N-myc downstream-regulated gene 1 is mutated in hereditary motor and sensory neuropathy-Lom

Abstract

Hereditary motor and sensory neuropathies, to which Charcot-Marie-Tooth (CMT) disease belongs, are a common cause of disability in adulthood. Growing awareness that axonal loss, rather than demyelination per se, is responsible for the neurological deficit in demyelinating CMT disease has focused research on the mechanisms of early development, cell differentiation, and cell-cell interactions in the peripheral nervous system. Autosomal recessive peripheral neuropathies are relatively rare but are clinically more severe than autosomal dominant forms of CMT, and understanding their molecular basis may provide a new perspective on these mechanisms. Here we report the identification of the gene responsible for hereditary motor and sensory neuropathy-Lom (HMSNL). HMSNL shows features of Schwann-cell dysfunction and a concomitant early axonal involvement, suggesting that impaired axon-glia interactions play a major role in its pathogenesis. The gene was previously mapped to 8q24.3, where conserved disease haplotypes suggested genetic homogeneity and a single founder mutation. We have reduced the HMSNL interval to 200 kb and have characterized it by means of large-scale genomic sequencing. Sequence analysis of two genes located in the critical region identified the founder HMSNL mutation: a premature-termination codon at position 148 of the N-myc downstream-regulated gene 1 (NDRG1). NDRG1 is ubiquitously expressed and has been proposed to play a role in growth arrest and cell differentiation, possibly as a signaling protein shuttling between the cytoplasm and the nucleus. We have studied expression in peripheral nerve and have detected particularly high levels in the Schwann cell. Taken together, these findings point to NDRG1 having a role in the peripheral nervous system, possibly in the Schwann-cell signaling necessary for axonal survival.

Figure 1

a, Diagram of the BAC, PAC, and cosmid (HCT) clone contig of the HMSNL region. Known genes are represented by arrows pointing in the direction of their transcription. b, Positions of polymorphic microsatellites used to construct haplotypes on disease chromosomes. The HMSNL critical interval is flanked by markers pJ10 and 369CA3 and contains the WISP1 and NDRG1 genes.

Figure 2

Scale diagram of the genomic and cDNA organization of NDRG1, with an illustration of the HMSNL mutation. a, Genomic organization. The NDRG1 gene consists of 16 exons spanning 60 kb of genomic sequence. A CpG island overlaps with the first exon and the 5′ end of intron 1. A potential promoter with a TATA box, a GC box similar to the N-myc binding region of mouse Ndr1, and a TATA-less enhancer were located 39 bp, 65 bp, and 81 bp upstream, respectively, of the first exon. The 5′ UTR is split between the first two exons. b, cDNA structure. Translation starts from nucleotide position 123. c, HMSNL mutation: C→T substitution at base 564 results in a stop signal (TGA) at codon 148 in exon 7 (R148X). All nucleotide positions are given relative to the published sequence (GenBank accession number D87953).

Figure 3

RT-PCR using specific NDRG1 primers in hNT2 cells that are not differentiated (lane 1), hNT2 cells induced to neuronal differentiation (lane 2), in vitro cultured nonmyelinating Schwann cells (lane 3), total adult peripheral nerve (lane 4), and fetal brain (lane 5). NDRG1 specificity was confirmed by transfer of the RT-PCR products to a membrane and by back-hybridization with the PCR product.