Mutation in the novel nuclear-encoded mitochondrial protein CHCHD10 in a family with autosomal dominant mitochondrial myopathy

Abstract

Mitochondrial myopathies belong to a larger group of systemic diseases caused by morphological or biochemical abnormalities of mitochondria. Mitochondrial disorders can be caused by mutations in either the mitochondrial or nuclear genome. Only 5% of all mitochondrial disorders are autosomal dominant. We analyzed DNA from members of the previously reported Puerto Rican kindred with an autosomal dominant mitochondrial myopathy (Heimann-Patterson et al. 1997). Linkage analysis suggested a putative locus on the pericentric region of the long arm of chromosome 22 (22q11). Using the tools of integrative genomics, we established chromosome 22 open reading frame 16 (C22orf16) (later designated as CHCHD10) as the only high-scoring mitochondrial candidate gene in our minimal candidate region. Sequence analysis revealed a double-missense mutation (R15S and G58R) in cis in CHCHD10 which encodes a coiled coil-helix-coiled coil-helix protein of unknown function. These two mutations completely co-segregated with the disease phenotype and were absent in 1,481 Caucasian and 80 Hispanic (including 32 Puerto Rican) controls. Expression profiling showed that CHCHD10 is enriched in skeletal muscle. Mitochondrial localization of the CHCHD10 protein was confirmed using immunofluorescence in cells expressing either wild-type or mutant CHCHD10. We found that the expression of the G58R, but not the R15S, mutation induced mitochondrial fragmentation. Our findings identify a novel gene causing mitochondrial myopathy, thereby expanding the spectrum of mitochondrial myopathies caused by nuclear genes. Our findings also suggest a role for CHCHD10 in the morphologic remodeling of the mitochondria.

Figure 1

Electron micrographs from the index patient shows a variety of ultrastructural abnormalities in the presence of normal myofibrillar architecture (a-d). Note the accumulation of glycogen (black arrows) and lipids (white arrow heads). There is also an increased number of mitochondria containing abnormal circular cristae (black arrow heads) and marginalization of mitochondria (panels b and c). Selected area (square) from panel (b) is shown in higher power in panel (d).

Figure 2

MitoCarta which is based on the published Maestro algorithm was used to identify the candidate mitochondrial gene. Chromosome 22 open reading frame 16 (C22orf16) was identified as the only high scoring mitochondrial gene in the candidate region.

Figure 3

Mutations of CHCHD10 in patients with mitochondrial myopathy. Sequencing of CHCHD10 in the index patient identified a double missense mutation (R15S; G58R) in cis in the coding region (a and b). Both mutations co-segregated with disease phenotype in this large mitochondrial myopathy pedigree (a). The first, a C to A substitution at position 43 at the level of coding DNA (c.43C>A), is predicted to result in an amino acid substitution of arginine by a serine at codon 15 at the protein level (p.R15S) (b). The second mutation at nucleotide position 172 (c.172G>C) is predicted to cause a substitution of a glycine by an arginine at codon 58 (p.G58R) (b). (c) Evolutionary conservation of amino acids in the mutated region of CHCHD10 in various species. Mutated amino acids are highlighted in yellow. Nonconserved residues are shown in red.

Figure 4

The coiled-coil helix coiled-coil helix protein (CHCHD10) is encoded by four exons. Both mutations occur in exon 2 (a). Predicted structural and functional domains of CHCHD10, a protein of 142 amino acids, are shown (b). The only predicted structural and functional domain is a CHCH domain (102-133). Sequence analysis revealed an N-terminal mitochondrial localization signal (M; amino acids 1-16).

Figure 5

Co-localization of CHCHD10 with mitochondria. HEK-293 cells were transfected with either wild-type (WT) CHCHD10 (a-c), or mutant (R15S/G58R) CHCHD10 (d-f). CHCHD10 is myc-tagged (green). CHCHD10 colocalizes with the mitochondrial marker, DsRed-Mito (red). Merged images are shown in yellow. Scale bars, 10 μm.

Figure 6

Confocal images of mitochondria labeled by transient transfection of DsRed-Mito in HEK-293 cells expressing either wild-type (WT) or mutant (R15S, G58R or R15S/G58R) CHCHD10 (a). Quantitative image analysis of mitochondrial length is shown in panel (b) (n>10 cells in at least three independent experiments; *, p < 0.05 versus WT; n.s., not significant; error bars represent means ± S.E.)