Genetic mapping of the {beta}1- and {gamma}-subunits of the human skeletal muscle L-type voltage-dependent calcium channel on chromosome 17q and exclusion as candidate genes for malignant hyperthermia susceptibility

Malignant hyperthermia susceptibility (MHS) is an autosomaldominant disorder of skeletal muscle which manifests as a life-threateninghypermetabolic crisis triggered by commonly-used inhalationanaesthetics and depolarizing muscle relaxants. Defects in theryanodine receptor (RYR1) protein have been proposed to underlyMHS, but significant genetic heterogeneity in MHS has recentlybeen demonstrated. In order to investigate the potential rolesplayed by other skeletal muscle calcium channels in MHS, weisolated cosmids containing the gene encoding the ß₁subunitof skeletal muscle L-type voltage-dependent calcium channel(CACNLB1). We identified a new, highly polymorphic dinucleotiderepeat motif close to this gene, and linkage analysis placedthe marker proximal to the HOX2B locus, previously localizedto chromosome segment 17q21–q22. We recently identifieda novel marker within the     

Localization of the gene encoding the {alpha}2/{delta}-subunits of the L-type voltage-dependent calcium channel to chromosome 7q and analysis of the segregation of flanking markers in malignant hyperthermia susceptible families

Malignant hyperthermia susceptibility (MHS) is an autosomaldominant disorder of skeletal muscle which manifests as a potentiallyfatal hypermetabolic crisis triggered by commonly used anaestheticagents. This demonstration of genetic heterogeneity in MHS promptedthe investigation of the roles played by calcium regulatoryproteins other than the ryanodine receptor (RYR1), which isknown to be linked to MHS in fewer than half of the EuropeanMHS families studies to date. Previously, we have excluded thegenes encoding the skeletal muscle L-type voltage-dependentcalcium channel     

The oculopharyngeal muscular dystrophy locus maps to the region of the cardiac {alpha} and {beta} myosin heavy chain genes on chromosome 14q11.2-q13

Oculopharyngeal muscular dystrophy (OPMD) is a late-onset autosomaldominant muscular dystrophy which presents typically after theage of 50 with progressive eyelid drooping and an increasingdifficulty in swallowing. Though OPMD has a world-wide incidence,it is more common in the French Canadian population. We haveidentified a homogeneous group of families and studied 166 polymorphicmarkers as part of a genome search before establishing linkageto chromosome 14. We determined that the OPMD locus maps toa less than 5 cM region of chromosome 14q11.2–q13. Themaximum two—point lod score in three French Canadian familiesof 14.73 (     

Detection of polymorphisms in the estradiol 17{beta}-hydroxysteroid dehydrogenase II gene at the EDH17B2 locus on 17q11-q21

The human estradiol 17ß-hydroxysteroid dehydrogenaseII (17ß-HSD II) gene has been assigned by somaticcell hybridization to chromosome 17q11–q21, near the regionof assignment of the gene BRCA1, which is involved in hereditarybreast-ovarian cancer. The nucleotide sequence of 17ß-HSDII was completely determined in four unrelated individuals.Direct sequencing of PCR fragments that span the complete 17ß-HSDII gene revealed a total of 11 allelic variants which were dueto single base substitutions. The presence of these variantswas then studied in twenty six additional unrelated individuals.There were nine frequent and two rare polymophisms. Seven ofthe 11 polymorphisms were in complete linkage disequilibrium.These polymorphisms in the 17ß-HSD II gene providemarkers that can be used for the genetic mapping of this locus,and may be used to establish whether 17ß-HSD II isa candidate gene for hereditary breast-ovarian cancer.     

Pregnancy: Cervical ripening with the cytokines interleukin 8, interleukin 1{beta} and tumour necrosis factor {alpha} in guinea-pigs

It has been suggested that the collagenolytic enzymes releasedfrom white blood cells which infiltrate the pregnant human uterinecervix at term are responsible for connective tissue changeswhich take place during the ripening process. Similarly, aninfiltration of inflammatory cells occurs in pregnant guinea-pigseither spontaneously at term or at preterm after treatment withthe antiprogestin onapristone. The objective of this study wasto evaluate the effects of the inflammatory cytokines interleukin8 (IL-8), interleukin 1 (IL-1), tumour necrosis factor (TNF-)and a combination of IL-1 and TNF- on cervical ripening in guinea-pigsduring advanced pregnancy. The cytokines were applied locally(intracervically) in a gel for 2 days and the effects were assessedon the third day by both extensibility measurements and morphologicalevaluation. IL-8 treatment on days 42 and 43 post coitum (p.c)and on days 48 and 49 p.c. (term: day 67± 3 p.c.) significantly(P < 0.05) increased cervical extensibility at both stagesof pregnancy. Although IL-1 treatment (days 42 and 43 p.c.)led to a slight increase in cervical extensibility, this effectwas not statistically significant. An electron microscope studyperformed on days 48 and 49 p.c. revealed a pronounced cervicalripening accompanied by the dissolution of collagen fibres,stromal oedema and the infiltration of polymorphonuclear leukocytesin all cytokine-treated groups. The morphological effects ofIL-8 and IL-1 were indistinguishable from those observed duringnormal cervical ripening at term. In contrast, TNF-, both aloneand in combination with IL-1, brought about a severe inflammatoryreaction with a massive infiltration of lymphocytes, marcophagesand polymorphonuclear leukocytes at the investigated dose. Weconclude that the local application of the inflammatory cytokinesIL-8, IL-1 and TNF- produces cervical ripening without inducinglabour in pregnant guinea-pigs; the morphological effects ofIL-8 and IL-1 being similar to the physiological cervical ripening.Our data support the view that cytokines, particularly IL-8,may play an important role during physiological, pathologicaland induced cervical ripening and could be clinically usefulas an adjunct to labour and delivery.     

Evidence for a calcium regulated, bidirectional intronic promoter in the murine TCR V{alpha}1 gene

Previous studies of the TCR chain gene have located promoterelements 5' to the start of the various V genes. The only fullycharacterized enhancer for the entire chain gene (V, J andC genes) has been located {small tilde}3 kb from the 3' endof C. We now report the existence of additional regulatory elementslocated in the introns of several murine V genes (V1, V3 andVB6.2.16). In the case of V1, this element appears to be a promoterwith bidirectional activity that is not T cell specific. Interestingly,upstream of the promoter in the antisense strand, an open readingframe has been found that codes for a small molecular weightprotein ({small tilde}60 amino acids) that contains a prollne-richregion and a tyrosine-isoleucine motif that has homology toIgß (the B29 gene product). A rabbit antiserum madeagainst this sequence has confirmed its existence by Westernblot and immunoprecipitation. Thus this V1 intronic promoterhas the potential not only to induce the formation of a truncatedV1 gene product, but also regulates the expression of a smallmolecular weight protein that may be involved in lymphocyteantigen receptor signaling. The activity of this promoter isregulated by changes in intracellular calcium. In the presenceof ionomycin the promoter is down-regulated in the sense directionand its activity is enhanced in the antisense direction. Thisresult suggests that this promoter can act differentially toproduce two very different gene products. The bidirectionalV1 promoter appears to be the first in the Ig superfamily toinduce potentially functional proteins in both directions.     

The major peripheral myelin protein zero gene: structure and localization in the cluster of Fc{gamma} receptor genes on human chromosome 1q21.3 - q23

We have characterized the human gene encoding the major peripheralmyelin protein zero (PO) and assigned it, by In situ hybridization,to the q21.3–q23 region of human chromosome 1. This regionis known to contain a cluster of Interspersed genes coding forthe related human leukocyte receptors of the Fc portion of theImmunoglobulin G (Fc     

Binding of peptides to HLA-DQ molecules: peptide binding properties of the disease-associated HLA-DQ({alpha}1*0501, {beta}1*0201) molecule

Peptide binding to DQ molecules has not previously been described.Here we report a biochemical peptlde-blndlng assay specificfor the BQ2 [I.e. DQ(1^*0501, ß1^*0201)] molecule. Thismolecule was chosen since It shows a strong association to diseasessuch as celiac disease and insulin-dependent diabetes mellitus.Initially we radlolabelled some selected peptides and testedthem for binding to affinity-purified DQ2 molecules. One ofthe peptides, a Mycobacterium bovis (MB) 65 kDa 243–255Ypeptide, displayed a good slgnal-to-noise ratio and was thuschosen as an indicator peptide in the DQ2 binding assay. TheMB 65 kDa 243–255Y peptide bound to DQ2 In a strictlypH-dependent fashion, with optimal binding around pH 5 and onlyweak binding at pH 7.4. The association of the MB 65 kDa 243–255Ypeptide to DQ2 was slow, but once formed, the peptide-HLA complexeswere very stable. The binding of peptides to DQ2 was specific,as shown in Inhibition experiments with a panel of 47 peptides,differing in length, sequence, and origin. The binding of peptidesto DR3 was tested in a similar assay with a Mycobacterium tuberculosis65 kDa 3–13 peptide as the binding indicator. DQ2 andDR3 molecules bound to different sets of peptides. However,the peptide binding to DQ2 and DR3 showed, In general, similarcharacteristics with respect to pH dependence and kinetic parameters,Indicating that the overall rules for peptide binding to DQmolecules are the same as those previously shown for human DRand murlne I-A and I-E molecules.     

Identification of the Arg1086His mutation in the alpha subunit of the voltage-dependent calcium channel (CACNA1S) in a North American family with malignant hyperthermia

Individuals from a large North American population were screened for the presence of the mutation in the alpha1 subunit of the voltage-dependent calcium channel (CACNA1S) that has recently been associated with malignant hyperthermia (MH). This Arg1086His mutation was screened for in 154 MH normal (MHN) individuals and 112 MH susceptible (MHS) individuals, who were diagnosed by the North American protocol of the in vitro contracture test. PCR and restriction enzyme analysis was used to test for the mutation. The Arg1086His mutation in the CACNA1S was not found in any of the MHN individuals. In contrast, two related individuals (grandfather and grandson, father and son of the MH proband) among the MHS group exhibited this mutation. However, a third MHS individual in the same family (granddaughter, cousin of the grandson) did not exhibit this mutation. These results indicate that this mutation may be associated with MH in this family. Genetic alterations in the CACNA1S associated with MH are present in approximately 1% of this North American MHS population.     

Screening of SNPs at 18 positional candidate genes, located within the GD-1 locus on chromosome 14q23-q32, for susceptibility to Graves' disease: a TDT study

Graves' disease (GD) is a complex autoimmune thyroid disorder with a strong genetic component. Genome-wide screens resolved several susceptibility loci that contribute to the development of GD. One of the susceptibility loci (GD-1 locus) was mapped on chromosome 14q31. However, a susceptibility gene located within the GD-1 locus remains undefined. Here we screen eighteen single nucleotide polymorphisms (SNPs), each is situated at a corresponding positional candidate gene, located within the GD-1 susceptibility locus on chromosome 14q23-q32, for predisposition to GD using the transmission disequilibrium test in 126 simplex Russian families affected with GD. Among SNPs tested, a significant preferential transmission of the Ala allele (41 transmissions vs. 17 nontransmissions, corrected P=0.031) of the Thr92Ala SNP within the DIO2 gene, encoding type II iodothyronine deiodinase, from parents to affected children was found in a Russian family data set. The Thr92Ala SNP of the DIO2 gene and the D727E substitution of the thyrotropin receptor (TSHR) gene have been found to be in pair-wise linkage disequilibrium. The A92/E727 haplotype showed significant preferential transmission from parents to affected sibling (17 transmissions vs. 8 nontransmissions, P=0.039) in simplex families. This suggests that the Thr92Ala variant of the DIO2 gene is associated or may be in linkage disequilibrium with a functional DIO2 polymorphism which involves in the development of GD in a Russian population.