High-resolution melting analysis of 15 genes in 60 patients with cytochrome-c oxidase deficiency

Abstract

Cytochrome-c oxidase (COX) deficiency is one of the common childhood mitochondrial disorders. Mutations in genes for the assembly factors SURF1 and SCO2 are prevalent in children with COX deficiency in the Slavonic population. Molecular diagnosis is difficult because of the number of genes involved in COX biogenesis and assembly. The aim of this study was to screen for mutations in 15 nuclear genes that encode the 10 structural subunits, their isoforms and two assembly factors of COX in 60 unrelated Czech children with COX deficiency. Nine novel variants were identified in exons and adjacent intronic regions of COX4I2, COX6A1, COX6A2, COX7A1, COX7A2 and COX10 using high-resolution melting (HRM) analysis. Online bioinformatics servers were used to predict the importance of the newly identified amino-acid substitutions. The newly characterized variants updated the contemporary spectrum of known genetic sequence variations that are present in the Czech population, which will be important for further targeted mutation screening in Czech COX-deficient children. HRM and predictive bioinformatics methodologies are advantageous because they are low-cost screening tools that complement largescale genomic studies and reduce the required time and effort.

The subunit composition and function of mammalian cytochrome c oxidase

“…The first case of a disease (early-onset leukodystrophic encephalopathy, myopathy, and growth retardation) based on mutations in a nuclear-coded gene of a COX subunit (VIb-1) was described in two children which carried a homozygous c.221G/A substitution in exon 2 of COX6B1, causing a p.R19H amino acid change (Massa et al, 2008). In four patients with COX deficiency non-synonymous mutations were identified in three structural genes (p.R85W (COX4I2), p.R71H (COX5A), p.K31del (COX7A1)) and in one assembly factor (p.R431W (COX10)) (Vondrackova et al, 2012). A further mitochondrial disease, manifesting with encephalomyopathy, hydrocephalus and hypertrophic cardiomyopathy due to a missense p.R20C mutation in the COX6B1 gene, coding for COX subunit VIb-1, was recently described (Abdulhag et al, 2015).…”

Multisystem mitochondrial diseases due to mutations in mtDNA-encoded subunits of complex I

“…In patients 1-8 and 10, total genomic DNA isolated from muscle biopsy or cultivated skin fibroblasts (P2) and mtDNA (NC_012920) was sequenced as described previously [6]. In patients 9 and 11-13, the mtDNA mutation m.13513G > A was detected by PCR-RFLP using mismatch primers (F: 5′-GTTTGCGGTTTCGA TGATGTGAT-3′; R: 5′-AACCATACC-TCTCAC TTCAACCTCCC-3′) and Bsp143I endonuclease (Thermo Fisher Scientific, Waltham, Massachusetts, USA).…”

Tissue‐ and Condition‐Specific Isoforms of Mammalian Cytochrome c Oxidase Subunits: From Function to Human Disease

“…Alternatively, COX4-2 may be expressed in a minor pancreatic cell type. Another heterozygous missense mutation was reported in a patient with COX deficiency but not functionally confirmed [164]. …”

“… 1 Note that additional heterozygous mutations have been identified in individual patients with COX deficiency in COX4I2 , COX5a , and COX6a2 but have not been functionally confirmed as disease causing [164]. …”