Analysis in a large hyperkalemic periodic paralysis pedigree supports tight linkage to a sodium channel locus.

Hyperkalemic periodic paralysis (HYPP) is an autosomal dominant muscle disease with electrophysiological abnormalities suggesting a defect in a voltage-gated sodium channel (NaCh) gene. A human NaCh gene was recently shown to cosegregate with the disease allele in a family with HYPP. Using an independent clone, we have demonstrated close genetic linkage between an NaCh gene and the HYPP locus in another family. With physiological data demonstrating abnormal NaCh function in HYPP patients, the absence of any obligate recombinations in the two families strengthens the argument that this NaCh gene is the site of the defect in this disorder.     

Linkage of hyperkalaemic periodic paralysis in Quarter horses to the horse adult skeletal muscle sodium channel gene

A genetic disease observed in certain Quarter horses is hyperkalaemic periodic paralysis (HYPP). This disease causes attacks of paralysis which can be induced by ingestion of potassium. Recent studies have shown that HYPP in humans is due to single base changes within the adult skeletal muscle sodium channel gene. A large Quarter horse pedigree segregating dominant HYPP was studied to determine if mutations of the sodium channel gene are similarly responsible for HYPP in horses. We used cross-species, PCR-mediated, cDNA cloning and sequencing of the horse adult skeletal muscle sodium channel alpha-subunit gene to identify a polymorphism, and then used this polymorphism to see if the horse sodium channel gene was genetically linked to HYPP in horses. The sodium channel gene was indeed found to be tightly linked to HYPP (LOD = 2.7, theta = 0). Our results suggest that HYPP in horses involves the same gene as the clinically similar human disease, and indicates that these are homologous disorders. The future identification of the specific sodium channel mutation causing HYPP in Quarter horses will permit the development of accurate molecular diagnostics of this condition, as has been recently shown for humans.     

Molecular genetic and genetic correlations in sodium channelopathies: Lack of founder effect and evidence for a second gene

We present a correlation of molecular genetic data (mutations) and genetic data (dinucleotide-repeat polymorphisms) for a cohort of seven hyperkalemic periodic paralysis (HyperPP) and two paramyotonia congenita (PC) families from diverse ethnic backgrounds. We found that each of three previously identified point mutations of the adult skeletal muscle sodium-channel gene occurred on two different dinucleotide-repeat haplotypes. These results indicate that dinucleotide-repeat haplotypes are not predictive of allelic heterogeneity in sodium channelopathies, contrary to previous suggestions. In addition, we identified a HyperPP pedigree in which the dominant disorder was not linked to the sodium-channel gene. Thus, a second locus can give rise to a similar clinical phenotype. Some individuals in this pedigree exhibited a base change causing the nonconservative substitution of an evolutionarily conserved amino acid. Because this change was not present in 240 normal chromosomes and was near another HyperPP mutation, it fulfilled the most commonly used criteria for being a mutation rather than a polymorphism. However, linkage studies using single-strand conformation polymorphism–derived and sequence-derived haplotypes excluded this base change as a causative mutation: these data serve as a cautionary example of potential pitfalls in the delineation of change-of-function point mutations.     

Identification of a mutation in the gene causing hyperkalemic periodic paralysis.

DNA from seven unrelated patients with hyperkalemic periodic paralysis (HYPP) was examined for mutations in the adult skeletal muscle sodium channel gene (SCN4A) known to be genetically linked to the disorder. Single-strand conformation polymorphism analysis revealed aberrant bands that were unique to three of these seven patients. All three had prominent fixed muscle weakness, while the remaining four did not. Sequencing the aberrant bands demonstrated the same C to T transition in all three unrelated patients, predicting substitution of a highly conserved threonine residue with a methionine in a membrane-spanning segment of this sodium channel protein. The observation of a distinct mutation that cosegregates with HYPP in two families and appears as a de novo mutation in a third establishes SCN4A as the HYPP gene. Furthermore, this mutation is associated with a form of HYPP in which fixed muscle weakness is seen.     

Paroxysmal dystonic choreoathetosis: tight linkage to chromosome 2q.

Paroxysmal dystonic choreoathetosis (PDC) is characterized by attacks of involuntary movements that last up to several hours and occur at rest both spontaneously and following caffeine or alcohol consumption. We analyzed a Polish-American kindred with autosomal dominant PDC and identified tight linkage between the disorder and microsatellite markers on chromosome 2q (maximum two-point LOD score 4.77; recombination fraction 0). Our results clearly establish the existence of a locus for autosomal dominant PDC on distal chromosome 2q. The fact that three other paroxysmal neurological disorders (periodic ataxia with myokymia and hypo- and hyperkalemic periodic paralysis) are due to mutation in ion-channel genes raises the possibility that PDC is also due to an ion-channel gene mutation. It is noteworthy that a cluster of sodium-channel genes is located on distal chromosome 2q, near the PDC locus. Identifying the PDC locus on chromosome 2q will facilitate discovery of the PDC gene and enable investigators to determine whether PDC is genetically homogeneous and whether other paroxysmal movement disorders are also genetically linked to the PDC locus.     

Evidence for a single pedigree source of the hyperkalemic periodic paralysis susceptibility gene in Quarter Horses

The pedigree origin of a base pair substitution in the horse muscle sodium channel gene that confers susceptibility to the muscle disease hyperkalemic periodic paralysis (HYPP) was investigated with a set of 978 Quarter Horses. The horses were chosen at random, based on a collection of blood samples taken between 1989 and 1991 to meet parentage testing requirements, primarily but not exclusively from breeding stallions. The frequency of Quarter Horses positive for the base pair substitution, all heterozygotes, was 4-4%, which corresponds to an allelic frequency of 0.02. All horses positive for the gene traced to a single previously identified stallion as first, second or third generation descendants. A higher frequency of the HYPP susceptibility trait than expected by random occurrence was found among his descendants in this study.     

Hypokalemic Periodic Paralysis and the Dihydropyridine Receptor (CACNLIA3): Genotype/Phenotype Correlations for two Predominant Mutations and Evidence for the Absence of a Founder Effect in 16 Caucasian Families

Hypokalemic periodic paralysis (hypoPP) is an autosomal dominant disorder belonging to a group of muscle diseases involving the abnormal function of ion channels. This group of muscle diseases also comprises hyperkalemic periodic paralysis and paramyotonia congenita, both sodium-channel diseases, and myotonia congenita, a chloride-channel disorder. HypoPP is characterized by acute attacks of muscle weakness concomitant with a fall in blood potasium levels. We recently localized the hypoPP locus (hypoPP1) to chromosome 1q31-32, in an interval where the α1 subunit of the dihydropyridine receptor calcium channel (CACNL1A3) also maps. Subsequently, deleterious mutations in the voltage-sensor segment S4 were found, establishing the dihydropyridine receptor CACNL1A3 as the causative gene for hypoPP. In this paper, we report the study of 16 hypoPP families of Caucasian origin. We found only two mutations—Arg528His and Arg1239His—that cosegregated with hypoPP, each in half of the families. Analysis of the clinical characteristics of both groups of families demonstrated that incomplete penetrance is a distinctive feature of the Arg528His mutation. Using dinucleotide repeats contained within or close to the dihydropyridine receptor gene, in conjunction with evidence of a de novo Arg1239His mutation, we show that a founder effect is unlikely to account for the two predominant mutations.     

Linkage data suggesting allelic heterogeneity for paramyotonia congenita and hyperkalemic periodic paralysis on chromosome 17

Summary Paramyotonia congenita (PC), an autosomal dominant non-progressive muscle disorder, is characterised by cold-induced stiffness followed by muscle weakness. The weakness is caused by a dysfunction of the sodium channel in muscle fibre. Parts of the gene coding for the -subunit of the sodium channel of the adult human skeletal muscle (SCN4A) have been localised on chromosome 17. To investigate the role of this gene in the etiology of PC, a linkage analysis in 17 well-defined families was carried out. The results (z=20.61, =0.001) show that the mutant gene responsible for the disorder is indeed tightly linked to the SCN4A gene. The mutation causing hyperkalemic periodic paralysis (HyperPP) with myotonia has previously been mapped to this gene locus by the same candidate gene approach. Thus, our data suggest that PC and HyperPP are caused by allelic mutations at a single locus on chromosome 17.Dedicated to Professor P. E. Becker on the occasion of his 83rd birthday.     

Mutations in an S4 segment of the adult skeletal muscle sodium channel cause paramyotonia congenita.

The periodic paralyses are a group of autosomal dominant muscle diseases sharing a common feature of episodic paralysis. In one form, paramyotonia congenita (PC), the paralysis usually occurs with muscle cooling. Electrophysiologic studies of muscle from PC patients have revealed temperature-dependent alterations in sodium channel (NaCh) function. This observation led to demonstration of genetic linkage of a skeletal muscle NaCh gene to a PC disease allele. We now report the use of the single-strand conformation polymorphism technique to define alleles specific to PC patients from three families. Sequencing of these alleles defined base pair changes within the same codon, which resulted in two distinct amino acid substitutions for a highly conserved arginine residue in the S4 helix of domain 4 in the adult skeletal muscle NaCh. These data establish the chromosome 17q NaCh locus as the PC gene and represent two mutations causing the distinctive, temperature-sensitive PC phenotype.     

De novo pathogenic SCN8A mutation identified by whole-genome sequencing of a family quartet affected by infantile epileptic encephalopathy and SUDEP

Individuals with severe, sporadic disorders of infantile onset represent an important class of disease for which discovery of the underlying genetic architecture is not amenable to traditional genetic analysis. Full-genome sequencing of affected individuals and their parents provides a powerful alternative strategy for gene discovery. We performed whole-genome sequencing (WGS) on a family quartet containing an affected proband and her unaffected parents and sibling. The 15-year-old female proband had a severe epileptic encephalopathy consisting of early-onset seizures, features of autism, intellectual disability, ataxia, and sudden unexplained death in epilepsy. We discovered a de novo heterozygous missense mutation (c.5302A>G [p.Asn1768Asp]) in the voltage-gated sodium-channel gene SCN8A in the proband. This mutation alters an evolutionarily conserved residue in Nav1.6, one of the most abundant sodium channels in the brain. Analysis of the biophysical properties of the mutant channel demonstrated a dramatic increase in persistent sodium current, incomplete channel inactivation, and a depolarizing shift in the voltage dependence of steady-state fast inactivation. Current-clamp analysis in hippocampal neurons transfected with p.Asn1768Asp channels revealed increased spontaneous firing, paroxysmal-depolarizing-shift-like complexes, and an increased firing frequency, consistent with a dominant gain-of-function phenotype in the heterozygous proband. This work identifies SCN8A as the fifth sodium-channel gene to be mutated in epilepsy and demonstrates the value of WGS for the identification of pathogenic mutations causing severe, sporadic neurological disorders.     

A gene for autosomal recessive limb-girdle muscular dystrophy in Manitoba Hutterites maps to chromosome region 9q31-q33: evidence for another limb-girdle muscular dystrophy locus.

Characterized by proximal muscle weakness and wasting, limb-girdle muscular dystrophies (LGMDs) are a heterogeneous group of clinical disorders. Previous reports have documented either autosomal dominant or autosomal recessive modes of inheritance, with genetic linkage studies providing evidence for the existence of at least 12 distinct loci. Gene products have been identified for five genes responsible for autosomal recessive forms of the disorder. We performed a genome scan using pooled DNA from a large Hutterite kindred in which the affected members display a mild form of autosomal recessive LGMD. A total of 200 markers were used to screen pools of DNA from patients and their siblings. Linkage between the LGMD locus and D9S302 (maximum LOD score 5.99 at recombination fraction .03) was established. Since this marker resides within the chromosomal region known to harbor the gene causing Fukuyama congenital muscular dystrophy (FCMD), we expanded our investigations, to include additional markers in chromosome region 9q31-q34.1. Haplotype analysis revealed five recombinations that place the LGMD locus distal to the FCMD locus. The LGMD locus maps close to D9S934 (maximum multipoint LOD score 7.61) in a region that is estimated to be approximately 4.4 Mb (Genetic Location Database composite map). On the basis of an inferred ancestral recombination, the gene may lie in a 300-kb region between D9S302 and D9S934. Our results provide compelling evidence that yet another gene is involved in LGMD; we suggest that it be named "LGMD2H."     

脂肪分化相关蛋白基因多态性与优质鸡屠宰性状和肌内脂肪含量的相关性研究

对脂肪分化相关蛋白(Adipocyte Differentiation-Related Protein, ADFP)基因的外显子进行SNPs 检测, 探讨其作为鸡脂肪性状候选基因的可能性。实验以四川省畜牧科学研究院和大恒家禽育种有限公司培育的优质肉鸡新品系为素材, 采用PCR-SSCP的方法进行SNPs 检测和基因型的分析。结果找到3个SNPs位点: 4 079位由A→T(位点A)、4 843位由C→T(位点B)和7 070位由A→G(位点C)。单位点基因型对屠宰性状的遗传效应分析表明, 位点A的基因型对腿肌率、腹脂重、腹脂率和肌内脂肪含量有显著性影响(P < 0.05), 位点B的基因型对活重和屠体重均有显著性影响(P < 0.05), 位点C的基因型对胸肌重和肌内脂肪含量有显著性影响(P < 0.05), 对胸肌率有极显著性影响(P < 0.01)。初步推断ADFP基因可能是影响鸡脂肪性状的主效基因或与主效基因连锁, 推测可以利用多态位点A和C对鸡腹脂重、腹脂率和肌内脂肪含量进行标记辅助选择。     

Clinical and genetic heterogeneity in laminopathies

Mutations in the LMNA gene encoding lamins A/C are responsible for more than ten different disorders called laminopathies which affect various tissues in an isolated (striated muscle, adipose tissue or peripheral nerve) or systemic (premature aging syndromes) fashion. Overlapping phenotypes are also observed. Associated with this wide clinical variability, there is also a large genetic heterogeneity, with 408 different mutations being reported to date. Whereas a few hotspot mutations emerge for some types of laminopathies, relationships between genotypes and phenotypes remain poor for laminopathies affecting the striated muscles. In addition, there is important intrafamilial variability, explained only in a few cases by digenism, thus suggesting an additional contribution from modifier genes. In this regard, a chromosomal region linked to the variability in the age at onset of myopathic symptoms in striated muscle laminopathies has recently been identified. This locus is currently under investigation to identify modifier variants responsible for this variability.     

EphB2 SNPs and sporadic prostate cancer risk in African American men

The EphB2 gene has been implicated as a tumor suppressor gene somatically altered in both prostate cancer (PC) and colorectal cancer. We have previously shown an association between an EphB2 germline nonsense variant and risk of familial prostate cancer among African American Men (AAM). Here we set out to test the hypothesis that common variation within the EphB2 locus is associated with increased risk of sporadic PC in AAM. We genotyped a set of 341 single nucleotide polymorphisms (SNPs) encompassing the EphB2 locus, including known and novel coding and noncoding variants, in 490 AA sporadic PC cases and 567 matched controls. Single marker-based logistical regression analyses revealed seven EphB2 SNPs showing statistically significant association with prostate cancer risk in our population. The most significant association was achieved for a novel synonymous coding SNP, TGen-624, (Odds Ratio (OR) =?0.22; 95% Confidence Interval (CI) 0.08-0.66, p?=?1×10(-5)). Two other SNPs also show significant associations toward a protective effect rs10465543 and rs12090415 (p?=?1×10(-4)), OR?=?0.49 and 0.7, respectively. Two additional SNPs revealed trends towards an increase in risk of prostate cancer, rs4612601 and rs4263970 (p?=?0.001), OR?=?1.35 and 1.31, respectively. Furthermore, haplotype analysis revealed low levels of linkage disequilibrium within the region, with two blocks being associated with prostate cancer risk among our population. These data suggest that genetic variation at the EphB2 locus may increase risk of sporadic PC among AAM.     

Neuronal sodium-channel alpha1-subunit mutations in generalized epilepsy with febrile seizures plus

Generalized epilepsy with febrile seizures plus (GEFS+) is a familial epilepsy syndrome characterized by the presence of febrile and afebrile seizures. The first gene, GEFS1, was mapped to chromosome 19q and was identified as the sodium-channel beta1-subunit, SCN1B. A second locus on chromosome 2q, GEFS2, was recently identified as the sodium-channel alpha1-subunit, SCN1A. Single-stranded conformation analysis (SSCA) of SCN1A was performed in 53 unrelated index cases to estimate the frequency of mutations in patients with GEFS+. No mutations were found in 17 isolated cases of GEFS+. Three novel SCN1A mutations-D188V, V1353L, and I1656M-were found in 36 familial cases; of the remaining 33 families, 3 had mutations in SCN1B. On the basis of SSCA, the combined frequency of SCN1A and SCN1B mutations in familial cases of GEFS+ was found to be 17%.     

Paramyotonia congenita mutations reveal different roles for segments S3 and S4 of domain D4 in hSkM1 sodium channel gating

Mutations in the gene encoding the voltage-gated sodium channel of skeletal muscle (SkMl) have been identified in a group of autosomal dominant diseases, characterized by abnormalities of the sarcolemmal excitability, that include paramyotonia congenita (PC) and hyperkalemic periodic paralysis (HYPP). We previously reported that PC mutations cause in common a slowing of inactivation in the human SkMl sodium channel. In this investigation, we examined the molecular mechanisms responsible for the effects of L1433R, located in D4/S3, on channel gating by creating a series of additional mutations at the 1433 site. Unlike the R1448C mutation, found in D4/S4, which produces its effects largely due to the loss of the positive charge, change of the hydropathy of the side chain rather than charge is the primary factor mediating the effects of L1433R. These two mutations also differ in their effects on recovery from inactivation, conditioned inactivation, and steady state inactivation of the hSkMl channels. We constructed a double mutation containing both L1433R and R1448C. The double mutation closely resembled R1448C with respect to alterations in the kinetics of inactivation during depolarization and voltage dependence, but was indistinguishable from L1433R in the kinetics of recovery from inactivation and steady state inactivation. No additive effects were seen, suggesting that these two segments interact during gating. In addition, we found that these mutations have different effects on the delay of recovery from inactivation and the kinetics of the tail currents, raising a question whether this delay is a reflection of the deactivation process. These results suggest that the S3 and S4 segments play distinct roles in different processes of hSkM1 channel gating: D4/S4 is critical for the deactivation and inactivation of the open channel while D4/S3 has a dominant role in the recovery of inactivated channels. However, these two segments interact during the entry to, and exit from, inactivation states.     

Correlation analysis of relationships between polymorphisms of high quality chicken myogenin gene and slaughter and meat quality traits]

PCR-SSCP technique was designed in this study to investigate the effect of MyoG on quality meat chicken (developed by Sichuan Dahen Poultry Breeding Company using local breeds). Four mutations at base position in promoter were detected among individuals in each line, i.e. T/C in locusA and T/A, T/C and A/G in locus B . The least square analysis showed that there were a significant difference between genotypes and breast muscle percentage and some carcass traits (P<0.05for locus A. There were a significant difference (P <0.05) in breast muscle weight between ACAA and AB geno- types; a significant difference (P <0.05) in leg muscle percentage between CC and AC for locus B, and a extremity signifi-cant difference (P <0.01) in the frequency of genotype muscle Fibre Density for Bothlocus A to locus B. There was no sig-nificant difference (P >0.05) in the other triats. It was concluded from the results that MyoG gene is the major gene affect-ing the muscle fiber traits of chicken or it links with the candidate gene, and the mutation could be used as the molecular genetic marker to select the chickens for Meat Quality traits.