Spinal muscular atrophy with pontocerebellar hypoplasia is caused by a mutation in the VRK1 gene

Abstract

The spinal muscular atrophies (SMAs) are a genetically and clinically heterogeneous group of disorders characterized by degeneration and loss of anterior horn cells in the spinal cord, leading to muscle weakness and atrophy. Spinal muscular atrophy with pontocerebellar hypoplasia (SMA-PCH, also known as pontocerebellar hypoplasia type 1 [PCH1]) is one of the rare infantile SMA variants that include additional clinical manifestations, and its genetic basis is unknown. We used a homozygosity mapping and positional cloning approach in a consanguineous family of Ashkenazi Jewish origin and identified a nonsense mutation in the vaccinia-related kinase 1 gene (VRK1) as a cause of SMA-PCH. VRK1, one of three members of the mammalian VRK family, is a serine/threonine kinase that phosphorylates p53 and CREB and is essential for nuclear envelope formation. Its identification as a gene involved in SMA-PCH implies new roles for the VRK proteins in neuronal development and maintenance and suggests the VRK genes as candidates for related phenotypes.

Figure 1

Linkage and Homozygosity Mapping of Markers on 14q32 to SMA-PCH in a Consanguineous Family Affected individuals are shown in black. STR genotypes in the critical region are shown below each sampled individual, where numbers indicate amplicon size (bp). The disease-associated haplotype is shaded in yellow or yellow-orange. Only IV-12 is homozygous by descent for the region boxed in red.

Figure 2

Muscle Biopsy of the Proband at Age 3 Yrs Frozen hematoxylin-eosin-stained section shows bundles of larger-diameter fibers adjacent to bundles of very small fibers. Intermingled with the large fibers are a few angular fibers, singly and in small groups. There is no abundance of central nuclei, necrosis, regeneration, or other fiber structural changes. Collagen encircling atrophic fibers is observed.

Figure 3

Brain CT Scan of the Proband's Sister at Age 7 Yrs, in 1989 (A) Cerebellar vermis hypoplasia, communicating fourth ventricle and cisterna magna (arrow). (B) Cerebellar vermis hypoplasia (arrow). (C) Large cisterna magna and cerebellar hemisphere hypoplasia.

Figure 4

Homozygous VRK1 Nonsense Mutation in SMA-PCH (A) Genomic sequence of the VRK1 exon 12 region in informative family members. The proband is homozygous for a C>T transition that creates a TGA termination codon. (B) Schematic representation of the VRK1 protein, including the ATP binding site, kinase domain, and nuclear leader sequence (NLS). Amino acid sequence of the NLS is indicated below, with the R358X mutation shown in red. (C) Species conservation of amino acids in the R358X region. (D) Allele-specific expression of VRK1 in R358X carriers. VRK1 exons 8 (ex8) and 12 (ex12) were sequenced in both genomic DNA and cDNA of a carrier of the R358X mutation. cDNA was extracted from an Epstein-Barr virus (EBV)-transformed B cell line. Exon 8 contains a known polymorphism (rs2230532) that also segregates in this family, and the exon 12 R358X mutation corresponds to 1072C>T in the cDNA (NM_003384). Top: genomic DNA sequences, showing equal biallelic content in both exon 8 and exon 12 sequences. Bottom: Unequal allelic expression in cDNA, where the wild-type allele predominates over the mutant (>70%).