A Novel Variant of ATP5MC3 Associated with Both Dystonia and Spastic Paraplegia

Abstract

Background: In a large pedigree with an unusual phenotype of spastic paraplegia or dystonia and autosomal dominant inheritance, linkage analysis previously mapped the disease to chromosome 2q24-2q31.

Objective: The aim of this study is to identify the genetic cause and molecular basis of an unusual autosomal dominant spastic paraplegia and dystonia.

Methods: Whole exome sequencing following linkage analysis was used to identify the genetic cause in a large family. Cosegregation analysis was also performed. An additional 384 individuals with spastic paraplegia or dystonia were screened for pathogenic sequence variants in the adenosine triphosphate (ATP) synthase membrane subunit C locus 3 gene (ATP5MC3). The identified variant was submitted to the "GeneMatcher" program for recruitment of additional subjects. Mitochondrial functions were analyzed in patient-derived fibroblast cell lines. Transgenic Drosophila carrying mutants were studied for movement behavior and mitochondrial function.

Results: Exome analysis revealed a variant (c.318C > G; p.Asn106Lys) (NM_001689.4) in ATP5MC3 in a large family with autosomal dominant spastic paraplegia and dystonia that cosegregated with affected individuals. No variants were identified in an additional 384 individuals with spastic paraplegia or dystonia. GeneMatcher identified an individual with the same genetic change, acquired de novo, who manifested upper-limb dystonia. Patient fibroblast studies showed impaired complex V activity, ATP generation, and oxygen consumption. Drosophila carrying orthologous mutations also exhibited impaired mitochondrial function and displayed reduced mobility.

Conclusion: A unique form of familial spastic paraplegia and dystonia is associated with a heterozygous ATP5MC3 variant that also reduces mitochondrial complex V activity.

Figure 1.. ATP5MC3 mutation and conservation analysis.

(A) c.318C>G mutation in exon 4 of ATP5MC3 resulting in (B) p.Asn106Lys substitution in the c subunit. (C) Electropherogram demonstrating heterozygous mutation near the start of exon 4. (D) ARNP motif conserved among c subunit genes APT5MC1–3, Drosophila, and bacteria including G. bethesdensis. Gln is substituted for Asn in S. simulans and E. coli.

Figure 2.. Decreased complex V function in ATP5MC3-associated disease.

A significant decrease was observed in relative ATP synthase activity in fibroblast cell lines derived from proband and affected father compared to two unrelated control fibroblast cell lines, with an associated decrease in cellular ATP and oxygen consumption in patient fibroblast cells lines. (* p-value <0.01).

Figure 3.. Drosophila geotaxis assay.

(A) Mean distance traveled in geotaxis assay for transgenic flies with arm-GAL4 driver with or without UAS constructs (see also supplementary video). Wild type flies compared to UAS-WT overexpression flies showed no significant difference. Flies carrying p.N102K and p.N102E expression constructs traveled significantly shorter distances. (B) Aging flies showed decreasing activity with age but demonstrated consistent differences between transgenic lines. (** p-value <0.001).

Figure 4.. Drosophila complex V activity in transgenic flies.

Relative ATP synthase activity among control and UAS-GAL4 transgenic flies. Significant reductions were noted in flies containing both arm-GAL4 driver and UAS constructs with either p.N102E or p.N102K mutations, but not with wild type. WT1 is a yellow mutant, which retains normal neuromuscular function. WT2 contains the CyO (curly wing) balancer used to create UAS-GAL4 transgenic lines. (* p-value <0.01; ** p-value <0.001).