomyopathy, hepatopathy, renal involvement including

Fanconi syndrome orglomerulopathy, diabetes mellitus orpancreatic exocrine failure, bone marrow failure, and

gastrointestinal dysmotility or chronic intestinal pseudo-

obstruction.

Stroke-Like Lesions in NonvascularDistributionMetabolic strokes are the result of a significant, sudden

compromise in energy production. Unlike vascular strokes,they do not follow a vascular territory and usually occur in

younger individuals. Although these lesions are most

typical of mtDNA point mutations causing mitochondrialencephalomyopathy with lactic acidosis and stroke-like

episodes, nuclear gene mutations, such as polymerase

gamma (POLG), can cause secondary mtDNA changes,such as mtDNA depletion and multiple deletions, leading

to a metabolic stroke-like presentation.6 The neuroimaging

apparent diffusion coefficient map can show a mixture ofhypointensity and hyperintensity, suggestive of both cyto-

toxic and vasogenic edema.7

Case example: A 16-year-old female presents with acutevision loss and constant left leg twitching. The result of

electroencephalogram (EEG) is consistent with epilepsia

partialis continua (EPC) with epileptiform discharges from

the right centro-parieto-temporal and right occipito-temporal region. Brain magnetic resonance imaging (MRI)

reveals T2 prolongation in right occipital, pulvinar, pre-

frontal gyrus, cerebellum, and splenium of corpus callo-sum. Result for testing of mtDNA point mutations is

negative. Further nuclear diagnostic testing reveals com-

pound heterozygosity of 2 known pathogenic mutationsin POLG.

Leigh Syndrome (Bilateral Symmetric BasalGanglia Lesions)Leigh syndrome is named after Denis Leigh (pronouncedLee), a British neuropathologist, who described subacute

necrotizing encephalomyelopathy in a 7-month-old in 1951.8

Historically, it has been a finding seen at autopsy, but isnow commonly identified on neuroimaging of the brain

with calcifications on computed tomography or MRI with

typical focal, bilateral, and symmetric T2 hyper- T1hypointense lesions with a characteristic involvement of

periaqueductal gray, pons, midbrain, brainstem, cerebel-

lum, basal ganglia, and rarely white matter involvement.9,10

Symptoms and MRI findings can fluctuate based on

metabolic stress with typical worsening seen during acute

illness, fasting, or surgery.Symptoms of Leigh syndrome include encephalopathy

with global developmental delay, seizures, basal ganglia or

brainstem dysfunction with prominent movement disorder,dysphagia and feeding difficulties, somnolence, nystagmus,

respiratory involvement including apnea, and ataxia. The

movement disorder can include choreathetosis, dyskinesia,and dystonia and can be difficult to treat. Feeding

difficulties and increased calorie burn from constant move-

ments often lead to failure to thrive. Many of these childrenare at a high risk for aspiration and should be monitored

for swallowing difficulties with a consideration on feeding

tube placement. Seizures are present in 40% of patientswith Leigh syndrome. Other neurologic symptoms include

neuropathy, myopathy, and optic atrophy. Systemic symp-

toms may include diabetes, short stature, cardiomyopathy,anemia, renal failure, vomiting, and diarrhea.11 Some

Genetics for the neurologist 183

patients may be diagnosed with cerebral palsy or hypoxic-

ischemic encephalopathy and have a relatively static course,

although usually Leigh syndrome is a progressive disease.12

Leigh syndrome is typically seen more in the pediatric

population, but reports of adults with Leigh syndrome are

seen throughout the literature.13,14 Adults may presentwith different features than those seen in the pediatric

population, such as more prominent cranial nerve dysfunc-

tion and cerebellar and long-tract signs instead of devel-opmental delay, failure to thrive, elevated lactate level,

imaging of the abnormalities in basal ganglia, and COX

deficiency.15

Leigh syndrome is caused by both nuclear- and mito-

chondrial-encoded mutations, including pyruvate dehydro-

genase complex deficiency, RC complex deficiency (inparticular, SURF1, an assembly factor for complex IV),

and PDSS2, causing primary coenzyme Q10 (CoQ10)

deficiency.16,17

Case example: a 5-year-old with long-standing cerebral

palsy and seizure disorder presents to the emergency room

with difficulty in breathing. He acutely develops a feverwith temperature of 1041F and becomes encephalopathic.

Biochemical testing shows lactic acidosis. MRI scan of the

brain reveals an increased signal on MRI in bilateral basalganglia. Two weeks later, he is no longer able to track

visually and lost verbal expression. Diagnostic testing

reveals 2 mutations in the autosomal recessive gene,COX10, which is a known cause of complex IV deficiency.

Encephalopathy With HepatopathyEncephalopathy is a nonspecific clinical symptom but

combined with a finding of hepatopathy should increasethe suspicion of mitochondrial disease as well as other

inborn errors of metabolism. The most common causes of

encephalopathy with hepatopathy are the mtDNA deple-tion syndromes (MDDS).18 Alpers-Huttenlocher syndrome,

also known as Alpers or AHS, is one of the MDDS

phenotypes and represents a prototypical nuclear genemutation in POLG, causing a breadth of clinical pheno-

types. The clinical spectrum of POLG is discussed sepa-

rately later. Other MDDS gene mutations that includeencephalopathy-hepatopathy symptoms are DGUOK,

MPV17, TWINKLE, and SUCLG1.19 These hepatocerebral

phenotypes are characterized by early-onset neurologicabnormalities and progressive liver failure. Biochemical

testing reveals hypoglycemia and lactic acidosis, as well

as multiple RC complex deficiencies and mtDNA depletion(reduction in mtDNA copy number).20

Case example: a 3-year-old girl presents acutely with

episodes of altered mental status, abdominal pain withbloating during feeds. EEG during her altered mental status

episode is normal. She has a history of failure to thrive and

refractory gastroesophageal reflux. Her result of neuroima-ging is normal. Her serum lactates are on borderline level,

and her urine organic acids show dicarboxylic acids. Her

gastroenterologist orders a liver biopsy for elevated transa-minases. Liver biopsy demonstrates complex I þ III þ IV

deficiency. Two heterozygous mutations in the autosomal

recessive gene, DGUOK, are determined to be the etiology

of her symptoms.

Cognitive Decline (Mitochondrial Dementia)Regression, or loss of previously achieved milestones, is a

red-flag sign of childhood neurodegenerative disease.

Regression can be slowly progressive or acute whentriggered by mitochondrial stressors. Epilepsy and stroke-

like episodes can also trigger a cognitive decline.

Cognitive impairment may be obvious on clinical exam-ination or may be so focal and well compensated that

neuropsychological testing is necessary to determine any

deficits. The range of cognitive deficits seen in mitochon-drial disease is wide and can have more of a psychiatric

presentation with frank psychosis, acute confusional states,

and behavioral changes or mimic other neurodegenerativedementias seen in Alzheimer or Parkinson disease.21

Childhood POLG and Leigh syndrome have altered mental

status as a common symptom.Dementia often starts with specific focal cognitive deficits

and not a global decline. Dementia may be seen in isolation

at presentation but as with other mitochondrial diseases, ismore likely to be seen with other symptoms. Other central

nervous system symptoms that often accompany dementia

include epilepsy, stroke-like episodes, weakness, spasticity,movement disorders, ataxia, weakness, and migraine head-

ache. Result of neuroimaging may be normal or may show

global atrophy, basal ganglia calcifications, or areas ofincreased T2-weighted signal in the basal ganglia or white

matter.21

The nuclear mitochondrial diseases that have cognitivedecline as a common symptom include mitochondrial neu-

rogastrointestinal encephalomyopathy (MNGIE); POLG-

related disorders (various phenotypes); progressive externalophthalmoplegia (PEO; due to POLG, TWINKLE, TK2, or

ANT1); leukoencephalopathy with brainstem and spinal cord

involvement and lactate level elevation; Charcot-Marie-Toothdisease type 2; Leigh syndrome (due to SURF1); diabetes

insipidus, diabetes mellitus, optic atrophy, and deafness also

called Wolfram syndrome; and Mohr-Tranebjaerg (due toDDP1) also called deafness and dystonia syndrome.22

Cognitive impairment with decline is a frequent feature

of childhood and adult mitochondrial disease, most likelydue to Leigh syndrome in the younger age group. The

cognitive impairment progresses with duration of disease

and is seen as a symptom more frequently when the resultof neuroimaging shows abnormal. Treatment is supportive,

but CoQ10 deficiency should be investigated to offer

specific treatment.23

Case example: a 17-year-old male presents with a history

of myopathy. Previous muscle biopsy revealed COX-

negative fibers; biochemical testing confirms complex IVdeficiency. He begins to have difficulty in reading compre-

hension, attention, and memory. MRI and EEG reports are

normal. Neuropsychological testing is performed to deter-mine specific deficits and to recommend school

A. Goldstein, P. Bhatia, and J.M. Vento184

accommodations. Sequencing of the mtDNA genome in

muscle is normal, and further nuclear genetic analysis does

not reveal pathogenic mutations in known nuclear-encodedcomplex IV genes.

EpilepsyThe brain has an extremely high-energy requirement and is

therefore frequently involved in children who have mito-chondrial disease. Epilepsy is the main childhood mani-

festation of mitochondrial encephalopathy.24 Epilepsy may

be the presenting feature of mitochondrial disease, but asseen with other neurologic symptoms, it is often part of a

multisystem clinical picture. Other features, such as cog-

nitive decline, may evolve over time but be interpreted asan epileptic encephalopathy. Common findings in epilepsy

that should raise the suspicion of mitochondrial disease

include EPC, myoclonus, and status epilepticus, especiallyif the seizures are explosive in onset.24 Associated systemic

symptoms commonly seen with mitochondrial epilepsy

include sensorineural hearing loss, retinopathy, cardiomyo-pathy or arrhythmia, diabetes mellitus, hepatopathy, and

renal tubulopathy.24

Seizures are quite common in mitochondrial disease,with a reported incidence of 35%-60% in patients with

biochemical or molecularly confirmed mitochondrial dis-

ease.24 In a retrospective study, patients with idiopathicepilepsy admitted to the pediatric epilepsy monitoring unit

were reviewed for previous testing and 28% had biochem-

ical abnormalities suggestive of mitochondrial dysfunction,and in those with multifocal epileptiform discharges,

biochemical abnormalities were seen in 75%.25

Patients with epilepsy and mitochondrial disease havemixed seizure types. One review of patients with confirmed

RC defects revealed a spectrum of epilepsy phenotypes

ranging from Ohtahara syndrome to Landau-Kleffner syn-drome, with seizure types ranging from generalized to

partial. Seizure onset was in young children, and the

neuroimaging of the majority showed abnormality.26

Another review of patients with seizures and mitochon-

drial disease demonstrated that seizures were preceded by

multisystemic symptoms, such as global delay, failure tothrive, or ataxia in the majority of patients, and most had

multiple seizure types. The epilepsy phenotypes ranged

from neonatal refractory status epilepticus with multiorganfailure to infantile spasms, myoclonic epilepsy, recurrent

status epilepticus, and EPC. For one-third of the patients,

serum and urine biochemical testing was completelynormal and diagnosis was made on liver biopsy RC testing.

These authors suggested that epilepsy was a poor prog-

nostic sign in this population, with a 45% mortality rate,and half of their patients dying within 9 months of the

onset of their epilepsy.27

Common mitochondrial etiologies of epilepsy includePOLG, other disorders of mtDNA maintenance (PEO1,

RRM2B, and SUCLA2), complex I deficiency, complex

deficiencies caused by disordered assembly (FOXRED1,BCS1L, SCO2, and TMEM70) or abnormal protein

structure (NDUFV1, NDUFS4, NDUFA1, SDHA, and

MTATP6), disorders of CoQ10 biosynthesis (PDSS2,

COQ2, COQ6, COQ9, and ADCK3), disorders of mito-chondrial translation (RARS2 and TFSM), or disordered

mitochondrial import of small molecules (SLC25A22).24

Myoclonic epilepsy in mitochondrial disease, in particu-lar caused by POLG mutations, can mimic a progressive

myoclonic epilepsy syndrome (such as neuronal ceroid-

lipofuscinosis) due to other associated symptoms, whichinclude cognitive decline, ataxia, motor incoordination,

and pigmentary retinopathy. POLG-related myoclonic epi-

lepsy has been shown to be more resistant to therapy withantiepileptic drugs compared with others, such as complex

I assembly factor gene mutations.24 Therefore, determining

the underlying etiology can have important implications fortreatment.

The treatment of mitochondrial epilepsy can be difficult

and require multiple antiepileptic drugs. The ketogenic dietmay be considered and appears to be helpful in terms of

reducing seizure frequency, but further studies are needed

to determine whether a subgroup of patients wouldrespond better than another. While the ketogenic has been

utilized in different epilepsy types and different RC

diseases, significant side effects, such as metabolic acidosisand persistent hypoglycemia, have been reported.28-31. In

particular, the ketogenic diet is specifically contraindicated

in pyruvate carboxylase deficiency.32 The use of valproicacid (VPA) has been very controversial. Avoidance of VPA-

induced hepatotoxicity and concern for Alpers syndrome

existed long before the POLG gene was discovered.Prospective testing for POLG of patients before the use of

VPA has been suggested by several groups.33,34

Case example: a 17-month-old with liver failure isreferred for neurologic examination. He had normal devel-

opment and neuroimaging. VPA was started after his

second seizure, when he could not tolerate his first antic-onvulsant. Within 3 months, he developed fulminant

hepatic failure. Molecular analysis of POLG revealed 2

heterozygous mutations in trans.

AtaxiaMitochondrial ataxias can present at any age and are

clinically characterized by a cerebellar syndrome of ataxia,

dysarthria, and nystagmus.35 Sensory ataxia from peripheralneuropathy or spinal cord involvement may also be present.

Cerebellar atrophy can be found as the neuroimaging

correlates with this symptom. Additional neuroimagingcharacteristics on MRI include involvement of the basal

ganglia and white matter or increased T2-weighted signal in

the cerebellar cortex.35 Imaging may be normal despite thepresence of cerebellar signs on examination. Other systemic

symptoms that accompany mitochondrial ataxia may include

encephalomyopathy, epilepsy, muscle weakness, regression,hearing loss, ophthalmoplegia, short stature, ataxia, optic

atrophy, increased muscle tone, and stroke-like episodes.35

Nuclear mitochondrial syndromes in which ataxia maybe present include Leigh syndrome; sensory ataxic

Genetics for the neurologist 185

neuropathy, dysarthria, ophthalmoplegia; spinocerebellar

ataxia and epilepsy (SCAE); Alpers-Huttenlocher syn-

drome; X-linked sideroblastic anemia with ataxia;infantile-onset spinocerebellar ataxia; mitochondrial reces-

sive ataxia syndrome (MIRAS); myoclonus epilepsy myo-

pathy sensory ataxia (MEMSA); and leukoencephalopathywith brainstem and spinal cord involvement with lactic

acidosis.36 Ataxia is less frequently seen in chronic PEO

(CPEO) (both recessive and dominant forms); MNGIE;diabetes insipidus, diabetes mellitus, optic atrophy, and

deafness; CoQ10 deficiency; autosomal dominant optic

atrophy and deafness, dilated cardiomyopathy with ataxia;and pyruvate dehydrogenase complex (PDC) deficiency.36

Friedreich ataxia is a secondary mitochondrial disease due

to mutations in frataxin leading to RC deficiencies. Clinicalfindings include cerebellar ataxia, spasticity, cardiomyopa-

thy, dysarthria, and diabetes.37

Case example: a 16-year-old boy presents with intermit-tent difficulty in walking. His examination reveals ataxia

and nystagmus, with mild dysarthria. His MRI report shows

cerebellar atrophy. The serum lactate level was elevated 5times greater than the upper limit of normal, with elevated

pyruvate. Skin fibroblasts reveal PDC deficiency.

Ocular SymptomsPatients with mitochondrial disease should be monitored ona regular basis by an ophthalmologist familiar with findings

associated with mitochondrial disease. The evaluation

should include an ophthalmologic examination, includingvisual acuity testing and a fundoscopic examination, to

screen for optic neuropathy and pigmentary retinopathy.38,39

Visual field testing can reveal abnormalities due tocerebral strokes or lesions by detecting field cuts and signs

of optic neuropathy. The external examination should

detect and monitor ptosis and ophthalmoplegia, whichcan be seen in CPEO due to POLG, POLG2, ANT1, or

TWINKLE mutations.40,41 Optic atrophy and retinal degen-

eration can be seen due to the metabolically active retinalpigment epithelium and retinal ganglion cell layer. Visual

evoked potentials may be abnormal in mitochondrial

disease where the axons of the retinal ganglion cells areinvolved, but the photoreceptors are spared (result of

electroretinography is normal). Electroretinography can

detect cone dysfunction, which has been reported in somenuclear mitochondrial diseases.42

Mitochondrial ptosis can have asymmetric onset and be

slowly progressive with little diurnal variation. When it isaccompanied by pigmentary retinopathy changes or PEO,

mitochondrial disease should be suspected over myasthenia

gravis or other conditions. Pigmentary retinopathy inmitochondrial disease has a perimacular distribution, with-

out drusen, and should not affect vision.42

Case example: a 15-year-old boy with a history ofgeneralized epilepsy and short stature presents with

migraine headaches and difficulty concentrating in school.

On neurologic examination, his eyelids appear to bedroopy. His parents concur that he has appeared more

sleepy than usual. He is seen by an ophthalmologist, and

ptosis is confirmed. Tissue biopsy reveals complex IV

deficiency. Mutations are identified in the autosomalrecessive gene, TWINKLE (C10orf2).

Sensorineural Hearing LossImpaired hearing is a common clinical finding in mito-chondrial disease, due to the high-energy demands of the

auditory apparatus. Seldom seen in isolation, the hearing

loss is usually seen in the context of a multisystem disorder,and in fact, the hearing loss may be missed due to other

clinical symptoms needing attention. Patients with mito-

chondrial disease should undergo testing for hearing losson a routine basis. The hearing loss is amenable to treat

with amplified devices, such as hearing aids and cochlear

implantation. Some of the nuclear mitochondrial genemutations that cause hearing loss include the MDDS

involving SUCLA2, SUCLG1, and TWINKLE.41

Case example: a 4-year-old boy presents with cerebralpalsy and developmental delay. Brain MRI is consistent

with Leigh syndrome, and skin biopsy showed complex IV

deficiency. A complex IV nuclear panel in blood revealedmutations in SURF1. With a diagnosis of mitochondrial

disease, he had brainstem auditory evoked responses and

was found to have bilateral hearing loss. He received acochlear implant, and his encephalopathy has improved.

Genetic Classification ofMitochondrial DiseasesThe clinical suspicion of mitochondrial disease based onthe above-mentioned red-flag neurologic symptoms opens

up a diagnostic odyssey for both the clinician and the

patient. Biochemical screening tests may include serumlactate, pyruvate, amino acids, acylcarnitine profile, carni-

tine battery, CoQ10 levels, and urine organic acids. Tissue

sampling may include skin biopsy for further biochemicalanalysis of RC defects or defects in pyruvate metabolism

(PDC and pyruvate carboxylase). Muscle or liver biopsy can

be analyzed for RC defects, mtDNA copy number to lookfor depletion syndromes, mtDNA sequencing to look for

mutations or deletions, and CoQ10 quantitation.

The confirmatory molecular diagnosis of mitochondrialdisease is challenging due to the presence of 2 separate but

interacting genomes: the mtDNA and the nuclear DNA

(nDNA). Both of these genomes must undergo DNAreplication and protein synthesis to generate the complex

(I-V) subunits and assembly factors involved in the RC. The

small, circular mtDNA contributes to only a fraction ofmitochondrial disease. Only 13 subunits of the RC are

mtDNA-encoded, whereas nDNA encodes the remaining

74 subunits of complexes I-V.43 In addition, nDNA alsoencodes CoQ10 and cytochrome c and important subunits

in the RC.44 Overall, nDNA encodes about 1500 proteins

targeted to the mitochondria.45 Therefore, most inheritedmitochondrial disease is nuclear encoded46, and as such,

Table 2 Nuclear Mitochondrial Genes and Associated Clinical Symptoms

InheritancePattern*

Stroke-like

Lesions

BilateralBasal

GangliaLesions

Encephalopathy Epilepsy CognitiveDecline

Ataxia OcularSigns

SensorineuralHearing Loss

Other NeurologicSigns

Other SystemicFindings

Complex I deficiencyNDUFA1 XL X X X X Hypotonia,

choreoathetosisNDUFAF2 AR X X XNDUFAF4 AR X X X CardiomyopathyNDUFS3 AR† X DystoniaNDUFS4 AR X Hypotonia Failure to thriveNDUFS6 AR Fatal neonatal

lactic acidosisNDUFS7 AR† X X DysarthriaNDUFS8 AR† X Poor feeding,

cardiomyopathyNDUFV1 AR X X X X Leukoencephalopathy

Complex II deficiencySDHA AR XSDHAF1 AR X Leukoencephalopathy

Complex III deficiencyBCSIL AR X X X X Bjornstad

syndrome‡,GRACILE

syndrome§.Hepatopathy, renal

disease

Complex IV deficiencyCOX10 AR X X X X X X Hypotonia, pyramidal

signsRenal tubulopathy,

cardiomyopathySCO1 AR X X Leukoencephalopathy,

neuropathyNeonatal

hepatopathySCO2 AR X Myopathy, neuropathy,

leukoencephalopathyFatal neonatal

cardiomyopathySURF1 AR X X X Failure to thriveCOX15 AR X X X Leukoencephalopathy,

hypotoniaFatal

cardiomyopathyCOX6B1 AR X Weakness, ataxia,

leukoencephalopathyFASTKD2 AR X X Hemiplegia, asym-

metric cerebral atrophy

A.Goldstein,P.Bhatia,andJ.M

.Vento186

Complex V deficiencyTMEM70 AR X X X X Hypotonia

Cardio-myopa-

thy,IUGR

Coenzyme Q 10 deficiencyPDSS1 AR X X Neuropathy Cardiac valvu-

lopathy, livedoreticularis

PDSS2 AR X X X X Myopathy NephropathyCoQ2 AR X X Myopathy NephropathyCoQ6 X X X NephropathyCoQ9 AR X Myopathy Nephropathy,

cardiomyopathyADCK3 AR X X X X

Mitochondrial DNA depletion syndromesPOLG AD/AR X X X X X X Myopathy, neuropathy,

leukoence phalopathyHepatopathy

RRM2B AD/AR X X X X Myopathy, neuropathy NephropathySUCLA2 AR X X X X Myopathy, dystonia Lactic acidosisSUCLG1 AR X X X Myopathy, dystonia Infantile lactic

acidosisSUCLG2 X X MyopathyC10orf2/

TwinkleAD/AR x X X X X Neuropathy, athetosis Hepatic

involvementDGUOK AR x X Neuropathy Hepatic involve-

ment (neonatalonset, severe)

MPV17 AR x Neuropathy Infantile hepatopa-thy, hypoglycemia

TK2 AR X X Congenital musculardystrophy

TYMP AR X X X Myopathy, neuropathy,leukoence phalopathy

Gastrointestinaldysmotility, anor-

exia, MNGIE

OthersTAZ XL X Weakness Cardiomyopathy

(Barth syndrome)k,neutropenia

OPA1 AD X X X Myopathy, neuropathyOPA3 AD/AR X X Chorea, spastic

paraparesis

Geneticsfor

theneurologist

187

Table 2 (continued )

InheritancePattern*

Stroke-like

Lesions

BilateralBasal

GangliaLesions

Encephalopathy Epilepsy CognitiveDecline

Ataxia OcularSigns

SensorineuralHearing Loss

Other NeurologicSigns

Other SystemicFindings

PDHA1 XL X X Spastic quadriplegiaSLC25A4/

ANT1AD, AR X Cardiomyopathy

(AR)POLG2 AD XDARS2 X Leukoence phalopathyDDP1/ TIMM8A XL X X X Deafness and dys-

tonia (Mohr-Tra-nebjaerg

syndrome)z

MFN2 AD Weakness, peripheralneuropathy (Charcot-Marie-Tooth disease

2A2)PDHA1 XL X X HypotoniaFRDA AR X Friedreich ataxia#,

neuropathyCardiomyopathy,

diabetesABC7 XL X Ataxia Sideroblastic

anemia

*AD ¼ autosomal dominant; AR ¼ autosomal recessive; IUGR ¼ intrauterine growth restriction; XL ¼ X-linked.†Likely inheritance pattern; not yet confirmed.‡Bjornstad syndrome: congenital sensorineural hearing loss, pili torti(twisted hair) (OMIM # 262000).§GRACILE syndrome: Growth retardation, aminoaciduria, cholestasis, iron overload, lactic acidosis, and early death (OMIM # 603358).kBarth syndrome: X-linked disease characterized by dilated cardiomyopathy with endocardial fibroelastosis, a predominantly proximal skeletal myopathy, growth retardation, neutropenia, and organic

aciduria, particularly excess of 3-methylglutaconic acid (OMIM # 302060).zMohr-Tranebjaerg syndrome: progressive syndrome with sensorineural hearing loss, cortical blindness, dystonia, fractures, and mental deficiency (OMIM # 304700).#Friedreich syndrome: Neurodegenerative disorder characterized by progressive gait and limb ataxia with associated limb muscle weakness, absent lower limb reflexes, extensor plantar responses,

dysarthria, and decreased vibratory sense and proprioception ( OMIM # 229300).

A.Goldstein,P.Bhatia,andJ.M

.Vento188

Genetics for the neurologist 189

the inheritance pattern for nuclear mitochondrial genes

follows mendelian laws: autosomal dominant, autosomal

recessive, or X-linked.47

Studies of nDNA can be conducted less invasively than

tissue biopsies through blood and have evolved from

single-gene tests to panels for a particular clinical syndromeor RC defect (eg, complex I deficiency panel) to next-

generation sequencing, which includes hundreds of rele-

vant nuclear mitochondrial genes. With the discovery ofnuclear genes causing mitochondrial disease, clinical phe-

notypes and prognosis are becoming more defined. Table 2

offers an overview of the relevant nuclear mitochondrialgenes and their clinical manifestations.4,24,48,49

The list of nuclear genes associated with mitochondrial

dysfunction is constantly expanding,50 especially with theadvent of commercially available next-generation sequen-

cing. The first identified nuclear gene mutation responsible

for mitochondrial disease was SDHA, which is found insiblings with Leigh syndrome,51 and to date, mutations

have been reported in more than 100 nuclear-encoded

genes.48 In the near future, this number is expected toexpand. Advances in sequencing technology have already

allowed clinicians to better diagnose and treat their

patients, especially in the setting of nuclear mutations ininfantile mitochondrial disease.1

Genes of nDNA are required by the mitochondria for

many functions, including RC complex structural proteinsand assembly factors; mtDNA maintenance (replication,

maintenance, and translation); CoQ10 biosynthesis; main-

tenance of the lipid milieu of the inner mitochondrialmembrane; solute carrier across the inner mitochondrial

membrane; fission and fusion; and mitochondrial biogen-

esis.52,53 Mutations disrupting mtDNA maintenance canproduce single or multiple mtDNA deletions or and cause

the clinical syndromes of MDDS and MNGIE.

Historically, mitochondrial disease has largely beenclassified by clinical symptoms, biochemical testing, and

tissue biopsies. The advent of next-generation sequencing

has enabled molecular diagnosis to be an importantdiagnostic element in the characterization of mitochondrial

syndromes and allowed for more accurate family counsel-

ing. With likely hundreds of nuclear genetic etiologies formitochondrial disease, a classification system is helpful for

neurologists to understand the pathophysiology of clinical

symptoms.

RC DysfunctionNuclear genes are responsible for most of the structural

proteins and assembly factors in the RC. Table 2 sum-

marizes the more common gene mutations grouped byeach RC complex along with the clinical manifestations. A

single nuclear gene mutation can lead to single biochemical

abnormality (ie, NDUFSV1 mutation causing isolatedcomplex I deficiency) or may cause multiple RC complex

deficiencies (ie, POLG mutations causing combined com-

plex I þ IV deficiencies). In pediatric patients withmitochondrial disease, up to 25% of cases have multiple

RC deficiencies, usually due to mutations involving mtDNA

maintenance (POLG), mtDNA deletion (Kearns-Sayre syn-

drome), mitochondrial translation defects, or a CoQ10biosynthesis defect.54

CoQ10 DeficiencyCoQ10, also called ubiquinone, is a potent antioxidant as

well as an electron carrier in the RC between complexes I þII to complex III. CoQ10 is synthesized in the mitochon-

dria and involves at least 12 genes. Defects in the nuclear

genes are inherited in an autosomal recessive manner. Adeficiency of CoQ10 may appear on tissue biopsy analysis

as a combined RC defect involving complexes I þ III or II

þ III.55

Clinically, 6 major phenotypes have been identified13,56:

�

encephalomyopathic form with seizures and ataxia;

�

multisystem infantile form with encephalopathy, cardi-

omyopathy, and renal failure;

� predominantly cerebellar form with ataxia and cerebellar

atrophy;

�

Leigh syndrome with growth retardation; � isolated myopathic form; and

�

steroid-resistant nephrotic syndrome.

There are 6 primary biosynthetic genes: PDSS1, PDSS2,

COQ2, COQ6, COQ9, and ADCK3 (CABC1 and COQ8);and involvement of 2 other genes: ETFDH and APTX; that

can lead to secondary deficiencies. Testing for the majority

of these genes is clinically available and an importantcondition as primary biosynthetic disorders are amenable

to treat with supplemental CoQ10.56

Defects of mtDNA TranslationThe nDNA encodes proteins that are essential for mtDNAprotein synthesis. A mutation in one of these genes, RARS2,

has been described as a progressive clinical syndrome of

lactic acidosis and pontocerebellar hypoplasia, and severeintractable epilepsy.57 Other translational genes that have

been characterized are included in Table 2.

Disorders of mtDNA MaintenancemtDNA Depletion Syndromes (MDS)MDS are a genetically heterogenous group of disorders with

the same molecular end result of reduced amount of

mtDNA (known as copy number) in specific tissues dueto mutations in genes that affect the mitochondrial nucleo-

tide pools or the mtDNA replication fork and lead to

decreased mtDNA copy number (depletion of mtDNA).58

Symptoms of mtDNA depletion syndrome vary by age but

include specific phenotypes of encephalomyopathy, hepa-

topathy, and isolated myopathy. Infants tend to have moreprominent lactic acidosis, failure to thrive, and hypotonia.

Children tend to have prominent epilepsy and liver

involvement. Muscle weakness tends to have a bimodalpresentation in childhood and then in adulthood.

A. Goldstein, P. Bhatia, and J.M. Vento190

Prominent symptoms with increasing age include migraine-

like headaches, ataxia, polyneuropathy, cognitive impair-

ment, psychiatric symptoms, and gastrointestinalsymptoms.41

mtDNA depletion syndromes have become an important

cause of inherited metabolic disorders, especially in chil-dren, but also in adults. The manifestations vary from a

tissue-specific mtDNA depletion to widespread multisyste-

mic disorders. Nine genes are known to underlie this groupof disorders, and many disease genes are still unidentified.

However, the disease mechanisms seem to be intimately

associated with mtDNA replication and nucleotide poolregulation. We review here the current knowledge on the

clinical and molecular genetic features of mtDNA depletion

syndromes. These syndromes have been divided into 3clinical categories: myopathic (TK2), encephalomyopathic

(SUCLA2, SUCLG1, RRM2B, and TYMP), and hepatocer-

ebral (DGUOK, MPV17, POLG, and C10orf2). However,these categories are not discrete, and they can have clinical

overlap, especially as the disease progresses. POLG-related

disorders would be reviewed in most detail, followed by abrief description of the other mitochondrial depletion

syndromes.

POLG-Related DisordersPOLG gene mutations are the most common cause ofinherited mitochondrial diseases in children and adults.

POLG encodes for human POLG, the only polymerase

involved in mtDNA replication and repair; thus, abnorm-alities can lead to mtDNA mutations or multiple deletions

or both. Currently, more than 150 mutations within POLG

are known, with a map of mutations updated on a regularbasis available at /http://tools.niehs.nih.gov/polg/S.59

Strict genotype-phenotype correlation is not possible, as

the clinical presentation is not solely dependent on thePOLG genotype.60,61 Like other mitochondrial disorders,

they involve multiple organ systems, mainly central and

peripheral nervous system, liver, muscle, and other organs,such as the gastrointestinal system.60 Disease progression of

POLG-related disorders is highly unpredictable. Neurologic

manifestations in younger patients mainly consist ofseizures, lactic acidosis, and hepatic failure. However, older

individuals mainly present with myopathy, sensory ataxia,

and CPEO. Neuroimaging findings include an increasedsignal on T2-weighted imaging of the posterior thalami,

dentate nuclei, inferior olives, and occipital cortex.

POLG-related disorders are classified as 6 recognizablephenotypes with symptom overlap, which actually creates a

disease spectrum.

1.

AHS: AHS is the most severe phenotype of the POLG-related disorders.62,63 Many children with AHS have

typical development before the onset of symptoms. AHS

typically presents before 4 years of age, but may presentthrough early adulthood with seizures, developmental

delay with episodic regression, and hepatopathy due to

anticonvulsants. Anticonvulsants, especially VPA, maycause the liver enzymes to elevate, raising the index of

suspicion of POLG. Many mutations in POLG have been

reported and may present in either an autosomal

recessive or dominant fashion. The clinical trial ofrefractory seizures, episodic psychomotor regression,

and a characteristic hepatopathy has been described as

the classic presentation.64 However, the clinical pheno-type and progression are highly variable. The disease

typically starts in infancy or in early childhood with

seizures. Although no seizure semiology is typical forAHS, as the disease progresses, seizures evolve into

more complex and refractory syndromes, such as focal

status epilepticus, EPC, or multifocal myoclonic epi-lepsy. Characteristic EEG findings in the beginning may

be unilateral occipital rhythmic high-amplitude slow

activity and superimposed polyspikes.65 The seizure fociusually shift on EEG with time. The response to antic-

onvulsants is also variable. VPA must be used with

extreme caution when the etiology of seizures isunknown, as it can precipitate liver dysfunction in

AHS.33,66 Cognitive decline is characteristic and may

be the result of neuronal gliosis, refractory seizures, andhigh dosages of various antiepileptic medications. The

characteristic early occipital lobe involvement leads to

migraine with visual auras (in older children) andcortical blindness, and it may also serve as the initial

seizure focus. Other characteristic neurologic symptoms

include hypotonia, sensory ataxia or cerebellar ataxia orboth, extrapyramidal movements, peripheral neuropa-

thy, and progressive spastic paraparesis (due to the

destruction of cerebral cortex). Hepatic involvement isalso highly variable. It can progress to fulminant end-

stage liver disease in few months. The disease progres-

sion can be variable, marked by periods of stability. Lifeexpectancy from onset of symptoms ranges from

3 months to 12 years.60 Brain computed tomography

or MRI may be initially normal or demonstrate gliosis inoccipital lobe regions and generalized atrophy. Cerebel-

lum, basal ganglia, thalamus, and brainstem are sequen-

tially involved.67

2.

MEMSA: MEMSA includes a spectrum of disorders with

epilepsy, myopathy (distal and proximal), and ataxia

without ophthalmoplegia.68 It may include some of thepatients previously classified as having MIRAS and

SCAE. Subclinical sensory polyneuropathy leading to

ataxia, starting in teenage years, is usually the first signof the disease. Subsequently, epilepsy develops, begin-

ning as focal but rapidly progressing to refractory

generalized type, accompanied by progressive encepha-lopathy. Myoclonus and myopathy may also be present.

Conspicuous absence of ragged red fibers from muscle

biopsy clearly distinguishes this from myoclonic epi-lepsy with ragged red fibers syndrome.68

3.

Ataxia neuropathy spectrum (ANS): ANS is an auto-

somal recessive POLG disorder, in which multipledeletions in mtDNA occur.69 This spectrum, previously

referred to as sensory ataxic neuropathy, dysarthria,

ophthalmoplegia, encompasses several clinical syn-dromes that include ataxia, neuropathy (sensory, motor,

Genetics for the neurologist 191

or mixed), bulbar dysfunction, and PEO without myo-

pathy (in contrast to MEMSA).70 The disease onset is

usually in early teenage years to late third decade. LikeMEMSA, it includes many patients previously classified

as MIRAS or SCAE. Epileptic encephalopathy may be a

component, but disease course is usually less progres-sive. Psychiatric disturbance is common but may be

underrecognized especially in adult-onset mitochondrial

disease. Migrainous headaches may precede symptomsby many years. Other features include myoclonus,

blindness, hearing loss, and varying degree of liver

failure. Muscle pathology is often normal.

4. Childhood myocerebrohepatopathy spectrum (MCHS):

MCHS is the rarest form of POLG spectrum. Disease

onset is between first few months of life and 3 years. Thedisease course is rapidly progressive with a fatal out-

come. Neurologic manifestations include developmental

delay, encephalopathy, dementia, myopathy, and hypo-tonia. In contrast to AHS, seizures are not a common

manifestation in MCHS. Other features include failure to

thrive, lactic acidosis, liver failure, renal tubular acido-sis, pancreatitis, cyclic vomiting, and hearing loss.71

Brain MRI can show generalized atrophy. Early disease

onset, hepatopathy, encephalopathy, and a fatal outcomemake this disorder resembles AHS. However, severe

myopathy, specific liver pathology, and nonspecific MRI

brain findings help differentiate MCHS from AHS.63

5.

Autosomal recessive PEO: adult-onset PEO without

systemic involvement is the hallmark of autosomal

recessive PEO.72 Molecular genetic confirmation ofPOLG is required for diagnosis, as isolated PEO can

also be seen in thyroid disorders or myasthenia gravis.

PEO may be the first manifestation of other moresystemic disorders in the POLG spectrum like

ANS.73,74 Other associated features may include neuro-

pathy, myopathy, and sensory ataxia.75

6.

Autosomal dominant PEO (adPEO): adPEO is another

adult-onset disorder, which manifests as progressive

weakness of extraocular eye muscles,72,73 along withproximal myopathy and wasting leading to exercise

intolerance. CPEO þ (CPEO-plus) manifests as sensor-

ineural hearing loss, axonal neuropathy, cerebellarataxia, depression, parkinsonism, hypogonadism,76

and cataracts. Mutations in POLG1, ANT1, or C10orf2

(TWINKLE) may result in multiple mtDNA deletionsleading to this disorder.77

MNGIEMNGIE is an autosomal recessive disorder caused by

mutations in the nuclear TYMP gene coding for the enzymethymidine phosphorylase. Accumulation of thymidine is

toxic and leads to mtDNA instability and RC dysfunction.

Symptoms include severe gastrointestinal dysmotility,cachexia, ptosis and external ophthalmoplegia, peripheral

sensorimotor neuropathy, and a diffuse leukoencephalo-

pathy.78 Diagnosis is made by the detection of elevatedthymidine or decreased thymidine phosphorylase levels in

blood. Confirmatory molecular testing can be performed

with TYMP sequencing. MNGIE is rare, but treatment via

allogenic hematopoetic stem cell transplantation is availableas part of a research protocol, making MNGIE perhaps the

only mitochondrial disorder that is potentially curable.78

Solute Transport Across the InnerMitochondrial MembraneSolute transporters must shuttle specific molecules across

the relatively impermeable inner mitochondrial membrane.SLC25A22 is the mitochondrial glutamate carrier. Muta-

tions in this gene have been associated with neonatal or

early infantile epileptic encephalopathy with burst suppres-sion EEG (Otohara syndrome).79

Symptoms include myoclonic and focal seizures from

first few days of life, microcephaly, hypotonia, and globaldevelopmental delay. Neuroimaging may reveal cerebellum

and corpus callosum hypoplasia, abnormal gyral pattern in

temporoparietal area, and hypomyelination of temporalpoles.24

Another important solute transporter is ANT1

(SLC25A4), which transports ATP out of the mitochondrialmatrix in exchange for adenosine diphosphate. Mutations

in ANT1 can cause adPEO, cardiomyopathy, and skeletal

myopathy.80 Solute transporters are a common, growinggroup of disorders that may be amenable to specific

treatments.

ConclusionsClinical suspicion for nuclear mitochondrial disorders iswarranted when there are neurologic features and multi-

system involvement. Clinical features, biochemical studies,

and neuroradiologic features can help direct diagnostictesting. Determination of a molecular etiology is not always

possible; however, rapidly evolving genomic technology

will greatly improve diagnostic capabilities. Expandedmultigene panels and genomic testing will allow clinicians

to provide more accurate recurrence risk counseling to

families. Additionally, it is expected to improve our under-standing of the spectrum and natural history of the vast

range of mitochondrial disease in time. The advent of

extensive genetic testing may require a clinician to use apatient’s genotype to help determine symptomatology.

Thorough examination, family history, and functional and

biochemical testing will remain critical elements to inter-pret test results and counsel families. Therefore, the

clinician must be aware not only of the clinical diagnostic

signs and the clinical phenotypes of mitochondrial disease,but also of the different diagnostic testing available. We

hope this paper serves as a resource for the clinician who

must gather the clinical information, send or process thebiochemical and molecular data, and arrive at a diagnosis

that not only best fits the clinical symptoms, but allows for

further treatment options and genetic counseling of thefamily in the context of a nuclear mitochondrial disorder.

A. Goldstein, P. Bhatia, and J.M. Vento192

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