Linkage analysis of neurofibromatosis type I, using chromosome 17 DNA markers.

The gene for von Recklinghausen neurofibromatosis type 1 (NF1) has recently been mapped to the pericentromeric region of human chromosome 17. To further localize the NF1 gene, linkage analysis using chromosome 17 DNA markers was performed on 11 multigeneration families with 175 individuals, 57 of whom were affected. The markers used were D17Z1 (p17H8), D17S58 (EW301), D17S54 (EW203), D17S57 (EW206), D17S73 (EW207), CRI-L946, HOX-2, and growth hormone. Tight linkage was found between NF1 and D17Z1, D17S58, and D17S57 with a recombination fraction of zero. One recombinant was detected between NF1 and D17S73, showing linkage with a 10% recombination fraction. No linkage was detected between NF1 and CRI-L946 or between HOX-2 and growth hormone. Our data are consistent with the proposed gene order pter D17S58-D17Z1-NF1-D17S57-D17S73 qter.     

Close flanking markers for neurofibromatosis type I (NF1).

A genetic linkage study with 16 polymorphic DNA markers spanning the region 17p11-17q24 in 22 NF1 families is presented. Close linkage between NF1 and eight pericentromeric markers (HHH202, EW206, CRI-L946, EW203, EW301, FG2, p17H8, and CRI-L581) has been found, probe HHH202 being the closest marker to NF1. Genetic heterogeneity has been excluded. The study of multiply informative meioses suggests that the probes HHH202 and RW206 are flanking markers for NF1. The most likely order on the basis of multiply informative meioses and multipoint mapping is pter-pA10.41-EW301-cen-HHH202-NF1-EW206-++ +EW207-qter.     

A refined genetic map of the region of chromosome 17 surrounding the von recklinghausen neurofibromatosis (NF1) gene

The von Recklinghausen neurofibromatosis (NF1) gene has been mapped to the pericentromeric region of chromosome 17. We conducted linkage analyses of NF1 by using 10 polymorphic DNA markers from this chromosomal region. We ascertained 20 American Caucasian NF1 families (163 individuals, 98 NF1 affected) in Michigan and Ohio and also studied a large family ascertained primarily in North Carolina. The following markers were used in this study: HHH202, TH17.19, D17Z1, ERBA1, EW203, EW206, EW207, EW301, CRI-L581, and CRI-L946. NF1 did not recombine with either TH17.19 or HHH202 in any of the informative meioses surveyed (maximum lod scores of 17.04 and 7.21, respectively, at a recombination fraction of .00), indicating that these markers map very close to the NF1 gene. We also report evidence of three instances of recombination between NF1 and the centromeric marker D17Z1 (maximum lod score of 13.43 at a recombination fraction of .04), as well as two crossovers between pairs of marker loci. We find no evidence of locus heterogeneity, and our results support the localization of the NF1 gene to proximal chromosome 17q.     

Genetic linkage studies of chromosome 17 RFLPs in von Recklinghausen neurofibromatosis (NF1)

Recent localization of the gene for von Recklinghausen neurofibromatosis (NF1) to chromosome 17 has led to studies to identify additional tightly linked probes that can be used in defining the primary genetic defect in NF1. We have examined and obtained blood for DNA linkage studies on over 250 individuals from 10 multigeneration neurofibromatosis families. We have analyzed 130 members in 7 families with the available chromosome 17 NF1 linked probes, pE51, D17S71, and D17Z1, as well as two probes generated from our own chromosome 17/19 enriched library (LDR92, LDR152A). Tight linkage was found between NF1 and the centromeric probe D17Z1 (theta = 0.04) and between NF1 and D17S71 (theta = 0.08). A definite recombinant was seen for the D17Z1 marker, which previously had not exhibited crossingover. Chromosome 17 DNA markers pE51, LDR92, and LDR152A gave slightly positive scores, which were not statistically significant.     

Characterization of a translocation within the von Recklinghausen neurofibromatosis region of chromosome 17

The genetic defect causing von Recklinghausen neurofibromatosis (NF1) has been mapped to the proximal long arm of chromosome 17 by linkage analysis. Flanking markers have been identified, bracketing NF1 in 17q11.2 and laying the foundation for isolating the disease gene. Recently, a family in which a mother and her two children show both the symptoms of NF1 and the presence of a balanced translocation, t(1;17)(p34.3;q11.2), has been identified. We have examined the possibility that the translocation has occurred in or near the NF1 gene by constructing a somatic cell hybrid line containing the derivative chromosome 1 (1qter-p34.3::17q11-qter). On chromosome 1, the breakpoint occurred between SRC2 and D1S57, which are separated by 14 cM. The translocation breakpoint was localized on chromosome 17 between D17S33 and D17S57, markers that also flank NF1 within a region of 4 cM. These data are consistent with the possibility that the translocation event is the cause of NF1 in this pedigree. Consequently, the isolation of the translocation breakpoint, by approach from either the chromosome 1 or the chromosome 17 side, may facilitate the identification of the NF1 gene.     

The gene for von Recklinghausen neurofibromatosis (NF1) maps to the pericentromeric region of chromosome 17 in Chinese families

Linkage analysis of six Chinese families with neurofibromatosis type 1 (NF1) confirms the location of the NF1 gene to the region of the proximal long arm of chromosome 17, as in Caucasian populations. The diagnosis of NF1 was made according to internationally accepted criteria. The markers used were D17S71, D17S58, D17S33, and EVI2A. The overall odds in favor of NF1 lying within this linkage group in the families studied are over 150,000:1, with a maximum location score of 5.112 for the interval D17S58-EVI2A.     

The order of loci in the pericentric region of chromosome 17, based on evidence from physical and genetic breakpoints.

Previous genetic analyses of chromosome 17 markers and NF1 (Fain et al. 1987) were extended in an attempt to order marker loci that map physically to 17cen----17q12. Three additional markers (HHH202, CRI-L581, and CRI-L946) were included in the analyses. Recombinants within the cluster of seven unordered marker loci were identified by pairwise analyses for each family and by examining the within-sibship segregation patterns for different markers. Changes in the segregation pattern for different loci define genetic breakpoints. Given that interference is complete in the region, markers with the same segregation pattern lie on one side of the breakpoint, while markers with different segregation patterns lie on opposite sides of the breakpoint. If the order of boundary markers is known, markers on each side of a breakpoint can be oriented in relation to the centromere. The order cen-(HHH202/NF1)-(EW207)-(EW203/CRI-L581)- (CRI-L946)-(HOX-2/NGFR)-qter was inferred by combining information from physical breakpoints in a panel of mouse/human hybrids and information from genetic breakpoints found in 16 NF1 families.     

Neurofibromatosis type 2 appears to be a genetically homogeneous disease

Neurofibromatosis type 2 (NF2) is an autosomal dominant syndrome characterized by the development of vestibular schwannomas and other tumors of the nervous system, including cranial and spinal meningiomas, schwannomas, and ependymomas. The presence of bilateral vestibular schwannomas is sufficient for the diagnosis. Skin manifestations are less common than in neurofibromatosis type 1 (NF1; von Recklinghausen disease). The apparent clinical distinction between NF1 and NF2 has been confirmed at the level of the gene locus by linkage studies; the gene for NF1 maps to chromosome 17, whereas the gene for NF2 has been assigned (in a single family) to chromosome 22. To increase the precision of the genetic mapping of NF2 and to determine whether additional susceptibility loci exist, we have performed linkage analysis on 12 families with NF2 by using four polymorphic markers from chromosome 22 and a marker at the NF1 locus on chromosome 17. Our results confirm the assignment of the gene for NF2 to chromosome 22 and do not support the hypothesis of genetic heterogeneity. We believe that chromosome 22 markers can now be used for presymptomatic diagnosis in selected families. The NF2 gene is tightly linked to the D22S32 locus (maximum lod score 4.12; recombination fraction 0). A CA-repeat polymorphism at the CRYB2 locus was the most informative marker in our families (lod score 5.99), but because the observed recombination fraction between NF2 and CRYB2 was 10 cM, predictions using this marker will need to be interpreted with caution.     

The gene for a novel epidermal antigen maps near the neurofibromatosis 1 gene.

Recently the M17S1 gene, encoding an epidermal antigen thought to play a role in cell adhesion, was mapped to chromosome bands 17q11-q12, placing it in the vicinity of the gene for the genetic disorder neurofibromatosis 1 (NF1). The pleomorphic cutaneous lesions of NF1 and the precedent for other genes being embedded within the NF1 gene prompted us to investigate whether the M17S1 gene mapped near, or within, the NF1 gene. Genetic linkage analyses revealed that M17S1 was tightly linked to NF1 and mapped within the interval bounded by D17S58 and D17S54. Physical mapping of an M17S1 cDNA on somatic cell hybrids, yeast artificial chromosomes, and an NF1 patient with a deletion involving an entire NF1 allele demonstrated that M17S1 is located at least 180 kb centromeric to the NF1 gene. The distance between the genes suggests that M17S1 is unlikely to contribute to the NF1 phenotype since a gross chromosomal rearrangement would be required to disrupt expression of both genes.     

Tightly linked markers for the neurofibromatosis type 1 gene

Relationships among genetic markers in the region of the neurofibromatosis type 1 (NF1) gene on chromosome 17 were investigated by linkage studies in a large sample set of affected families and in a panel of 58 normal families. A new marker, pHHH202 (D17S33), was included along with two markers known to be closely linked to NF. The maximum likelihood estimate of the recombination rate between the pHHH202 and NF1 loci was found to be O. Multilocus analysis suggested the following marker order: pA10-41-(p3-6, pHHH202); the NF1 gene fell with equal likelihood between either pA10-41-p3-6 or p3-6-pHHH202. The odds against NF1 being outside this cluster of tightly linked markers were greater than 15:1.     

Genetic linkage mapping of chromosome 17 markers and neurofibromatosis type 1

The von Recklinghausen neurofibromatosis (NF1) gene has been localized to the pericentromeric region of chromosome 17. We have screened six multigenerational families with multiple, tightly linked markers to aid in mapping this region of the chromosome. More than 150 members in six families were typed with probes including HHH202, D17Z1, EW203, EW206, EW207, EW301, pA10-41, D17S37, and D17S36. Two-point lod scores for NF1 versus all markers were calculated. HHH202 demonstrated the tightest linkage to NF1 with theta = .0, z = 3.86 (95% confidence limits [CL] of theta = .0-.13), suggesting that HHH202 be considered as a potential candidate marker for use in carrier detection and prenatal diagnosis. Pairwise marker-to-marker lod scores were used in examining the most likely order of subsets of the markers. Of those tested, the most likely order was (pter)-pA10-41-EW301-D17Z1-HHH202-NF1-E W206-EW207-EW203-(qter). In addition, we have ascertained an NF1 x NF1 half-cousin mating in which there are four affected family members who are potentially homozygous for the disease gene. Two of these four individuals have been sampled and typed for marker loci. When their D17Z1 genotypes are considered, the probability that both these individuals are heterozygous is 85%.     

Confirmation of the localization of the human GABAA receptor alpha 1-subunit gene (GABRA1) to distal 5q by linkage analysis.

The GABAA receptor is the major inhibitory neurotransmitter receptor in the mammalian brain. To date, 14 genes that encode subunits of this receptor have been identified; these appear to be scattered throughout the human genome and are under investigation as candidate loci for a number of neurological and psychiatric disorders. We report here a highly polymorphic (dC-dA)n repeat within the human alpha 1-subunit gene (GABRA1). Typing of this marker in the Centre d'Etude du Polymorphisme Humain (CEPH) panel of families confirms the previous assignment of the GABRA1 locus to the distal portion of chromosome 5q by demonstrating linkage to the markers CRI-L45 (D5S61) (Zmax = 11.00, theta max = 0.15), CRI-V1022 (D5S54) (Zmax = 7.25, theta max = 0.20), and CRI-P148 (D5S72) (Zmax = 5.71, theta max = 0.24).     

Physical mapping of the von Recklinghausen neurofibromatosis region on chromosome 17

The von Recklinghausen neurofibromatosis (NF1) locus has been linked to chromosome 17, and recent linkage analyses place the gene on the proximal long arm. NF1 probably resides in 17q11.2, since two unrelated NF1 patients have been identified who possess constitutional reciprocal translocations involving 17q11.2 with chromosomes 1 and 22. We have used a somatic-cell hybrid from the t(17;22) individual, along with other hybrid cell lines, to order probes around the NF1 locus. An additional probe, 17L1, has been isolated from a NotI linking library made from flow-sorted chromosome 17 material and has been mapped to a region immediately proximal to the translocation breakpoint. While neither NF1 translocation breakpoint has yet been identified by pulse-field gel analysis, an overlap between two probes, EW206 and EW207, has been detected. Furthermore, we have identified the breakpoint in a non-NF1 translocation, SP-3, on the proximal side of the NF1 locus. This breakpoint has been helpful in creating a 1,000-kb pulsed-field map, which includes the closely linked NF1 probes HHH202 and TH17.19. The combined somatic-cell hybrid and pulsed-field gel analysis we report here favors the probe order D17Z1-HHH202-TH17.19-CRYB1-17L1-NF1- (EW206, EW207, EW203, L581, L946)-(ERBB2, ERBA1). The agreement in probe ordering between linkage analysis and physical mapping is excellent, and the availability of translocation breakpoints in NF1 should now greatly assist the cloning of this locus.     

Regional mapping panel for human chromosome 17: Application to neurofibromatosis type 1

A somatic cell hybrid mapping panel was constructed to localize cloned DNA sequences to any of 15 potentially different regions of human chromosome 17. Relatively high-resolution mapping is possible for 50% of the chromosome length in which 12 breakpoints are distributed over approximately 45 megabases, with an average spacing estimated at 1 breakpoint every 2-7 megabases. This high-resolution capability includes the pericentromeric region of 17 to which von Recklinghausen neurofibromatosis (NF1) has recently been mapped. Using 20 cloned genes and anonymous probes, we have tested the expected order and location of panel breakpoints and confirmed, refined, or corrected the regional assignment of several cloned genes and anonymous probes. Four markers with varying degrees of linkage to NF1 have been physically localized and ordered by the panel: the loosely linked markers myosin heavy chain 2 (25 cM) to p12----13.105 and nerve growth factor receptor (14 cM) to q21.1----q23; the more closely linked pABL10-41 (D17S71, 5 cM) to p11.2; and the tightly linked pHHH202 (D17S33) to q11.2-q12. Thus, physical mapping of linked markers confirms a pericentromeric location of NF1 and, along with other data, suggests the most likely localization is proximal 17q.     

No evidence for linkage of familial hypertrophic cardiomyopathy and chromosome 14q1 locus D14S26 in a chinese family: evidence for genetic heterogeneity

Summary To understand the molecular basis of familial hypertrophic cardiomyopathy (FHC) in the Chinese population, a family with FHC was investigated. Nineteen family members who were 16 years of age or older were examined by M-mode or two-dimensional echocardiography. Eight members were diagnosed to be affected echocardiographically or clinically. Lymphocytes isolated from 20 family members were successfully transformed into permanent lymphoblastoid cell lines by Epstein-Barr virus. Three genomic DNA probes (CRI-L436, CRI-L329, and pSC14) that were derived from chromosome 14q1 loci and demonstrated to be linked closely to FHC were used to probe this family. Using the techniques of restriction fragment length polymorphism (RFLP) and linkage analysis, the probe CRI-L436, which recognized locus D14S26, was found informative in this family. The lod scores were -2.0 at = 0.025 and -1.49 at = 0.05. Thus, there was no evidence of linkage between the locus D14S26 and the gene for FHC in the pedigree studied. In addition, polymerase chain reaction (PCR) amplification did not indicate a mutation on exon 13 of the cardiac myosin heavy chain gene as previously reported. Our data suggest that FHC is a genetically heterogeneous disease.     

A genomewide screen in multiplex rheumatoid arthritis families suggests genetic overlap with other autoimmune diseases

Rheumatoid arthritis (RA) is an autoimmune/inflammatory disorder with a complex genetic component. We report the first major genomewide screen of multiplex families with RA gathered in the United States. The North American Rheumatoid Arthritis Consortium, using well-defined clinical criteria, has collected 257 families containing 301 affected sibling pairs with RA. A genome screen for allele sharing was performed, using 379 microsatellite markers. A nonparametric analysis using SIBPAL confirmed linkage of the HLA locus to RA (P < .00005), with lambdaHLA = 1.79. However, the analysis also revealed a number of non-HLA loci on chromosomes 1 (D1S235), 4 (D4S1647), 12 (D12S373), 16 (D16S403), and 17 (D17S1301), with evidence for linkage at a significance level of P<.005. Analysis of X-linked markers using the MLOD method from ASPEX also suggests linkage to the telomeric marker DXS6807. Stratifying the families into white or seropositive subgroups revealed some additional markers that showed improvement in significance over the full data set. Several of the regions that showed evidence for nominal significance (P < .05) in our data set had previously been implicated in RA (D16S516 and D17S1301) or in other diseases of an autoimmune nature, including systemic lupus erythematosus (D1S235), inflammatory bowel disease (D4S1647, D5S1462, and D16S516), multiple sclerosis (D12S1052), and ankylosing spondylitis (D16S516). Therefore, genes in the HLA complex play a major role in RA susceptibility, but several other regions also contribute significantly to overall genetic risk.     

A microsatellite polymorphism associated with the PLC1 (phospholipase C) locus: identification, mapping, and linkage to the MODY locus on chromosome 20.

A highly polymorphic (dC-dA)n.(dG-dT)n dinucleotide repeat at the PLC1 locus on human chromosome 20 has been identified. Primers flanking the dinucleotide repeat were used for PCR amplification of the repeat region in 37 informative kindreds from the Centre d'Etude du Polymorphisme Humain. Two-point linkage analysis indicates that PLC1 is closely linked to several chromosome 20 markers, including D20S16 (Zmax = 41.25; theta = 0.07), D20S17 (Zmax = 42.81; theta = 0.09), and ADA (Zmax = 57.24; theta = 0.05). Multipoint linkage analysis places the PLC1 locus between D20S18 and D20S17, 11.2 and 6.6 cM, respectively, from these loci (sex-averaged distances). In addition, the PLC1 gene shows linkage to the maturity-onset diabetes of the young (MODY) locus on chromosome 20 with a lod score of 4.57 at theta = 0.089.     

Localization of the mutation in an extended family with Charcot-Marie-Tooth neuropathy (HMSN I).

Hereditary motor and sensory neuropathy type I (HMSN I) or Charcot-Marie-Tooth (CMT) disease is an autosomal dominant peripheral neuropathy. In some CMT families linkage has been reported with either the Duffy blood group or the APOA2 gene, both located on chromosome 1q. More recently, linkage has been found in six CMT families with two chromosome 17p markers. We extensively analyzed a multi-generation Charcot-Marie-Tooth family by using molecular genetic techniques in order to localize the CMT gene defect. First, we constructed a continuous linkage group of 11 chromosome 1 markers and definitely excluded chromosome 1 as the site of mutation. Second, we analyzed the family for linkage with chromosome 17. The two-point lod scores obtained with D17S58 and D17S71 proved that this Charcot-Marie-Tooth family is linked to chromosome 17. Moreover, multipoint linkage results indicated that the mutation is most likely located on the chromosome 17p arm, distal of D17S71.     

Linkage analysis of von Recklinghausen neurofibromatosis to DNA markers on chromosome 17

Several recent studies indicate that the von Recklinghausen neurofibromatosis (NF1) gene is located near the centromere of chromosome 17 in some families. However, variable expressivity and a very high mutation rate suggest that defects at several different loci could result in phenotypes categorized as NF1. In order to assess this possibility and to map the NF1 gene more precisely, we have used two polymorphic DNA markers from chromosome 17 to screen several pedigrees for linkage to NF1. We ascertained a large Caucasian pedigree (33 individuals sampled, 17 NF1 affected) as well as eight smaller pedigrees and nuclear families (50 individuals sampled, 30 NF1 affected). Here, we report strong evidence of linkage of NF1 to the centromeric marker D17Z1 (maximum lod = 4.42) and a weaker suggestion of linkage to the ERBA1 oncogene (maximum lod = 0.57), both at a recombination fraction of zero. Since obligate cross-overs with NF1 were not observed for either marker in any of the informative families tested, the possibility of NF1 locus heterogeneity is not supported.     

No evidence of linkage between the locus for autosomal dominant retinitis pigmentosa and D3S47 (C17) in three Australian families

Summary A linkage analysis has been performed on three Australian families segregating for autosomal dominant retinitis pigmentosa (ADRP). No evidence of linkage has been found in any of the pedigrees studied between the locus D3S47 and the gene for ADRP. The D3S47 locus was found to show very close linkage with the ADRP gene in a large Irish pedigree. Our study together with a similar report on a British family indicates that there is genetic heterogeneity in this disease.