New genes and pathomechanisms in mitochondrial disorders unraveled by NGS technologies

Abstract

Next Generation Sequencing (NGS) technologies are revolutionizing the diagnostic screening for rare disease entities, including primary mitochondrial disorders, particularly those caused by nuclear gene defects. NGS approaches are able to identify the causative gene defects in small families and even single individuals, unsuitable for investigation by traditional linkage analysis. These technologies are contributing to fill the gap between mitochondrial disease cases defined on the basis of clinical, neuroimaging and biochemical readouts, which still outnumber by approximately 50% the cases for which a molecular-genetic diagnosis is attained. We have been using a combined, two-step strategy, based on targeted genes panel as a first NGS screening, followed by whole exome sequencing (WES) in still unsolved cases, to analyze a large cohort of subjects, that failed to show mutations in mtDNA and in ad hoc sets of specific nuclear genes, sequenced by the Sanger's method. Not only this approach has allowed us to reach molecular diagnosis in a significant fraction (20%) of these difficult cases, but it has also revealed unexpected and conceptually new findings. These include the possibility of marked variable penetrance of recessive mutations, the identification of large-scale DNA rearrangements, which explain spuriously heterozygous cases, and the association of mutations in known genes with unusual, previously unreported clinical phenotypes. Importantly, WES on selected cases has unraveled the presence of pathogenic mutations in genes encoding non-mitochondrial proteins (e.g. the transcription factor E4F1), an observation that further expands the intricate genetics of mitochondrial disease and suggests a new area of investigation in mitochondrial medicine. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi.

E4F1 controls a transcriptional program essential for pyruvate dehydrogenase activity

“…Strikingly, recent genetic studies designed to identify new mutated genes involved in unsolved cases of primary mitochondrial human disorders, led to the identification of a homozygous nonsynonymous mutation in the E4F1 gene of a patient showing reduced PDH complex activity, muscular defects, and lactate acidemia (36). This first indication that the E4F1-controlled program could be deregulated in a pathological situation provides an exciting clinical perspective to the present work.…”

COA7(C1orf163/RESA1) mutations associated with mitochondrial leukoencephalopathy and cytochrome c oxidase deficiency

“…We ruled out the presence of pathogenic mutations in mtDNA by Sanger sequencing. WES7 was then carried out. After standard filtering steps and assuming a recessive trait, we prioritised 13 genes with either one homozygous variant or two heterozygous variants.…”

E4F1-mediated control of pyruvate dehydrogenase activity is essential for skin homeostasis

“…It is worth noting, however, that a homozygous nonsynonymous mutation in the coding region of the E4f1 gene has been recently identified in a patient presenting clinical symptoms resembling those of Leigh syndrome patients (27). Although skin abnormalities have been reported only in some Leigh syndrome patients (28), they are part of the broad spectrum of clinical manifestations that are commonly observed in several mitochondrial disorders (29).…”