Reduced mitochondrial Ca2+ transients stimulate autophagy in human fibroblasts carrying the 13514A>G mutation of the ND5 subunit of NADH dehydrogenase

Abstract

Mitochondrial disorders are a group of pathologies characterized by impairment of mitochondrial function mainly due to defects of the respiratory chain and consequent organellar energetics. This affects organs and tissues that require an efficient energy supply, such as brain and skeletal muscle. They are caused by mutations in both nuclear-and mitochondrial DNA (mtDNA)-encoded genes and their clinical manifestations show a great heterogeneity in terms of age of onset and severity, suggesting that patient-specific features are key determinants of the pathogenic process. In order to correlate the genetic defect to the clinical phenotype, we used a cell culture model consisting of fibroblasts derived from patients with different mutations in the mtDNA-encoded ND5 complex I subunit and with different severities of the illness. Interestingly, we found that cells from patients with the 13514A4G mutation, who manifested a relatively late onset and slower progression of the disease, display an increased autophagic flux when compared with fibroblasts from other patients or healthy donors. We characterized their mitochondrial phenotype by investigating organelle turnover, morphology, membrane potential and Ca 2+ homeostasis, demonstrating that mitochondrial quality control through mitophagy is upregulated in 13514A4G cells. This is due to a specific downregulation of mitochondrial Ca 2+ uptake that causes the stimulation of the autophagic machinery through the AMPK signaling axis. Genetic and pharmacological manipulation of mitochondrial Ca 2+ homeostasis can revert this phenotype, but concurrently decreases cell viability. This indicates that the higher mitochondrial turnover in complex I deficient cells with this specific mutation is a pro-survival compensatory mechanism that could contribute to the mild clinical phenotype of this patient. Mitochondrial disorders include a wide range of pathological conditions characterized by defects in organelle homoeostasis and energy metabolism, in particular in the electron transport chain (ETC) complexes. They are mostly caused by mutations in nuclear-or mtDNA-encoded genes of the respiratory chain complexes leading to a variety of clinical manifestations, ranging from lesions in specific tissues, such as in Leber's hereditary optic neuropathy, to complex multisystem syndromes, such as myoclonic epilepsy with ragged-red fibers, Leigh syndrome or the mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes syndrome (MELAS). 1,2 Despite the detailed knowledge of the molecular defects in these diseases, their pathogenesis remains poorly understood. The heterogeneity of signs and symptoms depends on the diversity of the genetic background and on patient-specific compensatory mechanisms. Several studies investigated the consequences of nuclear DNA mutations on intracellular organelle physiology and Ca 2+ homeostasis. 3,4 Here we analyzed a cohort of patients with mutations in the mtDNA-encoded ND5 subunit of NADH dehydrogenase in order to correlate the clinical phenot...

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“…Scale bars, 10 μm. Effect of FATE1 expression on ER–mitochondria distance in H295R/TR N‐Flag FATE1 cells measured using a split‐GFP probe . Quantification is shown in the graph.…”

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