R. Mark Gardiner

SCN9A Mutations in Paroxysmal Extreme Pain Disorder: Allelic Variants Underlie Distinct Channel Defects and Phenotypes

Paroxysmal extreme pain disorder (PEPD), previously known as familial rectal pain (FRP, or OMIM 167400), is an inherited condition characterized by paroxysms of rectal, ocular, or submandibular pain with flushing. A genome-wide linkage search followed by mutational analysis of the candidate gene SCN9A, which encodes hNa(v)1.7, identified eight missense mutations in 11 families and 2 sporadic cases. Functional analysis in vitro of three of these mutant Na(v)1.7 channels revealed a reduction in fast inactivation, leading to persistent sodium current. Other mutations in SCN9A associated with more negative activation thresholds are known to cause primary erythermalgia (PE). Carbamazepine, a drug that is effective in PEPD, but not PE, showed selective block of persistent current associated with PEPD mutants, but did not affect the negative activation threshold of a PE mutant. PEPD and PE are allelic variants with distinct underlying biophysical mechanisms and represent a separate class of peripheral neuronal sodium channelopathy.

Spectrum of Mutations in the Batten Disease Gene, CLN3

Batten disease (juvenile-onset neuronal ceroid lipofuscinosis [JNCL]) is an autosomal recessive condition characterized by accumulation of lipopigments (lipofuscin and ceroid) in neurons and other cell types. The Batten disease gene, CLN3, was recently isolated, and four disease-causing mutations were identified, including a 1.02-kb deletion that is present in the majority of patients (The International Batten Disease Consortium 1995). One hundred eighty-eight unrelated patients with JNCL were screened in this study to determine how many disease chromosomes carried the 1.02-kb deletion and how many carried other mutations in CLN3. One hundred thirty-nine patients (74%) were found to have the 1.02-kb deletion on both chromosomes, whereas 49 patients (41 heterozygous for the 1.02-kb deletion) had mutations other than the 1.02-kb deletion. SSCP analysis and direct sequencing were used to screen for new mutations in these individuals. Nineteen novel mutations were found: six missense mutations, five nonsense mutations, three small deletions, three small insertions, one intronic mutation, and one splice-site mutation. This report brings the total number of disease-associated mutations in CLN3 to 23. All patients homozygous for mutations predicted to give rise to truncated proteins were found to have classical JNCL. However, a proportion of the patients (n = 4) who were compound heterozygotes for a missense mutation and the 1.02-kb deletion were found to display an atypical phenotype that was dominated by visual failure rather than by severe neurodegeneration. All missense mutations were found to affect residues conserved between the human protein and homologues in diverse species.

A model for Batten disease protein CLN3: Functional implications from homology and mutations

In an attempt to understand the molecular nature of Batten disease, we have examined the amino acid sequence of the affected CLN3 gene product (The International Batten Disease Consortium (1995) Cell 82, 949–957) and the site‐specific mutations which give rise to the biological defect. Homology searches and molecular modeling have led to the development of a model for the folding and disposition of the protein, possibly within a mitochondrial membrane. High homology with a yeast protein of unknown function suggests a strong evolutionary conservation of function. We speculate that a possible role for the protein may be in chaperoning the folding/unfolding or assembly/disassembly of other proteins, specifically subunit c of the mitochondrial ATP synthase complex.

Evaluation of the positional candidate gene CHRNA7 at the juvenile myoclonic epilepsy locus (EJM2) on chromosome 15q13-14

A previous study of 34 nuclear pedigrees segregating juvenile myoclonic epilepsy (JME) gave significant evidence of linkage with heterogeneity to marker loci on chromosome 15q13-14 close to the candidate gene CHRNA7 (Hum. Mol. Genet. 6 (1997) 1329). The aim of this work was to further evaluate the putative aetiological role of CHRNA7 in JME within the 34 families originally described, and to assess the contribution of this locus to a broader phenotype of idiopathic generalised epilepsy (IGE). Multipoint linkage analysis and intrafamilial association studies were performed with microsatellite markers that encompass both CHRNA7 and its partial duplication (CHRFAM7A). A maximum HLOD of 3.45 [alpha=0.58; (Zall=2.88, P=0.0008)] was observed 8 cM distal to D15S1360, a CHRNA7 intragenic marker. Significant exclusion lod scores were obtained across the region in 12 mixed phenotype JME/IGE families. Mutation screening of the CHRNA7 gene (and consequently exons 5-10 of CHRFAM7A) and its putative promoter sequence identified a total of 13 sequence variants across 23 of 34 JME-affected families. Two variants (c.1354G>A and c.1466C>T) are predicted to result in amino acid changes and one (IVS9+5G>A) is predicted to result in aberrant transcript splicing. However, none of the variants alone appeared either necessary or sufficient to cause JME in the families in which they occurred. In conclusion, linkage analyses continue to support the existence of a locus on chromosome 15q13-14 that confers susceptibility to JME but not to a broader IGE phenotype. Causal sequence variants in the positional candidate CHRNA7 have not been identified but the presence of multiple segmental duplications in this region raises the possibility of undetected disease-causing genomic rearrangements.

Genome-wide High-Density SNP-Based Linkage Analysis of Infantile Hypertrophic Pyloric Stenosis Identifies Loci on Chromosomes 11q14-q22 and Xq23

Infantile hypertrophic pyloric stenosis (IHPS) has an incidence of 1-8 per 1000 live births and is inherited as a complex sex-modified multifactorial trait with a striking male preponderance. Syndromic and monogenic forms exist, and two loci have been identified. Infants present with vomiting due to gastric-outlet obstruction caused by hypertrophy of the smooth muscle of the pylorus. A genome-wide SNP-based high-density linkage scan was carried out on 81 IHPS pedigrees. Nonparametric and parametric linkage analysis identified loci on chromosomes 11q14-q22 (Z(max) = 3.9, p < 0.0001; HLOD(max) = 3.4, alpha = 0.34) and Xq23 (Z(max) = 4.3, p < 0.00001; HLOD(max) = 4.8, alpha = 0.56). The two linked chromosomal regions each harbor functional candidate genes that are members of the canonical transient receptor potential (TRPC) family of ion channels and have a potential role in smooth-muscle control and hypertrophy.

Exploration of the Genetic Architecture of Idiopathic Generalized Epilepsies

Summary:  Purpose: Idiopathic generalized epilepsy (IGE) accounts for ∼20% of all epilepsies and affects about 0.2% of the general population. The etiology of IGE is genetically determined, but the complex pattern of inheritance suggests an involvement of a large number of susceptibility genes. The objective of the present study was to explore the genetic architecture of common IGE syndromes and to dissect out susceptibility loci predisposing to absence or myoclonic seizures.

Evaluation of CACNA1H in European patients with childhood absence epilepsy

CACNA1H was evaluated in a resource of Caucasian European patients with childhood absence epilepsy by linkage analysis and typing of sequence variants previously identified in Chinese patients. Linkage analysis of 44 pedigrees provided no evidence for a locus in the CACNA1H region and none of the Chinese variants were found in 220 unrelated patients.

Linkage analysis between childhood absence epilepsy and genes encoding GABAA and GABAB receptors, voltage-dependent calcium channels, and the ECA1 region on chromosome 8q

Childhood absence epilepsy (CAE) is an idiopathic generalised epilepsy (IGE) characterised by onset of typical absence seizures in otherwise normal children of school age. A genetic component to aetiology is well established but the mechanism of inheritance and the genes involved are unknown. Available evidence suggests that mutations in genes encoding GABA receptors or brain expressed voltage-dependent calcium channels (VDCCs) may underlie CAE. The aim of this work was to test this hypothesis by linkage analysis using microsatellite loci spanning theses genes in 33 nuclear families each with two or more individuals with CAE. Seventeen VDCC subunit genes, ten GABA(A)R subunit genes, two GABA(B) receptor genes and the ECA1 locus on 8q24 were investigated using 35 microsatellite loci. Assuming locus homogeneity, all loci gave statistically significant negative LOD scores, excluding these genes as major loci in the majority of these families. Positive HLOD scores assuming locus heterogeneity were observed for CACNG3 on chromosome 16p12-p13.1 and the GABRA5, GABRB3, GABRG3 cluster on chromosome 15q11-q13. Association studies are required to determine whether these loci are the site of susceptibility alleles in a subset of patients with CAE.

Genome wide high density SNP-based linkage analysis of childhood absence epilepsy identifies a susceptibility locus on chromosome 3p23-p14

Summary Childhood absence epilepsy (CAE) is an idiopathic generalised epilepsy (IGE) characterised by typical absence seizures manifested by transitory loss of awareness with 2.5–4 Hz spike-wave complexes on ictal EEG. A genetic component to the aetiology is well recognised but the mechanism of inheritance and the genes involved are yet to be fully established. A genome wide single nucleotide polymorphism (SNP)-based high density linkage scan was carried out using 41 nuclear pedigrees with at least two affected members. Multipoint parametric and non-parametric linkage analyses were performed using MERLIN 1.1.1 and a susceptibility locus was identified on chromosome 3p23-p14 (Zmean = 3.9, p < 0.0001; HLOD = 3.3, α = 0.7). The linked region harbours the functional candidate genes TRAK1 and CACNA2D2. Fine-mapping using a tagSNP approach demonstrated disease association with variants in TRAK1.

Expression of MUL, a gene encoding a novel RBCC family ring-finger protein, in human and mouse embryogenesis

We studied the expression of MUL, a gene encoding a novel member of the RING-B-Box-Coiled Coil family of zinc finger proteins that underlies the human inherited disorder, Mulibrey nanism. In early human and mouse embryogenesis MUL is expressed in dorsal root and trigeminal ganglia, liver and in epithelia of multiple tissues.