Delineation of C12orf65-related phenotypes: a genotype-phenotype relationship

Abstract

C12orf65 participates in the process of mitochondrial translation and has been shown to be associated with a spectrum of phenotypes, including early onset optic atrophy, progressive encephalomyopathy, peripheral neuropathy, and spastic paraparesis.We used whole-genome homozygosity mapping as well as exome sequencing and targeted gene sequencing to identify novel C12orf65 disease-causing mutations in seven affected individuals originating from two consanguineous families. In four family members affected with childhood-onset optic atrophy accompanied by slowly progressive peripheral neuropathy and spastic paraparesis, we identified a homozygous frame shift mutation c.413_417 delAACAA, which predicts a truncated protein lacking the C-terminal portion. In the second family, we studied three affected individuals who presented with early onset optic atrophy, peripheral neuropathy, and spastic gait in addition to moderate intellectual disability. Muscle biopsy in two of the patients revealed decreased activities of the mitochondrial respiratory chain complexes I and IV. In these patients, we identified a homozygous splice mutation, g.21043 T>A (c.282+2 T>A) which leads to skipping of exon 2. Our study broadens the phenotypic spectrum of C12orf65 defects and highlights the triad of optic atrophy, axonal neuropathy and spastic paraparesis as its key clinical features. In addition, a clear genotype-phenotype correlation is anticipated in which deleterious mutations which disrupt the GGQ-containing domain in the first coding exon are expected to result in a more severe phenotype, whereas down-stream C-terminal mutations may result in a more favorable phenotype, typically lacking cognitive impairment.

Figure 1

Pedigrees of the two families. Affected individuals are indicated by filled symbols.

Figure 2

(a) Left eye color fundus examination of patient A.IV-1 showing temporal pallor (arrow) of the optic disc accompanied by pink (normal) nasal region. (b) Visual field study of both eyes (Humphry Treshold 30-2) in patient A.IV-1 illustrating bilateral pseudo-temporal visual field loss (arrows). (c) Peripapillary RNFL of left eye. OCT study showing generalized thinning of retinal nerve fiber more dominant on the temporal side (green color – normal thickness, yellow color – borderline thickness, red color – clear thinning). (d) Fixed flexion contracture of the toes in patient A.IV-4. Note severe atrophy of dorsal feet muscles.

Figure 3

On T2 (FSE) sagittal view, (a) a short, hypoplastic corpus callosum is shown. The Rostrum is missing, and a posterior callosal hypogenesis is noted with an absence of the splenium. The optic chiasm is thin both in the sagittal (arrow head) as well as in the coronal (b) view indicating optic atrophy. Note the absence of the splenium on the T2WI axial view (c). There are bilateral high intensity foci (arrows) in the mesencephalon and the Thalamus, shown on T2 FLAIR image (d) with restricted diffusion, typical for mitochondrial disorders.

Figure 4

(a) Sequence analysis of C12orf65 in genomic DNA depicting a homozygous ‘AACAA' deletion in family A (left lower panel) and a g.21043T>A (c.282+2T>A) splice mutation in family B (right upper panel). (b) Left panel shows RT-PCR using cDNA using primers corresponding to positions c.124 and c.738 (NM_001143905.2) in exon 1 and exon 3, respectively. The wild-type expected 615-bp product is shown in the third lane from the right (labeled WT). Both parents harbor the wild-type product in addition to a shorter 305-bp product (sixth and seventh lanes from the right). Both affected individuals are homozygotes for the 305-bp product (fourth and fifth lanes from the right). In addition to the major wild-type and the mutant transcripts, a 646-bp product representing an alternative transcript (NM_001194995.1) is shown in both the mother and the affected patient (sixth and fourth lanes from the right, respectively). In patient B.V-3, residual 615-bp product is evident indicating the leaky nature of the splice mutation in family B (lane 4 from the right). Following sequencing of the 305-bp product, skipping of exon 2 was identified as depicted in the right side of part b. (c) A cartoon of the C12orf65 gene (upper panel) shows exons as blocks with light blue depicting non-translated regions and dark blue-translated regions. The black stars points to the c.413_417 delAACAA mutation in family A and the donor splice mutation in family B. Lower panel shows a scheme of the encoded protein. The C12orf65 mutations are depicted as black stars with their description underneath. The GGQ domain is marked in red and the amino-acid sequence resulting from the deleted 5-nt in family A is shown in orange within the dark green block on the C-terminal part of the protein.