Recessive axonal Charcot-Marie-Tooth disease due to compound heterozygous mitofusin 2 mutations

Abstract

Objective: Mutations in mitofusin 2 (MFN2) are the most common cause of axonal Charcot-Marie-Tooth disease (CMT2). Over 50 mutations have been reported, mainly causing autosomal dominant disease, though families with homozygous or compound heterozygous mutations have been described. We present 3 families with early-onset CMT2 associated with compound heterozygous MFN2 mutations. Transcriptional analysis was performed to investigate the effects of the mutations.

Methods: Patients were examined clinically and electrophysiologically; parents were also examined where available. Genetic investigations included MFN2 DNA sequencing and dosage analysis by multiplex ligation-dependent probe amplification. MFN2 mRNA transcripts from blood lymphocytes were analyzed in 2 families.

Results: Compound heterozygosity for MFN2 mutations was associated with early-onset CMT2 of varying severity between pedigrees. Parents, where examined, were unaffected and were heterozygous for the expected mutations. Four novel mutations were detected (one missense, one nonsense, an intragenic deletion of exons 7 + 8, and a 3-base pair deletion), as well as 2 previously reported missense mutations. Transcriptional analysis demonstrated aberrant splicing of the exonic deletion and indicated nonsense-mediated decay of mutant alleles with premature truncating mutations.

Conclusions: Our findings confirm that MFN2 mutations can cause early-onset CMT2 with apparent recessive inheritance. Novel genetic findings include an intragenic MFN2 deletion and nonsense-mediated decay. Carrier parents were asymptomatic, suggesting that MFN2 null alleles can be nonpathogenic unless coinherited with another mutation.

Figure. Pedigrees and transcript analysis

(A) Pedigrees. Arrow = index case; N = normal genotype. (B) Family 1 MFN2 mRNA PCR from exons 4 to 19. All bands were sequenced. Two transcripts were separated by agarose gel electrophoresis in 1/II-1 and 1/II-2; the upper band encodes the maternal c.647T>C mutation, the lower paternal band, encodes a shortened transcript in which exon 6 is spliced to exon 9, confirmed by sequencing. 1/I-1 was confirmed to heterozygously express c.647T>C. Con = normal control PCR; H₂O = water control PCR. (C) Family 2 MFN2 DNA and mRNA sequencing. The thymine allele at base 922 was not evident in 2/II-1 and 2/I-1 in the mRNA sequencing. This allele is likely subjected to nonsense-mediated decay. Because of the nonsense-mediated decay of her maternal allele, 2/II-1 only expresses a cytosine at base c.1556, while it is heterozygously expressed in 2/I-2.