Functional validation of CoQ deficiency fibroblast model with COQ7 mutations

Motivation: Coenzyme Q10 (CoQ10) deficiency syndrome comprises a heterogeneous group of mitochondrial disorders characterized by a decrease in CoQ10 content in cells and tissues. Primary CoQ deficiencies are rare genetic conditions caused by mutations in COQ genes, whose encoded proteins are directly linked to the final biochemical pathway of CoQ biosynthesis. Early diagnosis is both essential and one of the most challenging issues of this disease, mainly due to the variety of associated clinical manifestations. Here we present four clinical cases of primary CoQ10 deficiency, which is presumably caused by COQ7 mutations. The motivation for this work is to validate it in a cellular model based on primary cultures from patients' skin fibroblasts, in order to complete the previously started molecular diagnosis by whole-exome sequencing. Methods: Cultured patients fibroblasts was the biological starting material of our study. CoQ10 levels were measured by HPLC-ECD. In order to study mitochondrial function and respiration, oxygen consumption rate (OCR) was analysed using Seahorse technology. In addition, COQ7, several COQ proteins and other mitochondrial proteins expression were analysed by Western Blotting. Results: Patients' fibroblasts showed a basal level of CoQ10 lower than control fibroblasts (HDF). Moreover, the chromatogram revealed a peak corresponding to DMQ10, which was not seen in HDF. OCR showed mitochondrial respiration was affected in terms of maximal respiration and spare capacity with respect to control cells. Western blot analysis revealed the absence of COQ7 protein in patients' fibroblasts. Moreover, several COQ proteins, which are involved in the CoQ10 biosynthetic pathway, presented a moderate decreased expression. However, several mitochondrial related proteins maintained their physiological levels, such as VDAC, NDUFA9, UQCRCII or mtCOII. Conclusions: CoQ10 deficiency was confirmed in patients' fibroblasts. Since DMQ10 is the substrate of the reaction catalyzed by COQ7 protein, DMQ10 accumulation indicates that the COQ7 reaction is impaired. These results reveal that COQ7 mutation identified in patients' fibroblasts affects protein expression, CoQ10 levels and mitochondrial respiration. Finally, our data support the previous diagnosis obtained by exome analysis, proving that in these clinical cases, the CoQ10 deficiency is being produced by the absence of COQ7 protein

Secondary CoQ10 deficiency, bioenergetics unbalance in disease and aging

Coenzyme Q10 (CoQ10) deficiency is a rare disease characterized by a decreased accumulation of CoQ10 in cell membranes. Considering that CoQ10 synthesis and most of its functions are carried out in mitochondria, CoQ10 deficiency cases are usually considered a mitochondrial disease. A relevant feature of CoQ10 deficiency is that it is the only mitochondrial disease with a successful therapy available, the CoQ10 supplementation. Defects in components of the synthesis machinery caused by mutations in COQ genes generate the primary deficiency of CoQ10. Mutations in genes that are not directly related to the synthesis machinery cause secondary deficiency. Cases of CoQ10 deficiency without genetic origin are also considered a secondary deficiency. Both types of deficiency can lead to similar clinical manifestations, but the knowledge about primary deficiency is deeper than secondary. However, secondary deficiency cases may be underestimated since many of their clinical manifestations are shared with other pathologies. This review shows the current state of secondary CoQ10 deficiency, which could be even more relevant than primary deficiency for clinical activity. The analysis covers the fundamental features of CoQ10 deficiency, which are necessary to understand the biological and clinical differences between primary and secondary CoQ10 deficiencies. Further, a more in-depth analysis of CoQ10 secondary deficiency was undertaken to consider its origins, introduce a new way of classification, and include aging as a form of secondary deficiency.Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucía, Grant/Award Numbers: UPO-1259581, UPO-126247, UPO-1265673; Instituto de Salud Carlos III, Grant/Award Number: PI17/01286; Ministerio de Educación, Cultura y Deporte, Grant/Award Numbers: FPU14/04873, FPU16/0326

Cellular Models for Primary CoQ Deficiency Pathogenesis Study

This work was supported by Junta de Andalucia grants P18-RT-4572, UPO-126247, UPO1265673 and BIO-177, the Instituto de Salud Carlos III FIS grant FIS PI20/00541, the FEDER Funding Pro-gram from the European Union, and CIBERER (U729)-ISCIII, and Spanish Ministry of Science, Innovation and Universities grant RED2018-102576-T.Primary coenzyme Q10 (CoQ) deficiency includes a heterogeneous group of mitochondrial diseases characterized by low mitochondrial levels of CoQ due to decreased endogenous biosynthesis rate. These diseases respond to CoQ treatment mainly at the early stages of the disease. The advances in the next generation sequencing (NGS) as whole-exome sequencing (WES) and whole-genome sequencing (WGS) have increased the discoveries of mutations in either gene already described to participate in CoQ biosynthesis or new genes also involved in this pathway. However, these technologies usually provide many mutations in genes whose pathogenic effect must be validated. To functionally validate the impact of gene variations in the disease’s onset and progression, different cell models are commonly used. We review here the use of yeast strains for functional complementation of human genes, dermal skin fibroblasts from patients as an excellent tool to demonstrate the biochemical and genetic mechanisms of these diseases and the development of human-induced pluripotent stem cells (hiPSCs) and iPSC-derived organoids for the study of the pathogenesis and treatment approaches.Junta de Andalucia P18-RT-4572 UPO-126247 UPO1265673 BIO-177Instituto de Salud Carlos III European Commission FIS PI20/00541FEDER Funding Pro-gram from the European Union Instituto de Salud Carlos III U729Spanish Ministry of Science, Innovation and Universities RED2018-102576-

Secondary coenzyme Q10 deficiencies in oxidative phosphorylation (OXPHOS) and non-OXPHOS disorders

CoQ deficiency study group: et al.We evaluated the coenzyme Q₁₀ (CoQ) levels in patients who were diagnosed with mitochondrial oxidative phosphorylation (OXPHOS) and non-OXPHOS disorders (n = 72). Data from the 72 cases in this study revealed that 44.4% of patients showed low CoQ concentrations in either their skeletal muscle or skin fibroblasts. Our findings suggest that secondary CoQ deficiency is a common finding in OXPHOS and non-OXPHOS disorders. We hypothesize that cases of CoQ deficiency associated with OXPHOS defects could be an adaptive mechanism to maintain a balanced OXPHOS, although the mechanisms explaining these deficiencies and the pathophysiological role of secondary CoQ deficiency deserves further investigation.This work was supported by grants from the Instituto de Salud Carlos III (FIS: PI12/01683, PI14/00005, PI14/00028 and PI14/01962), the Explora Ciencia Program (SAF2013-50139-EXP), the Departamento de Ciencia, Tecnología, from Universidad del Gobierno de Aragón (Grupos Consolidados B33), the Fondo Europeo de Desarrollo Regional (FEDER Funding Program) from the European Union. The CIBERER is an initiative of the ISCIII. UCLH/UCL received a proportion of funding from the Department of Health sNIHR Biomedical Research Centers funding scheme.Peer Reviewe