A genome-wide search for chromosomal loci linked to bipolar affective disorder in the Old Order Amish.

The most characteristic features of bipolar affective disorder (manic-depressive illness) are episodes of mania (bipolar I, BPI) or hypomania (bipolar II, BPII) interspersed with periods of depression. Manic-depressive illness afflicts about one percent of the population, and if untreated, is associated with an approximately 20% risk of suicide. Twin, family and adoption studies provide compelling evidence for a partial genetic aetiology, but the mode(s) of inheritance has not been identified. Nonetheless, the majority of genetic linkage studies have assumed classical mendelian inheritance attributable to a single major gene. Although segregation analyses have yielded inconsistent results (with most studies rejecting a single locus inheritance model), the best single gene model is dominant inheritance if only BPI is considered. Reported linkages of bipolar affective disorder on chromosomes 11, 18, 21 and X have been difficult to substantiate, and additional studies are required for replication or exclusion of these regions. We now present the results of our genome-wide linkage analyses that provide evidence that regions on chromosomes 6, 13 and 15 harbour susceptibility loci for bipolar affective disorder, suggesting that bipolar affective disorder in the Old Order Amish is inherited as a complex trait.

Topoisomerase II alpha and topoisomerase II beta genes: characterization and mapping to human chromosomes 17 and 3, respectively.

Human cells contain two topoisomerase II isozymes named topo II alpha and topo II beta. The complementary DNAs for both enzymes have been cloned. The topo II alpha and topo II beta complementary DNAs hybridized to unique sequences of human, rodent, and chicken DNAs in Southern blots. The human topo II alpha gene has previously been mapped to chromosome 17. We confirmed the chromosomal location of topo II alpha and mapped the topo II beta gene to chromosome 3. In addition, topo II beta exhibits genetic polymorphism as has been reported for topoisomerases I and II alpha.

Update on the search for DNA markers linked to manic-depressive illness in the Old Order Amish.

In this report we describe our efforts to identify a gene involved in bipolar illness using a large, multigenerational Old Order Amish pedigree with many affected individuals. The original collection of cell lines from Amish pedigree 110 has been extended to include 169 individuals. We have used over 250 markers spaced at approximately 20 centiMorgans that detect restriction length fragment polymorphisms, but no LOD scores greater than 3 have been obtained from pairwise linkage analyses. We are expanding our collection of cell lines from both normal and affected family members and updating our diagnostic data as we continue our systematic screening of the genome for a gene involved in bipolar illness.

Family-based tests of association in the presence and absence of known linkage.

Correlation among sibling marker genotypes may invalidate the results of family-based tests of association in the presence of linkage. We apply an empirical variance estimation method, which is implemented in the program package FBAT, on Caucasian families with asthma in the presence and absence of linkage and compare the results with those obtained using the TDT (TDTEX-PAIRS) on the same data sets. Our results indicate that both tests generally perform comparably in either setting.

On the detection of linkage in multiple data sets: a comparison of various statistical approaches.

We contrast the pooling of multiple data sets with the compound HLOD (HLOD-C) and the posterior probability of linkage (PPL), two approaches that have been shown to have more power in the presence of genetic heterogeneity. We also propose and evaluate several multipoint extensions.

Phosphoenolpyruvate carboxykinase (GTP): characterization of the human PCK1 gene and localization distal to MODY on chromosome 20.

The human PCK1 gene encoding phosphoenolpyruvate carboxykinase (GTP) (PEPCK) was isolated and sequenced. There is 91% amino acid sequence identity (567/622 residues) between the human and the rat proteins, with conservation of intron/exon borders. A polymorphic dinucleotide microsatellite with the structure (CA)16(TA)5(CA) was identified in the 3' untranslated region of the cloned human PCK1 gene. This highly informative genetic marker has an estimated PIC value of 0.79 and heterozygosity of 0.81. Analysis of the RW pedigree demonstrated recombination between PCK1 and the MODY gene on chromosome 20. Multipoint linkage analysis of the reference pedigrees of the Centre d'Etude du Polymorphisme Humain localized PCK1 on the genetic map of chromosome 20 at a position distal to markers that are closely linked to MODY. PCK1 is part of a conserved linkage group on mouse Chromosome 2 with identical gene order but expanded length in the human genome.

A genetic linkage map of human chromosome 5 with 60 RFLP loci.

A genetic map of human chromosome 5 that contains 60 restriction fragment length polymorphism (RFLP) loci in one linkage group has been constructed. Segregation data using these markers and 40 large multigenerational families supplied by the Centre d'Etude du Polymorphisme Humain have been collected. Linkage analyses were performed with the program package CRI-MAP; using odds greater than 1000:1, 30 RFLP loci could be placed on the map. This genetic map spans 289 cM sex-equal, 353 cM in females, and 244 cM in males. While the relative rate of recombination for female meioses is nearly twice that of males over much of the chromosome, several instances of statistically significant excess male recombination were observed. The order of probes on the genetic map has been confirmed by their physical order as determined by somatic cell hybrid lines containing deletions of normal chromosome 5. There is concordance between the physical positions of markers and their genetic positions. Our most distal probes on the genetic map are cytologically localized to the most distal portions of the chromosome. This suggests that our genetic map spans most of chromosome 5.

Linkage analysis of maturity-onset diabetes of the young (MODY): genetic heterogeneity and nonpenetrance.

We have analyzed the inheritance of maturity-onset diabetes of the young (MODY) on chromosome 20 in a large multigeneration family, the R.-W. family, and in two other MODY families. Of the four branches of the R.-W. pedigree which have been studied, two have documented early onset of non-insulin-dependent diabetes mellitus (NIDDM), while there is no evidence of early onset in the other two branches. The early-onset branches have apparently inherited the same D20S16 allele from the affected parent, while another D20S16 allele was inherited in the two branches without evidence of early onset. A test for homogeneity, the M-test, using the results of two-point linkage analysis with D20S16 indicates heterogeneity between early- and late-onset branches of the R.-W. family (P less than or equal to .014). In addition, analysis strongly suggests that MODY as expressed in the EDI and WIS families is unlinked to loci on chromosome 20 (P less than or equal to .018-.004). Comparable results are seen when the data are analyzed by the HOMOG program. Three polymorphic loci-D20S16, D20S17, and ADA--show no recombination with the MODY locus when two-point linkage analysis is used in the early-onset branches of the family. The multipoint lod score in the early-onset branches of the R.-W. family is 10.16, with the most likely location being between D20S4 and D20S17. Multipoint linkage analysis using the CHROMPICS option of the program CRI-MAP has been used to follow inheritance of the MODY disease locus. This analysis has identified two cases of possible nonpenetrance in the early-onset branches of the family (odds of at least 156:1), as determined by the appearance of apparent isolated double crossovers at the MODY locus in these unaffected individuals.

Genetic analysis of eight loci tightly linked to neurofibromatosis 1.

The genetic locus for neurofibromatosis 1 (NF1) has recently been mapped to the pericentromeric region of chromosome 17. We have genotyped eight previously identified RFLP probes on 50 NF1 families to determine the placement of the NF1 locus relative to the RFLP loci. Thirty-eight recombination events in the pericentromeric region were identified, eight involving crossovers between NF1 and loci on either chromosomal arm. Multipoint linkage analysis resulted in the unique placement of six loci at odds greater than 100:1 in the order of pter-A10-41-EW301-NF1-EW207-CRI-L581-CRI-L946 -qter. Owing to insufficient crossovers, three loci--D17Z1, EW206, and EW203--could not be uniquely localized. In this region female recombination rates were significantly higher than those of males. These data were part of a joint study aimed at the localization of both NF1 and tightly linked pericentromeric markers for chromosome 17.

Framework multipoint map of the long arm of human chromosome 4 and telomeric localization of the gene for FSHD.

Mapping the long arm of Chromosome (Chr) 4 has assumed medical relevance with the establishment of linkage of facioscapulohumeral muscular dystrophy (FSHD) to distal 4q markers. We have constructed a multipoint linkage map using DNA markers that map to the long arm of Chr 4. Segregation data were collected for 17 DNA markers on the multigenerational CEPH mapping families, and data for one marker were taken from the published CEPH database. Genotypic information for six of these markers was also collected from a set of 24 families that exhibited inheritance of FSHD. Multipoint analyses allowed us to construct a map of 12 loci, connecting two previously separate linkage groups. Significant sex-specific differences in recombination were found for some genetic intervals. Four loci from the distal region of this map showed linkage with FSHD. A map using these terminal markers gave the strongest support for FSHD in the most distal position over all other possible positions.

Linkage analyses of five chromosome 4 markers localizes the facioscapulohumeral muscular dystrophy (FSHD) gene to distal 4q35.

The genetic locus for facioscapulohumeral muscular dystrophy (FSHD) has been mapped to chromosome 4. We have examined linkage to five chromosome 4q DNA markers in 22 multigenerational FSHD families. Multipoint linkage analyses of the segregation of four markers in the FSHD families and in 40 multigenerational mapping families from the Centre d'Etude du Polymorphisme Humaine enabled these loci and FSHD to be placed in the following order: cen-D4S171-factor XI-D4S163-D4S139-FSHD-qter. One interval, D4S171-FSHD, showed significant sex-specific differences in recombination. Homogeneity tests supported linkage of FSHD to these 4q DNA markers in all of the families we studied. The position of FSHD is consistent with that generated by other groups as members of an international FSHD consortium.

Genetic linkage map of 46 DNA markers on human chromosome 16.

We have constructed a genetic linkage map of human chromosome 16 based on 46 DNA markers that detect restriction fragment length polymorphisms. Segregation data were collected on a set of multigenerational families provided by the Centre d'Etude du Polymorphisme Humain, and maps were constructed using recently developed multipoint analysis techniques. The map spans 115 centimorgans (cM) in males and 193 cM in females. Over much of the chromosome there is a significantly higher frequency of recombination in females than males. Near the alpha-globin locus on the distal part of the short arm, however, there is a significant excess of male recombination. Twenty-seven (59%) of the markers on the map have heterozygosities greater than or equal to 0.50. The largest interval between loci on the sex-average map is 14 cM and the average marker spacing is 3 cM. Using loci on this map, one could detect linkage to a dominant disease on chromosome 16 with as few as 10-15 phase-known meioses.

Genetic characterization of the APC locus involved in familial adenomatous polyposis.

Familial adenomatous polyposis is a rare disease inherited in a Mendelian dominant fashion. It is characterized by the occurrence of more than 100 adenomatous polyps in the large bowels of affected individuals. The genetic defect responsible for adenomatous polyposis resides at a locus called APC which has been localized to the long arm of human chromosome 5. In this study, the APC locus was mapped with respect to 11 markers known to map to this chromosomal segment. Linkage of APC to four of these markers had been previously reported. Three additional markers are shown here to be linked to APC. By multipoint analysis, the APC locus maps to an interval bounded by D5S49 and D5S58. The refined map of the APC locus and the new markers described here improve the informativeness and accuracy of the presymptomatic diagnosis of familial adenomatous polyposis.

A clinically homogeneous group of families with facioscapulohumeral (Landouzy-Déjérine) muscular dystrophy: linkage analysis of six autosomes.

Facioscapulohumeral muscular dystrophy (FSHMD) is a neuromuscular disorder characterized by autosomal dominant inheritance and clinical onset in the muscles of the face and shoulder girdle. Using a set of RFLP markers spaced at approximately 20 centimorgans, we have begun a systematic search for markers linked to the disease. A total of 81 RFLP loci on six autosomes (1, 2, 5, 7, 10, and 16) have been examined for linkage to FSHMD in 13 families. With the computer program CRI-MAP, two-point and multipoint analyses have not resulted in any LOD score indicative of linkage to FSHMD. However, these analyses have allowed us to exclude 909 centimorgans (sex average) of our genetic maps in intervals where the LOD score is less than -2.0. We estimate our data have excluded 23% of the human genome.

A genetic linkage map of 32 loci on human chromosome 10.

We have constructed a genetic linkage map of human chromosome 10 based on DNA probes that detect 47 restriction fragment length polymorphisms (RFLPs) at 32 different loci. Segregation data were collected on a set of multigenerational families provided by the Centre d'Etude du Polymorphisme Humain and maps were constructed using recently developed multipoint analysis techniques. The length of the sex-averaged map is 178 cM and the sex-specific map lengths are 131 cM in males and 255 cM in females. Recombination is significantly higher in female meioses. The mean distance between loci is 5.6 cM for the sex-averaged map. The genetic map spans the length of the chromosome as judged by physical localization of probes by in situ hybridization techniques and mapping of the probes on human-hamster hybrid cell lines containing all or part of chromosome 10. The informativeness of two loci near the locus responsible for multiple endocrine neoplasia type 2A (MEN-2A) has been increased by isolation of cosmids that reveal additional RFLPs at these loci.

Linkage analyses of chromosome 18 markers do not identify a major susceptibility locus for bipolar affective disorder in the Old Order Amish.

Previously reported linkage of bipolar affective disorder to DNA markers in the pericentromeric region of chromosome 18 was reexamined in a larger homogeneous sample of Old Order Amish families. Four markers (D18S21, D18S53, D18S44, and D18S40) were examined in three kindreds containing 31 bipolar I (BP I) individuals. Although linkage findings were replicated in the one previously studied Amish pedigree containing four BP I individuals, linkage to this region was excluded in the larger sample. If a susceptibility locus for bipolar disorder is located in this region of chromosome 18, it is of minor significance in this population.

Multiplex sequencing of 1.5 Mb of the Mycobacterium leprae genome.

The nucleotide sequence of 1.5 Mb of genomic DNA from Mycobacterium leprae was determined using computer-assisted multiplex sequencing technology. This brings the 2.8-Mb M. leprae genome sequence to approximately 66% completion. The sequences, derived from 43 recombinant cosmids, contain 1046 putative protein-coding genes, 44 repetitive regions, 3 tRNAs, and 15 tRNAs. The gene density of one per 1.4 kb is slightly lower than that of Mycoplasma (1.2 kb). Of the protein coding genes, 44% have significant matches to genes with well-defined functions. Comparison of 1157 M. leprae and 1564 Mycobacterium tuberculosis proteins shows a complex mosaic of homologous genomic blocks with up to 22 adjacent proteins in conserved map order. Matches to known enzymatic, antigenic, membrane, cell wall, cell division, multidrug resistance, and virulence proteins suggest therapeutic and vaccine targets. Unusual features of the M. leprae genome include large polyketide synthase (pks) operons, inteins, and highly fragmented pseudogenes.

A genome-wide search for chromosomal loci linked to mental health wellness in relatives at high risk for bipolar affective disorder among the Old Order Amish.

Bipolar affective disorder (BPAD; manic-depressive illness) is characterized by episodes of mania and/or hypomania interspersed with periods of depression. Compelling evidence supports a significant genetic component in the susceptibility to develop BPAD. To date, however, linkage studies have attempted only to identify chromosomal loci that cause or increase the risk of developing BPAD. To determine whether there could be protective alleles that prevent or reduce the risk of developing BPAD, similar to what is observed in other genetic disorders, we used mental health wellness (absence of any psychiatric disorder) as the phenotype in our genome-wide linkage scan of several large multigeneration Old Order Amish pedigrees exhibiting an extremely high incidence of BPAD. We have found strong evidence for a locus on chromosome 4p at D4S2949 (maximum GENEHUNTER-PLUS nonparametric linkage score = 4.05, P = 5. 22 x 10(-4); SIBPAL Pempirical value <3 x 10(-5)) and suggestive evidence for a locus on chromosome 4q at D4S397 (maximum GENEHUNTER-PLUS nonparametric linkage score = 3.29, P = 2.57 x 10(-3); SIBPAL Pempirical value <1 x 10(-3)) that are linked to mental health wellness. These findings are consistent with the hypothesis that certain alleles could prevent or modify the clinical manifestations of BPAD and perhaps other related affective disorders.