Search

Iron–sulfur (Fe–S) clusters are essential cofactors for proteins that participate in fundamental cellular processes including metabolism, DNA replication and repair, transcriptional regulation, and the mitochondrial electron transport chain (ETC). ISCA2 plays a role in the biogenesis of Fe–S clusters and a recent report described subjects displaying infantile‐onset leukodystrophy due to bi‐allelic mutation of ISCA2. We present two additional unrelated cases, and provide a more complete clinical description that includes hyperglycinemia, leukodystrophy of the brainstem with longitudinally extensive spinal cord involvement, and mtDNA deficiency. Additionally, we characterize the role of ISCA2 in mitochondrial bioenergetics and Fe–S cluster assembly using subject cells and ISCA2 cellular knockdown models. Loss of ISCA2 diminished mitochondrial membrane potential, the mitochondrial network, basal and maximal respiration, ATP production, and activity of ETC complexes II and IV. We specifically tested the impact of loss of ISCA2 on 2Fe–2S proteins versus 4Fe–4S proteins and observed deficits in the functioning of 4Fe–4S but not 2Fe–2S proteins. Together these data indicate loss of ISCA2 impaired function of 4Fe–4S proteins resulting in a fatal encephalopathy accompanied by a relatively unusual combination of features including mtDNA depletion alongside complex II deficiency and hyperglycinemia that may facilitate diagnosis of ISCA2 deficiency patients.

Mutations in the mitochondrial genome, and in particular the mt-tRNAs, are an important cause of human disease. Accurate classification of the pathogenicity of novel variants is vital to allow accurate genetic counseling for patients and their families. The use of weighted criteria based on functional studies—outlined in a validated pathogenicity scoring system—is therefore invaluable in determining whether novel or rare mt-tRNA variants are pathogenic. Here, we describe the identification of nine novel mt-tRNA variants in nine families, in which the probands presented with a diverse range of clinical phenotypes including mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes, isolated progressive external ophthalmoplegia, epilepsy, deafness and diabetes. Each of the variants identified (m.4289T>C, MT-TI; m.5541C>T, MT-TW; m.5690A>G, MT-TN; m.7451A>T, MT-TS1; m.7554G>A, MT-TD; m.8304G>A, MT-TK; m.12206C>T, MT-TH; m.12317T>C, MT-TL2; m.16023G>A, MT-TP) was present in a different tRNA, with evidence in support of pathogenicity, and where possible, details of mutation transmission documented. Through the application of the pathogenicity scoring system, we have classified six of these variants as “definitely pathogenic” mutations (m.5541C>T, m.5690A>G, m.7451A>T, m.12206C>T, m.12317T>C, and m.16023G>A), whereas the remaining three currently lack sufficient evidence and are therefore classed as ‘possibly pathogenic’ (m.4289T>C, m.7554G>A, and m.8304G>A).

Distinguishing pathogenic from polymorphic changes poses significant problems for geneticists and despite 30 years of postgenomic experience this remains the case in mitochondrial genetics. Base substitutions in mitochondrial tRNA (mt-tRNA) genes are particularly difficult, but important, because they are common causes of pathology and associated with high rates of transmission. Providing accurate genetic advice to patients and their families is of paramount importance in disease prevention, and brings into sharp focus the factors used to distinguish pathogenic from polymorphic variants. We have reevaluated our pathogenicity scoring system for mt-tRNA mutations following a considerable increase in the number reported since the system was devised in 2004. This allowed us to address notable issues including the underestimation of "definitely pathogenic" mutations resulting from insufficient data collection. We illustrate the robustness of our revised scoring system using novel pathogenic and previously reported polymorphic changes and conclude that while clear evidence from single-fiber and/or trans-mitochondrial cybrid studies remains the gold standard for assigning pathogenicity, our scoring system is valuable for deciding which mt-tRNA mutations to investigate further using these labor-intensive techniques.