LOS ANGELES – Having just a small amount of heteroplasmy –– multiple mitochondrial genotype variants coexisting in the same cell –– may result in electron transport chain (ETC) abnormalities that contribute to mitochondrial dysfunction in people with single large-scale mitochondrial DNA deletion syndromes (SLSMDSs).
This finding, from a study conducted by Baylor Genetics researchers and presented at the annual meeting of the American College of Medical Genetics and Genomics, shows that low heteroplasmy plays a bigger role in SLSMDSs than previously thought and suggests that a combination of genetic and functional tests are needed to effectively diagnose these disorders.
"It was always thought that these patients had a relatively high degree of heteroplasmy for these mitochondrial deletions in order to have a phenotype," Christine Eng, chief medical officer of Baylor Genetics and a coauthor on the study, told GenomeWeb.
However, by correlating data between mitochondrial genome sequencing and functional ETC studies, the Baylor team found that patients with mitochondrial deletions involving even small levels of heteroplasmy and certain ETC deficiencies still resulted in the types of mitochondrial dysfunction characteristic of SLSMDSs.
The major disorders associated with SLSMDSs are chronic progressive external ophthalmoplegia (CPEO), Kearns-Sayre syndrome, and Pearson syndrome, each of which presents differently. CPEO is a late-onset disease characterized by ophthalmoplegia (paralysis or weakness of the eye muscles), ptosis (a drooping of the eyelid that covers all or part of the pupil), and varying degrees of skeletal muscle weakness. Kearns-Sayre syndrome is a more severe manifestation of CPEO, often manifesting before the age of 20. Kearns-Sayre can present all the symptoms observed in CPEO but typically with more pronounced muscle involvement.
Differing from the others, Pearson syndrome is an early-onset disease that typically manifests at infancy and initially presents with anemia and pancreatic insufficiency instead of muscle involvement. In many cases Pearson syndrome can spontaneously resolve after the first few years, and many surviving patients go on to be affected by later-stage Kearns-Sayre syndrome, suggesting that the two diseases share the same mechanism.
Xueyang Pan, a clinical biochemical genetics fellow at Baylor Genetics who led the study, explained that SLSMDS patients often show varying degrees of heteroplasmy, giving rise to the hypothesis that higher mutation loads may correlate with the severity of cellular dysfunction.
"However," he said, "we don't observe such a correlation, and so far, [we] don't have a clear idea about the threshold above which the mitochondrial deletion could cause a phenotype."
This led Pan and his colleagues to wonder whether low-level heteroplasmy might cause functional mitochondrial deficiencies. To investigate this, they conducted a retrospective study of all SLSMDS cases in Baylor Genetics' database where ETC functional testing and mitochondrial DNA (mtDNA) genome sequencing were performed on muscle samples.
From 1,118 cases, the team identified seven instances of CPEO and Kearns-Sayre syndrome that matched their criteria.
"Surprisingly, in all these cases we found a low heteroplasmy level with mitochondrial DNA deletions," Pan said. Additionally, clinical notes for these cases included cytochrome C oxidase (COX) deficiencies in muscle pathology biopsies. COX defects can lead to numerous dysfunctions, including loss of mental function, movement problems, hypertrophic cardiomyopathy, and eating difficulties.
The investigators amplified mtDNA samples with long-range PCR and sequenced them via next-generation sequencing, which revealed that all seven samples showed mtDNA deletions.
"All these deletions disrupt protein-coding genes involved in mitochondrial ETC functions," Pan said.
ETC deficiencies were then confirmed in functional studies that showed impaired activity of several ETC enzymes. Although most observed ETC abnormalities correlated with the deleted genes, some did not, which Pan said will be the topic of future work.
These findings, Pan said, underscore the clinical significance of low-level mitochondria deletions and highlight the value of integrating molecular and functional assessments for the accurate diagnosis of SLSMDSs.
"Low levels of heteroplasmy have led physicians to think [that] this was relatively not significant for their phenotype," said Eng. "Putting that together with functional data, however, brought more insight into the fact that low heteroplasmy can actually lead to clinical symptomatology."
As a result, the Baylor team suggests that physicians perform functional assessments on muscle samples to better verify the pathogenicity of mtDNA deletions in this set of syndromes.
"We learn a lot from the genome," Eng said, "but we think that there are ways of improving our understanding of the genome by adding additional functional studies."