Print Page
Genetics
For a list of labs performing testing, please visit genetests.org
Diseases from mutations in nuclear DNA (nDNA)
Mutations in nuclear encoded OXPHOS subunits and non-OXPHOS mitochondrial proteins, associated with some aspect of the biogenesis of the electron transport chain are becoming recognized as probably the most common cause of mitochondrial disease (Leonard and Shapira, 2000). This is due in part to the increased number of proteins synthesized by the nucleus compared to the mitochondria genome, approximately 850 compared to 13, respectively . More importantly, some of these proteins are responsible for the control of electron transport chain structure and function. Autosomal recessive inheritance of nuclear genetic defects is probably the most common etiology of pediatric patients with mitochondrial disorders .
Mutations involving subunits or ancillary proteins of the electron transport chain. mtDNA only encodes 13 subunits of the electron transport chain, while nDNA encodes 36 subunits of complex I, all 4 subunits of complex II, 10 subunits of complex III, 10 subunits of complex IV, and 14 subunits of complex V . In addition, nDNA also encodes coenzyme Q10 and cytochrome c.
As with mtDNA mutations, mutations in nuclear genes give rise to phenocopies and genocopies of syndromic and non-syndromic mitochondrial disorders. Although it is not completely clear, there are likely tissue specific expressions of some assembly factors as well as unknown variability in specific tissue expression of subunits and electron transport chain activity.
Complex I is the largest complex of the electron transport chain. Deficiencies in complex I are the most common electron transport defect, and nearly one-half of the patients present with Leigh syndrome with or without cardiomyopathy . Less common phenotypes of complex I deficiency include hepatopathy and tubulopathy, cardiomyopathy and cataracts, and lactic acidemia. Nuclear DNA mutations in complex I subunits have only been described in a few of the nuclear encoded subunits: NDUFS4 in a patient with encephalopathy and two patients with Leigh syndrome ; NDUFS7 and NDUFS8 mutations in patients with Leigh syndrome ; NDUFV1 in patients with leukodystrophy and myoclonic epilepsy ; and NDUFS2 in patients with hypertrophic cardiomyopathy and encephalomyopathy . Mutations in the subunit NDUFS1 have also been discovered in patients with complex I disease with hypotonia, seizures, ataxia and eyelid ptosis . In addition, mutations in genes NDUF8 and NDUFS4 have been shown to cause Leigh syndrome .
The genetic basis of greater than 50% patients with complex I dysfunction remains a mystery. Suggesting that other factors involved in the assembly or maintenance of this complex remains to be found .
Coenzyme Q10 shuttles electrons from complex I and II to complex III. Although uncommon, primary coenzyme Q10 deficiency can produce electron transport chain dysfunction. Family studies suggest an autosomal recessive inheritance pattern that segregates into at least 4 phenotypes: an encephalomyopathic form with exercise intolerance, mitochondrial myopathy, myogloburia, seizures, and ataxia , a multisystem infantile variant with encephalopathy, cardiomyopathy, ataxia, optic nerve atrophy, deafness, and nephrotic syndrome resulting in renal failure , Leigh syndrome with growth retardation, ataxia, and deafness , and an isolated myopathy . It is not clear how the reduction in coenzyme Q10 induces multiple phenotypes given the function within the electron transport chain. However, diagnosis is important as most all patients with coenzyme Q10 deficiency respond to coenzyme Q10 supplementation.
Complex II is the only electron transport chain complex whose four subunits are entirely encoded by nuclear genes. Deficiencies in complex II are rare and likely account for less than 5% of mitochondrial disease . The soluble proteins contain the succinate dehydrogenase activity while the membrane embedded subunits contain a cytochrome b and ubiquinone binding site. Missense mutations in one of the soluble subunits have been found in two families with Leigh syndrome . Another family was described with late onset degeneration with optic atrophy, ataxia, and myopathy .
Other patients with complex II deficiencies have been shown to have myopathy with exercise intolerance , isolated cardiomyopathy and progressive leukoencephalopathy . Investigations in these latter patients did not include DNA testing, so although by enzymology there were deficits in complex II, the location of the mutation(s) are unknown. Mutations in the large and small cytochrome b subunits have been demonstrated in autosomal dominant hereditary paraganglioma . Mutations in the soluble subunit, SDHB and the small cytochrome b subunit, SDHD, have been reported in patients with familial pheochromocytoma, with or without paraganglioma . A question to be answered is why some mutations clearly act as tumor suppressor genes with the other mutations give rise to Leigh syndrome.
Complex III has 10 subunits of nuclear origin and a single mtDNA transcribed product, cytochrome b. Patients presenting with myopathy, with and without myoglobinuria, have been shown to have mutations in the mtDNA product, cytochrome b . This sporatic form of mitochondrial myopathy is thought to arise from somatic mutations . A rare disorder, GRACILE syndrome (growth retardation, aminoaciduria, cholestasis, iron overload, lactacidosis, and early death) is an autosomal recessive disorder arising from a point mutation in a nuclear product, BCS1L . Mutations in this same protein can also produce the phenotype of neonatal proximal tubulopathy, hepatic cytolysis or failure, and encephalopathy . This inner mitochondrial protein is a chaperone necessary for the assembly of complex III. BCS1L appears to have an essential involvement in iron metabolism, however, this remains unknown .
Complex IV (COX) is the terminal enzyme in the mitochondrial electron transport chain and catalyzes the reduction of molecular oxygen by reduced cytochrome c. Ten of the 13 subunits are nuclear encoded. None of the nuclear encoded proteins have been associated with mitochondrial disease phenotypes described with COX deficiency . However, several nuclear gene products have been identified to be important in the biogenesis of the COX complex.
Mutations in the SURF-1 gene, a homologue of the yeast SHY1 gene, have been shown to give rise to Leigh syndrome. In fact, mutations in SURF-1 are the most common causes of Leigh syndrome with COX deficiency. Mutations resulting in premature stop codons have been found in all exons within the SURF-1 gene and appear to be the dominant type of mutation . SURF-1 mutations are not completely specific for Leigh syndrome as a few patients with non-Leigh syndrome with mutations in SURF-1 have been reported .
Mutations in SCO2 gene, a mitochondrial copper chaperone, have been found to cause fatal, early-onset hypertrophic cardiomyopathy with encephalopathy . The severity of this disorder is rather tissue specific to cardiac and skeletal muscle with little involvement in liver or fibroblasts. Interestingly, another mitochondrial copper chaperone, SCO1, has recently been found to give rise to hepatopathy and ketoacidotic coma but no cardiac symptoms . The findings, albeit with low patient numbers, suggest that SCO1 and SCO2 may have tissue specific copper delivery.
Two nuclear COX gene products involved with heme synthesis, COX10 and COX15, have been found to cause mitochondrial disease. A homozygous missense mutation in COX10 (heme A farnesyltransferase) was found in a patient with leukodystrophy and proximal tubulopathy . A misssense mutation in COX15 was found in a patient with early-onset fatal hypertrophic cardiomyopathy . COX15 is involved in the synthesis of heme A .
Two of the 16 subunits, a and A6L, are transcribed by the ATP6 and 8 genes within the mtDNA for complex V. Until recently, only mutations in the ATP6 gene have been reported to cause mitochondrial disease; NARP, MILS, or bilateral striatal necrosis. Data from Blue-Native electrophoresis indicated that several patients demonstrate isolated decrease in ATP synthase content. Howver, only two patients have been described with nuclear DNA defects, both have been assembly protein, ATP12 . Both patients showed early onset of signs and symptoms, dysmorphic features including large mouth and prominent nasal bridge, lactate elevations in urine and CSF, dygenesis of the corpus callosum and cortical atrophy, and the urine organic acids showed elevation of 3-methyl-glutaconic acid. 14 patients were shown to have decreased ATP synthase on Blue-Native electrophoresis, with only a single patient with the ATP12 mutation . This underscores the likelihood that further unknown nuclear factors involved in complex V expression.
Defects in maintenance of mitochondrial DNA (mtDNA) integrity
These disorders have been called defects of intergenomic signaling or nuclear-mitochondrial communication disorders. The consequences of these disorders are characterized by qualitative or quantitative alterations of mtDNA . The most common qualitative alteration is progressive external ophthalmoplegia (PEO), which is inherited as an autosomal dominant trait or recessive trait . Patients usually have ophthalmoparesis and exercise intolerance with onset between 18 to 40 years of age . Multiple genes are likely responsible for this disorder and some remain unknown, but at least 3 genes are involved. Mutations in a gene called Twinkle are associated with autosomal dominant PEO . The protein was named for the immunocytochemical staining pattern which looked like a star-like pattern in a night sky. The protein is associated with mitochondrial nucleoids, with highest expression in skeletal muscle. Another gene involved in autosomal dominant PEO is the missense mutation in the conserved polymerase domain of the mitochondrial gamma DNA polymerase (POLG1) . Interestingly, other missense mutations outside of both the polymerase and exonuclease domains were found in other families with apparently autosomal recessive PEO . Sporadic mutations in POLG1 have also been reported as the etiology of PEO . Mutations in the gene encoding a muscle-specific form of the adenine nucleotide translocator (ANT1) have been identified in some patients with autosomal dominant PEO .
Mutations in another gene, thymidine phosphorylase, cause an autosomal recessive disease of myoneurogastrointestinal encephalopathy (MNGIE) and PEO. This disorder produces PEO but with associated symptoms of dementia, progressive leukodystrophy, mitochondrial myopathy, periperhal neuropathy and prominent involvement of the gastrointestinal tract . Thymidine phosphorylase is widely expressed in human tissues, but paradoxically not in skeletal muscle in which multiple mtDNA deletions are present in some, but not all, patients . Both thymidine phosphorylase and adenine nucleotide translocator mutations affect the mitochondrial nucleotide pool and therefore may have similar pathogenic mutations .
Disorders of quantitative alterations of mtDNA are seen in severe or partial expression of mtDNA depletion. Clinically, these disorders are characterized by congenital or childhood forms of autosomal recessive myopathy or hepatopathy . Other tissues are often involved in both conditions, including the central nervous system. When liver and the central nervous system are primarily involved, mutations in the deoxyguanosine kinase gene have been described . In some patients, enzyme activities of complex I, III and IV are decreased in liver . The myopathic form is associated with mutations in the gene encoding thymidine kinase 2 . These latter patients usually present soon after birth with progressive weakness, hypotonia and areflexia. They die of respiratory failure within the first decade of life. In the pure form, only muscle seems to be involved .
Mutations in the POLG gene also cause the autosomal recessive disorder involving the liver and central nervous system, Alpers-Huttenlocher syndrome . The Alpers-Huttenlocher syndrome is part of the range of disorders comprising POLG mutations involving the brain and liver. Onset is between 1 month and 25 years of age . There is a clinical triad that is almost diagnostic consisting of refractory seizures, episodic psychomotor regression, cortical blindness and liver disease with micronodular cirrhosis . The clever and insightful investigation into the mechanism of this disorder has clarified much of POLG involvement in the broad clinical spectrum of POLG depletion disorders. There is evidence that POLG depletion is progressive and therefore, depletion or deletions test results cannot be used to exclude the diagnosis of POLG disease . Genetic studies have shown that mutations in the linker region plays an important role in the pathogenesis of Alpers syndrome in patients with POLG mutations, with 100% of the cases having the A467T and/or W748S substitutions . Mutations of two strongly pathological alleles of POLG seem to induce Alpers-Huttenlocker syndrome, while inheritance of one or two weaker POLG alleles give rise to non-Alpers-Huttenlocher syndrome that is usually later in onset and have more diverse phenotypes .
The unique nature of the symbiotic and semi-autonomous physiology of mitochondrion function gives rise to a wide range and variety of human disease. What was once a few rare diseases only described at Ground Rounds or as case reports in journals is now part of a growing occurrence of everyday clinic visits. This brief summary is only a partial list of mitochondrial disease. It is hoped that further research will help clarify why some patients have such varying phenotypes while others display phenocopies of particular signs and symptoms.
Diseases due to mutations in mtDNA
Human mtDNA is a 16,569 kb circular, double-stranded molecule, which encodes 37 genes, 2 rRNA genes, 22 tRNA genes, and 13 structural genes encoding the electron transport chain subunits. Over the course of time, it is speculated that mtDNA lost or adapted to use nDNA gene products needed for transcription, translation, and replication. Of the approximate 80 proteins that make up the electron transport chain, only 13 are encoded by the mtDNA. Complex II, coenzyme Q10, and cytochrome c are exclusively encoded by nDNA. In contrast, complex I, III, IV, and V contain some subunits encoded by mtDNA. There are 7 subunits of complex I (ND1-ND4, ND4L, ND5, and ND6), 1 subunit in complex III (cytochrome b), 3 subunits for complex IV (Cox I – Cox III), and subunits for complex IV (ATPase6 and ATPase 8). Although the mtDNA genome produces all the tRNAs and rRNAs necessary for intramitochondrial transcription, all of the regulatory machinery is produced by nDNA. The machinery for mtDNA replication and DNA repair are found in nDNA products.
Heteroplamsy and Threshold Effect. The advent of mitochondrial molecular pathology began in 1988 . Since this time, well over 100 pathological mutations within the mtDNA genome have been described. Each cell contains hundreds or thousands of mtDNA copies. Upon cell division, these copies distribute randomly among daughter cells. In normal tissues, all mtDNA molecules are identical (homoplasmy). Deleterious mutations of mtDNA usually affect some, but not all mtDNAs within a cell, tissue, or an individual (heteroplasmy). The clinical expression of a mtDNA mutation is largely determined by the relative proportion of normal and mutant mtDNA genomes in different tissues. The minimal number of mutant mtDNA genomes required to cause mitochondrial dysfunction in a particular organ varies between organs as a result of the energy demand of that organ or tissue (threshold effect). As the heteroplasmy approaches high proportions of mutated mtDNA, multiple organs begin to reach their threshold and the severity of mitochondrial dysfunction increases. This dynamic creates the phenotype of an individual with a mitochondrial disease.
Mitotic segregation. When the cell divides, the proportion of mutant mtDNA passed on in the daughter cells may shift and therefore, the phenotype may change. This process explains how certain patients with mtDNA disorders may actually manifest different mitochondrial diseases at different stages of their lives. When this process occurs during gamete production (meiotic segregation), each egg may have a different content of mutant mtDNA and explains why there might be sibling differences in disease expression.
Maternal inheritance. When the egg is fertilized, all paternal mtDNA is ubiquitinated and degraded. Therefore all mtDNA is derived from the mother (mitochondrial inheritance). A mother carrying a mtDNA mutation will pass it to all her children (boys and girls), but only her daughters will pass it on to all her children. There has been a single report of paternal mtDNA inheritance . This seems to be a unique circumstance and very likely an extremely rare occurrence .
A disease expressed in both boys and girls but with no evidence of paternal transmission, is strongly suggestive of a mtDNA point mutation. These diseases come in two flavors, those that impair mitochondrial protein synthesis and those that affect one of the 13 respiratory chain subunits encoded by mtDNA. Mutations that affect protein synthesis are usually mutations in the tRNA and rRNA genes and rearrangements of deletions or duplications.
Diseases of impaired protein synthesis: deletions and duplications. Although rarely can be inherited, single deletions are usually sporatic. The three most common are Pearson syndrome, Kearns-Sayre syndrome, and progressive ophthalmoplegia. Pearson syndrome is an often fatal multisystemic disease of infancy characterized by refractory anemia, vacuolization of marrow precursors and pancreatic fibrosis . The most common etiology is a mtDNA deletion of 4977 bp located between position 8488 and 13460 in the mtDNA molecule . The mtDNA deletions are usually more abundant in blood than in other tissues. Kearns-Sayre syndrome is a multisystemic progressive disorder characterized by impairment of eye movements (progressive external ophthalmoplegia), pigmentary retinopathy, heart block and onset before age 20 years . Other frequent signs are ataxia, dementia, short stature, diabetes mellitus, and deafness . Approximately 90% of patients with Kearns-Sayre syndrome have a large mtDNA deletion (1.3 – 10 kb) that is usually present in all tissues but often undetectable in blood cells, requiring the examination of muscle. Progressive external ophthalmoplegia, with or without proximal limb weakness, is characterized by paralysis of the extraocular muscles, ptosis and variably severe proximal limb weakness. These patients usually have a normal life span
Duplication of mtDNA can occur in either isolation or together with single deletions. These genotypes can be seen in patients with Kearns-Sayre syndrome or with diabetes mellitus and deafness. Duplications and duplication/deletions are rare and usually transmitted by maternal inheritance.
mtDNA point mutations. Well over 120 pathological mutations have been identified in mtDNA (DiMauro and Bonilla, 2003). Most of them are maternally inherited and phenotypically present with multisystemic manifestations. There are some that are sporadic and tissue specific. Two of the more common maternally inherited syndromes are mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) and myoclonus epilepsy with ragged red fibers (MERRF).
MELAS usually presents in children or young adults after normal early development . Symptoms include recurrent vomiting, migraine-like headache, and stroke-like episodes causing cortical blindness, hemiparesis, or hemianopia . MRI of the brain reveals ischemic events that do not correspond to a vascular distribution and do not conform to diffusion-weighted findings of arterial ischemic events . Approximately 80% have a mutation, A3243G in the tRNA Leu(UUR) gene (Goto et al., 2002). Phenocopies of MELAS have been found at other regions within the tRNA Leu, as well as other tRNA and protein-coding genes.
MERRF is characterized by myoclonus, generalized seizures, mitochondrial myopathy (ragged-red fibers), and cerebellar ataxia . Other signs include dementia, hearing loss, peripheral neuropathy, and multiple lipomas . Mutations in the tRNA Lys gene at positions A8344G, T8356C, and G8363A predominant . Approximately 80% or more have the A8344G mutation . Only a single patient with the complete syndrome harbored a mutation in a different tRNA, G611A in the tRNA Phe gene .
Mutations in tRNA are associated with multisystemic abnormalities. Almost any body system can be affected including the eye, hearing, endocrine, heart, gastrointestinal tract, and renal. Any combination of the systems and signs, especially if there are 3 or more systems involved without a clear etiology, should raise the suspicion of a mitochondrial disorder.
Defects of protein-coding genes. Although mutations in protein-encoding genes of the mtDNA can have multisystem abnormalities, many point mutations within these genes often escape the rules of mitochondrial genetics. These diseases affect single individuals and single tissues, most commonly skeletal muscle (DiMauro et al., 2003). Clinically, these patients may have exercise intolerance, recurrent myoglobinuria, and myalgia resulting from isolated defects of complex I, complex III, or complex IV. Isolated complex dysfunction is a result of mutations in genes encoding ND subunits, COX subunits, and cytochrome b . The lack of mitochondrial inheritance and isolated tissue expression suggests a de novo mutation early in stem cell development, after germ-layer differentiation. There may also be modifier nuclear genes that alter expression of the mitochondrial mutation. This is currently an area of active research.
Two common examples of defects in protein-coding gene mutations are neuropathy, ataxia, and retinitis pigmentosa (NARP)/maternally inherited Leigh syndrome (MILS) and Leber’s hereditary optic neuropathy (LHON). NARP most commonly affects children and young adults, and causes retinitis pigmentosa, dementia, seizures, ataxia, proximal neurogenic weakness and sensory neuropathy . Other clinical features can include short stature, sensorineural hearing loss, cardiac conduction defects, progressive external ophthalmoplegia and a mild anxiety disorder . Individuals with NARP can be stable for many years but suffer episodic deterioration during illness. NARP is part of a continuum of progressive neurodegenerative disorder, with NARP at one end of the spectrum and Leigh syndrome at the other.
Maternally inherited Leigh syndrome is a severe infantile encephalopathy characterized by hypotonia, spasticity, movement disorder, cerebellar ataxia, peripheral neuropathy and symmetrical lesions in the basal ganglia and the brainstem . Leigh syndrome or subacute necrotizing encephalomyelopathy, usually presents between 3 to 12 months of age, often followed by a viral illness. Decompensation during an illness is associated with psychomotor regression and lactic acidosis. Later onset can be seen in about 25% of patients after one year of age and into adulthood usually with slower progression . Strict criteria have been defined for this disorder with patients demonstrating progressive neurological disease with motor and intellectual developmental delay, signs and symptoms of brainstem and/or basal ganglia disease, raised lactate concentration in blood and/or cerebrospinal fluid and one or more of the following: characteristic MRI and/or CT findings, typical neuropathological changes and typical neuropathology in a similarly affected sibling . Before the advent of modern neuroimaging, the definitive diagnosis of Leigh syndrome was only made post mortem based on pathological findings . Findings are multiple focal symmetric necrotic lesions in the basal ganglia, thalamus, brainstem, dentate nuclei, and optic nerves. This is a spongiform appearance characterized by demyelination, gliosis, and vascular proliferation. Neuronal loss occurs but typically neurons are spared.
A variety of neurological symptoms are associated with Leigh syndrome . These include muscle weakness, hypo- or hyperreflexia, seizures, movement disorders, cerebellar ataxia, peripheral neuropathy, hypotonia, spasticity and dystonia. Respiratory symptoms due to brainstem abnormalities may include apnea, hyperventilation, or irregular respiration. Other brainstem induced abnormalities include difficulty in swallowing, persistent vomiting and abnormalities of thermoregulation. Ophthalmologic findings include eye movement abnormalities, retinitis pigmentosa, and optic atrophy. Systemic abnormalities include hypertropic cardiomyopathy , hepatomegaly or liver failure (Levine et al., 2003), or renal tubulopathy or diffuse glomerulocytic kidney damage .
The term Leigh-like disease is used for patients with signs and symptoms strongly suggestive of Leigh syndrome but who do not fulfill the above criteria. There might be atypical neuropathology such as variation in distribution of the lesion or the additional presence of unusual features, atypical neuroimaging, lack of elevated lactate in serum or CSF, or incomplete evaluation.
There are approximately 10% – 20% of individuals with maternally inherited Leigh syndrome with either the T8993G or the T8993C mutation in the MTATP6 gene (Makino et al., 1998). Both mutations are also associated with NARP. Greater than 50% of patients with NARP have a mutation at position 8993, at least those with lactate acidosis. In both Leigh syndrome and NARP the most common mutation is the T8993G transversion, but the T8993C has also been described . These mutations show the strongest genotype-phenotype correlation of any mtDNA mutations. The correlations between mutant load and disease severity is so strong that logistic regression models predicting the probability of a severe outcome can be performed . The T-to-G transversion is both clinically and biochemically more deleterious than the T-to-C change . Individuals with T8893G mutant loads below 60% are usually asymptomatic or only either have migraine headache or mild pigmentary retinopathy. There have been asymptomatic adults with mutant loads up to 75% . When the degree of heteroplasmy is between 70% – 90%, the NARP phenotype is present and when load is above 90%, the maternally inherited Leigh syndrome occurs. The less severe T9883C mutation requires a greater mutant load to express disease, with mutant loads of more than 90% needed to express a disease phenotype.
LHON is the second syndrome resulting from defects in mtDNA protein-coding genes. This disorder typically presents in young adults with painless acute or subacute loss of bilateral vision . Males are more commonly affected than females . Women tend to develop this disorder later in life and may be more severely affected. Typical age of onset is between 15 and 35 years but the range of onset can vary between 2 to 80 years . Visual loss typically begins painlessly and centrally in one eye. The second eye is usually affected weeks to months later. However, reports of simultaneous onset have also been reported. Rarely, loss of vision occurs after a prolonged interval or remains monocular . The acute phase begins with blurring of central vision and color desaturation. The most characteristic feature is an enlarging centrocecal scotoma. Visual acuities at the point of maximal visual loss range from no light perception to 20/20, with most patients deteriorating to acuities worse than 20/200 (Newman et al., 1995). Following the nadir, in some patients, acuity may improve. Improvement is more likely in patients with the T14484C mutation. Following the acute phase, individuals proceed into the atrophic phase of permanent legal blindness and large centrocecal scotoma.
LHON was the first maternally inherited mtDNA described at position 11778 affecting the ND4 subunit of complex I . The G to A transition mutation at position 11778 in the mtDNA remains the most common cause of LHON, accounting for over 70% of cases . Although over 40 other candidate point mutations have been found in patients with LHON only three common mutations, 90% of cases, distributed worldwide are found in most patients. These pathogenic mutations are 11778, 3460, and 14484 in the ND4, ND1, and ND6 subunits of complex I, respectively . There are 7 other primary LHON mutations but they have only been found in 2 or 3 families worldwide (Carelli et al., 2004). Other changes, called secondary/intermediary mutations, are population polymorphisms . Some of these mutations define the mtDNA haplogroup J and although debated, may play some modifying role .
An individual can only develop LHON when a primary mtDNA mutation is present. However, due to reduced penetrance, only approximately 50% of males and over 15% of females who harbor a primary LHON mutation develop blindness. Unknown environmental and genetic factors likely remain to be discovered that interact with primary mtDNA mutation to develop the disease.
