Exome sequencing identifies a DYNC1H1 mutation in a large pedigree with dominant axonal Charcot-Marie-Tooth disease

Abstract

Charcot-Marie-Tooth disease is characterized by length-dependent axonal degeneration with distal sensory loss and weakness, deep-tendon-reflex abnormalities, and skeletal deformities. It is caused by mutations in more than 40 genes. We investigated a four-generation family with 23 members affected by the axonal form (type 2), for which the common causes had been excluded by Sanger sequencing. Exome sequencing of three affected individuals separated by eight meioses identified a single shared novel heterozygous variant, c.917A>G, in DYNC1H1, which encodes the cytoplasmic dynein heavy chain 1 (here, novel refers to a variant that has not been seen in dbSNP131or the August 2010 release of the 1000 Genomes project). Testing of six additional affected family members showed cosegregation and a maximum LOD score of 3.6. The shared DYNC1H1 gene variant is a missense substitution, p.His306Arg, at a highly conserved residue within the homodimerization domain. Three mouse models with different mutations within this domain have previously been reported with age-related progressive loss of muscle bulk and locomotor ability. Cytoplasmic dynein is a large multisubunit motor protein complex and has a key role in retrograde axonal transport in neurons. Our results highlight the importance of dynein and retrograde axonal transport in neuronal function in humans.

Figure 1

Partial Pedigree Showing Mutation Status Open symbols represent unaffected individuals; filled squares represent affected males and filled circles affected females. Individual generations are numbered with roman numerals on the left. Individuals heterozygous for the p.His306Arg mutation are indicated as N/M. For the LOD-score calculation we assumed a rare autosomal-dominant model with a disease-allele frequency of 0.0001 and phenocopy rate of 0. The mutant DYNC1H1 allele is rare (frequency <0.001); because we only tested affected individuals to avoid issues of predictive testing and variable penetrance and because there are 12 meioses separating the nine sequenced individuals, the LOD score for the variant can be approximated as −log10(0.5¹²) = 3.6.

Figure 2

Schematic Representation of Human DYNC1H1 The N-terminal region of DYNC1H1 is represented by a horizontal black bar, and the stem domain (amino acids 53–1867) is indicated by a bracket above. Residues involved in DYNC1H1 dimerization (300–1140) are shown by a light blue bar; open boxes represent binding regions for intermediate (DYN1I; residues 448–703, solid line) and light-intermediate (DYN1LI; residues 651–802, broken line) chains, respectively. The motor domain (amino acids 1868–4646) is indicated by the orange shaded area; the seven ATPase domains are represented by circles, whereas the horizontal bar indicates the stalk region. The equivalent positions of mutations in three mouse models, Loa, Swl, and Cra1, are shown below the figure along with the p.His306Arg mutation identified in the family reported here. Note that the human protein contains two additional glycine residues at position 7 relative to mouse Dync1h1, that is numbering of equivalent residues in human DYNC1H1 is 2 higher than in mouse models. Phe582 is within a highly conserved domain responsible for binding the dynein intermediate chains as well as homodimerization. The 9 bp Swl deletion and Cra p.Tyr1057Cys mutation are outside the dynein intermediate-chain binding region but within the putative homodimerization domain. Numbering of amino acids is taken from Tynan et al. and the UniProtKB entry for human DYNC1H1 (identifier Q14204).