Definition of subchromosomal intervals around the myotonic dystrophy gene region at 19q

The localization to 19q of the gene causing myotonic dystrophy (DM) has been defined more precisely by refinement of the physical location of several linked markers. A somatic cell hybrid mapping panel from cells with t(1;19), t(12;19), and t(X;19) translocation products was constructed to define five different intervals across 19q. In addition, we have derived a series of cell hybrids by irradiation of a der(19)-only hybrid to further subdivide the cen-q13.1 region. Using an array of 36 cloned genes, anonymous DNAs, and enzyme markers, we have tested the location of the panel breakpoints and refined the regional assignment of several of these markers. All markers tightly linked to DM are localized mainly within 19q13.2, thus suggesting that the DM gene is also close to this region.     

Localization of a human Na, K-ATPase α subunit gene to chromosome 19q12åq13.2 and linkage to the myotonic dystrophy locus

The gene coding for a Na⁺,K⁺-ATPase α subunit (ATP1A3) has been localized to the q12åq13.2 region of human chromosome 19, potentially close to the myotonic dystrophy (DM) gene. In view of previous studies implicating a Na⁺,K⁺-ATPase in the pathology of DM, we have examined the possibility that ATP1A3 is a candidate for the DM locus. Although linked, several clear instances of recombination between ATP1A3 and DM rule out the possibility that mutations in ATP1A3 cause the disease. Examination of multiply informative pedigrees indicates the gene order DM-APOC2-ATP1A3.     

The human ryanodine receptor gene: Its mapping to 19q13.1, placement in a chromosome 19 linkage group, and exclusion as the gene causing myotonic dystrophy

The recent cloning of cDNA encoding the Ca++ release channel (ryanodine receptor) of human sarcoplasmic reticulum has enabled us to use somatic cell hybrids to localize the ryanodine receptor gene (RYR) to the proximal long arm of human chromosome 19. Studies with additional hybrids containing deletions or translocations in chromosome 19 enabled us to localize RYR to 19q13.1 in a region distal to GPI/MAG and proximal to D19S18/DNF11. On the basis that the myotonic dystrophy (DM) locus maps near this region and that myotonia could result from a defect in the ryanodine receptor, we examined the linkage between the DM locus and RYR. Our results, showing several DM-RYR recombinants, rule out an RYR defect as the cause of DM. However, localization of RYR to a region of human chromosome 19 which is syntenic to an area of pig chromosome 6 containing the HAL gene responsible for porcine malignant hyperthermia supports the candidacy of RYR for this disorder.     

Linkage analysis of the apolipoprotein C2 gene and myotonic dystrophy on human chromosome 19 reveals linkage disequilibrium in a French-Canadian population.

The gene for human apolipoprotein C2 (APOC2), situated on the proximal long arm of chromosome 19, is closely linked to the gene for the most common form of adult muscular dystrophy, myotonic dystrophy (DM). Six APOC2 RFLPs (TaqI, BglI, BanI, BamHI, NcoI, and AvaII) have been identified to date. We have conducted a comprehensive DM linkage study utilizing all six RFLPs and involving 50 families and 372 individuals. The most informative RFLPs are, in descending order, NcoI (lod = 6.64, theta = 0.05), BglI (lod = 6.12, theta = 0.05), AvaII (lod = 6.02, theta = 0.03), BanI (lod = 5.76, theta = 0.04), TaqI (lod = 4.29, theta = 0.06), and BamHI (lod = 1.75, theta = 0.01). A substantial increase in the lod scores over those seen with the individual RFLPs was obtained when the linkage of the entire APOC2 haplotype (composed of the six RFLPs) was studied (lod = 17.87, theta = 0.04). We have observed significant inter-APOC2 RFLP linkage disequilibrium. Consequently, the three most informative RFLPs have been found to be BanI, TaqI, and either BglI, AvaII, or NcoI polymorphisms. We also demonstrate linkage disequilibrium between DM and APOC2 in our French-Canadian population (standardized disequilibrium constant phi = .22, chi 2 = 5.12, df = 1, P less than 0.04). This represents the first evidence of linkage disequilibrium between APOC2 and the DM locus.     

Localisation of the myotonic dystrophy locus to 19q13.2–19q13.3 and its relationship to twelve polymorphic loci on 19q

Summary The order of fourteen polymorphic markers localised to the long arm of human chromosome 19 has been established by multipoint mapping in a set of 40 CEPH (Centre d'Étude de Polymorphisme Humain, Paris) reference families. We report here the linkage relationship of the myotonic dystrophy (DM) locus to twelve of these markers as studied in 45 families with DM. The resulting genetic map is supported by the localisation of the DNA markers in a panel of somatic cell hybrids. Ten of the twelve markers have been shown to be proximal to the DM gene and two, PRKCG and D19S22, distal but at distances of approximately 25 cM and 15 cM, respectively. The closest proximal markers are APOC2 (apolipoprotein C-II) and CKM (creatine kinase, muscle) approximately 3 cM and 2 cM from the DM gene respectively, in the order APOC2-CKM-DM. The distance between APOC2, CKM and DM (of the order of 2 million base pairs) and their known orientation should permit directional chromosome walking and jumping. The data presented here should enable us to determine whether or not new markers are distal to APOC2/CKM and thus potentially flank the DM gene.     

Localization of a human Na+,K+-ATPase alpha subunit gene to chromosome 19q12----q13.2 and linkage to the myotonic dystrophy locus

The gene coding for a Na+,K+-ATPase alpha subunit (ATP1A3) has been localized to the q12----q13.2 region of human chromosome 19, potentially close to the myotonic dystrophy (DM) gene. In view of previous studies implicating a Na+,K+-ATPase in the pathology of DM, we have examined the possibility that ATP1A3 is a candidate for the DM locus. Although linked, several clear instances of recombination between ATP1A3 and DM rule out the possibility that mutations in ATP1A3 cause the disease. Examination of multiply informative pedigrees indicates the gene order DM-APOC2-ATP1A3.     

The apolipoprotein CII gene: Subchromosomal localisation and linkage to the myotonic dystrophy locus

Summary The human apolipoprotein CII gene probe detects a restriction fragment length polymorphism located on chromosome 19. We have investigated the linkage of this polymorphism to the myotonic dystrophy locus in families. The two lici are closely linked with a maximum Lod score of 7.877 at 4% recombination. The close linkage and informativeness of the APOC2 polymorphism suggest that this probe may be of use for presymptomatic diagnosis of the myotonic dystrophy gene. The APOC2 gene was localised to the region 19p13–19q13 using somatic cell hybrids, providing further evidence that the myotonic dystrophy locus is situated in the central region of chromosome 19.     

Recombination events that locate myotonic dystrophy distal to APOC2 on 19q

We previously reported a recombination in an individual with myotonic dystrophy (DM) which placed the markers D19S19 and APOC2 on the same side of the DM locus. Haplotyping of this family with more recently characterized probes which are either tightly linked to DM or distal to the linkage group at q13.2 shows that the DM locus is distal to APOC2. This is confirmed by other recombinants where DM segregates with distal probes. Additional marker to marker recombinations in unaffected individuals are reported and support the order and orientation of the DM linkage group as pter-(INSR, LDLR,S9)-(S19,BCL3,APOC2)-(CKMM,DM)-(S22,+ ++PRKCG)-qter. The data presented here cannot determine whether DM is proximal or distal to CKMM. The consequences of this probe order for antenatal diagnosis and future research aiming to isolate the gene which is affected in DM are discussed.     

A multipoint linkage map around the locus for myotonic dystrophy on chromosome 19

Employing 16 polymorphic DNA markers as well as the chromosome 19 centromere heteromorphism, we have performed a genetic linkage study in 26 families with myotonic dystrophy. Fourteen of these markers had been assigned previously to one of five different intervals of the 19cen-19q13.2 segment by using somatic cell hybrids. For the long arm of chromosome 19, a genetic map that encompasses 9 polymorphic markers and the DM gene has been constructed. Our studies indicate that the DM and CKMM genes map distal to the ApoC2-ApoE gene cluster and to the anonymous polymorphic markers D19S15 and D19S16, but proximal to the D19S22 marker. The orientation of DM and CKMM remains to be determined.     

Identification of variable simple sequence motifs in 19q13.2-qter: markers for the myotonic dystrophy locus.

Variable simple sequence motifs (VSSMs), or microsatellites, were used for the genetic delimitation of the myotonic dystrophy (DM) region at 19q. Three simple sequence motifs were identified in and around the ERCC1 DNA-repair gene at 19q13.2-13.3 and one in the vicinity of the RRAS gene at 19q13.3-qter. A (TG)n repeat, situated within the ninth intron of the ERCC1 gene, was converted into a highly informative multiallelic marker using PCR-mediated DNA amplification and high-resolution gel analysis. The structurally similar sequence motif in the RRAS gene yielded a marker system with only two alleles. Use of these VSSMs for linkage analysis and haplotyping in a selected set of DM families revealed that the DM gene is distal but close to the ERCC1 locus and can be excluded from the CKM-ERCC1 interval at 19q13.2. The order for RRAS and other distally located markers was established as DM-D19S50-[RRAS,KLK]-D19S22-ter.     

A chromosome 19 clone from a translocation breakpoint shows close linkage and linkage disequilibrium with myotonic dystrophy

The gene for myotonic dystrophy (DM), the most common form of adult muscular dystrophy, is situated on the proximal long arm of chromosome 19. Although there exist markers that are tightly linked to the DM locus, its precise location is unknown. The identification and characterization of additional DNA probes closely linked to the DM locus continue to be priorities. In this study, we report on the linkage between a new DNA marker, designated p alpha 1.4P, and the DM locus in 50 families. The probe p alpha 1.4P was derived from a cloned breakpoint junction fragment from the chromosomal translocation t(14;19)(q32;q13.1). This translocation has been previously described in some cases of chronic lymphocytic leukemia. We have identified a BanI restriction fragment length polymorphism that is detected by p alpha 1.4P. Segregation analysis between this RFLP and DM revealed close linkage between the two loci (lod = 10.95, theta = 0). Furthermore, statistical evidence for linkage disequilibrium between p alpha 1.4P and the DM locus in a French Canadian population was found. Finally, by means of a somatic cell hybrid mapping panel, p alpha 1.4P was sublocalized to 19q12----19q13.2.     

Regional localisations and linkage relationships of seven RFLPs and myotonic dystrophy on chromosome 19

Summary We have studied the genetic linkage relationships of seven DNA polymorphisms on chromosome 19, with each other and with the myotonic dystrophy locus. The DNA sequences were localised to various regions of the chromosome using translocations in somatic cell hybrids. These results provide the basis for a linkage map of most of chromosome 19, and suggest that the myotonic dystrophy locus is close to the centromere.     

The syntenic relationship of proximal mouse chromosome 7 and the myotonic dystrophy gene region on human chromosome 19q

The syntenic relationship of the myotonic dystrophy (DM) gene region on human chromosome 19q and proximal mouse chromosome 7 was examined using an interspecific backcross between C3H/HeJ-gld/gld mice and Mus spretus. Segregation analyses were used to order homologs of nine human loci linked with the DM gene. Their order from the centromere was Prkcg, [Apoe, Atpa-2, Ckmm, D19S19h, Ercc-2], Cyp2b, Mag, Lhb. Two other murine loci, D7Rp2 and Ngfg, were also positioned within this interval. Homologs for five human chromosome 11 and 15 loci (Calc, Fes, Hras-1, Igflr, Tyr) were localized within an 18-cM span telomeric to Lhb. Comparison of the gene orders indicates an inversion extending from Prkcg through the interval between Mag and Lhb. This study establishes a detailed map of proximal mouse chromosome 7 that will be useful in identifying and determining whether new human chromosome 19 probes are linked to the DM region.     

D19S51 is closely linked with and maps distal to the myotonic dystrophy locus on 19q.

Recent genetic linkage studies have mapped the myotonic dystrophy (DM) locus to 19q13.3. All closely linked DM markers identified to date have been located on the centromeric side of the disease locus, with a relatively large genetic interval (9 cM) observed between the nearest distal marker and DM. We show here that the recently described marker p134C is tightly linked to DM (peak lod score 35.8 at peak recombination fraction .006) and confirm the previous suggestion that the p134C locus, D19S51 maps distal to the disease locus. D19S51 and the closest proximal flanking loci, ERCC1 and D19S115 (pE0.8), define a small genetic interval of less than 2 cM that contains the DM locus.     

Assignment of seven genes to distinct intervals on the midportion of human chromosome 19q surrounding the myotonic dystrophy gene region.

Hybridization studies using a panel of somatic cell hybrids with subchromosomal segments of 19q have localized the genes encoding hormone-sensitive lipase (LIPE), carcinoembryonic antigen (CEA), and small nuclear ribonucleoprotein polypeptide A (SNRPA) to various regions of 19q13.1; the cellular receptor for poliovirus sensitivity (PVS) to 19q13.2; and the genes coding for prostate-specific antigen (APS), human pancreatic kallikrein (KLK1), and small nuclear ribonucleoprotein 70-kD polypeptide (SNRP70) to 19q13.3----qter. Our results exclude several of these genes from being seriously considered as a candidate for the myotonic dystrophy gene on 19q.     

A reordering of human chromosome 19 long-arm DNA markers and identification of markers flanking the myotonic dystrophy locus

The gene for myotonic dystrophy (DM), the most common form of adult muscular dystrophy, has previously been mapped to the proximal long arm of chromosome 19. We have conducted linkage analysis on 53 DM families (comprising 421 individuals) using seven DM-linked DNA markers. This analysis, combined with our somatic cell hybrid mapping panel data, places the DM locus more distal on the chromosome 19 long arm than previously thought. Further, we have been able to unequivocally identify DNA markers that flank the disease locus. The definition of a 10-cM region of chromosome 19 that contains the DM locus should prove useful in both the search for the causative gene and the molecular diagnosis of DM.     

Physical and genetic characterization of the distal segment of the myotonic dystrophy area on 19q.

The mutation involved in myotonic dystrophy (DM) has been mapped to the region between the ERCC1 DNA repair gene and the anonymous D19S51 locus on 19q13.3. Starting at locus D19S112 (probe pX75b), which served as a novel entry site for this chromosome region, we have established a cosmid contig of approximately 200 kb. In the contig, a gene expressed in the brain and a highly informative, 12-allele (TG)n variable simple sequence motif (VSSM) were identified. With this marker, designated X75b-VSSM, a highly characteristic size distribution of alleles linked with DM, which differed significantly from that on normal chromosomes, was observed. Combining our physical mapping and genetic data, we show that the X75b-VSSM marker is the closest distal to DM, thus excluding the DM mutation from the entire telomeric portion of the ERCC1-D19S51 region.     

Establishment of the mouse chromosome 7 region with homology to the myotonic dystrophy region of human chromosome 19q

A number of genetic markers, including ATP1A3, TGFB, CKMM, and PRKCG, define the genetic region on human chromosome 19 containing the myotonic dystrophy locus. These and a number of other DNA probes have been mapped to mouse chromosome 7 utilizing a mouse Mus domesticus/Mus spretus interspecific backcross segregating for the genetic markers pink-eye dilution (p) and chinchilla (cch). The establishment of a highly syntenic group conserved between mouse chromosome 7 and human chromosome 19q indicates the likely position of the homologous gene locus to the human myotonic dystrophy gene on proximal mouse chromosome 7. In addition, we have mapped the muscle ryanodine receptor gene (Ryr) to mouse chromosome 7 and demonstrated its close linkage to the Atpa-2, Tgfb-1, and Ckmm cluster of genes. In humans, the malignant hyperthermia susceptibility locus (MHS) also maps close to this gene cluster. The comparative mapping data support Ryr as a candidate gene for MHS.     

Localization, by linkage analysis, of the cystinuria type III gene to chromosome 19q13.1.

Cystinuria is an autosomal recessive aminoaciduria in which three urinary phenotypes (I, II, and III) have been described. An amino acid transporter gene, SLC3A1 (formerly rBAT), was found to be responsible for this disorder. Mutational and linkage analysis demonstrated the presence of genetic heterogeneity in which the SLC3A1 gene is responsible for type I cystinuria but not for type II or type III. In this study, we report the identification of the cystinuria type III locus on the long arm of chromosome 19 (19q13.1), obtained after a genomewide search. Pairwise linkage analysis in a series of type III or type II families previously excluded from linkage to the cystinuria type I locus (SLC3A1 gene) revealed a significant maximum LOD score (zeta max) of 13.11 at a maximum recombination fraction (theta max) of .00, with marker D19S225. Multipoint linkage analysis performed with the use of additional markers from the region placed the cystinuria type III locus between D19S414 and D19S220. Preliminary data on type II families also seem to place the disease locus for this rare type of cystinuria at 19q13.1 (significant zeta max = 3.11 at theta max of .00, with marker D19S225).     

Mapping of facioscapulohumeral muscular dystrophy gene to chromosome 4q35-qter by multipoint linkage analysis and in situ hybridization

We have recently assigned the facioscapulohumeral muscular dystrophy (FSHD) gene to chromome 4 by linkage to the microsatellite marker Mfd 22 (locus D4S171). We now report that D4S139, a VNTR locus, is much more closely linked to FSHD. Two-point linkage analysis between FSHD and D4S139 in nine informative families showed a maximum combined lod score (Zmax) of 17.28 at a recombination fraction theta of 0.027. Multipoint linkage analysis between FSHD and the loci D4S139 and D4S171 resulted in a peak lod score of 20.21 at 2.7 cM from D4S139. Due to the small number of recombinants found with D4S139, the position of the FSHD gene relative to that of D4S139 could not be established with certainty. D4S139 was mapped to chromosome 4q35-qter by in situ hybridization, thus firmly establishing the location of the FSHD gene in the subtelomeric region of chromosome 4q. One small family yielded a negative lod score for D4S139. In the other families no significant evidence for genetic heterogeneity was obtained. Studies of additional markers and new families will improve the map of the FSHD region, reveal possible genetic heterogeneity, and allow better diagnostic reliability.