Familial isolated hyperparathyroidism is linked to a 1.7 Mb region on chromosome 2p13.3-14

BACKGROUND: Familial isolated hyperparathyroidism (FIHP) is an autosomal dominantly inherited form of primary hyperparathyroidism. Although comprising only about 1% of cases of primary hyperparathyroidism, identification and functional analysis of a causative gene for FIHP is likely to advance our understanding of parathyroid physiology and pathophysiology. METHODS: A genome-wide screen of DNA from seven pedigrees with FIHP was undertaken in order to identify a region of genetic linkage with the disorder. RESULTS: Multipoint linkage analysis identified a region of suggestive linkage (LOD score 2.68) on chromosome 2. Fine mapping with the addition of three other families revealed significant linkage adjacent to D2S2368 (maximum multipoint LOD score 3.43). Recombination events defined a 1.7 Mb region of linkage between D2S2368 and D2S358 in nine pedigrees. Sequencing of the two most likely candidate genes in this region, however, did not identify a gene for FIHP. CONCLUSIONS: We conclude that a causative gene for FIHP lies within this interval on chromosome 2. This is a major step towards eventual precise identification of a gene for FIHP, likely to be a key component in the genetic regulation of calcium homeostasis.

Identification of the semaphorin receptor PLXNA2 as a candidate for susceptibility to schizophrenia

The discovery of genetic factors that contribute to schizophrenia susceptibility is a key challenge in understanding the etiology of this disease. Here, we report the identification of a novel schizophrenia candidate gene on chromosome 1q32, plexin A2 (PLXNA2), in a genome-wide association study using 320 patients with schizophrenia of European descent and 325 matched controls. Over 25,000 single-nucleotide polymorphisms (SNPs) located within approximately 14,000 genes were tested. Out of 62 markers found to be associated with disease status, the most consistent finding was observed for a candidate locus on chromosome 1q32. The marker SNP rs752016 showed suggestive association with schizophrenia (odds ratio (OR) = 1.49, P = 0.006). This result was confirmed in an independent case-control sample of European Americans (combined OR = 1.38, P = 0.035) and similar genetic effects were observed in smaller subsets of Latin Americans (OR = 1.26) and Asian Americans (OR = 1.37). Supporting evidence was also obtained from two family-based collections, one of which reached statistical significance (OR = 2.2, P = 0.02). High-density SNP mapping showed that the region of association spans approximately 60 kb of the PLXNA2 gene. Eight out of 14 SNPs genotyped showed statistically significant differences between cases and controls. These results are in accordance with previous genetic findings that identified chromosome 1q32 as a candidate region for schizophrenia. PLXNA2 is a member of the transmembrane semaphorin receptor family that is involved in axonal guidance during development and may modulate neuronal plasticity and regeneration. The PLXNA2 ligand semaphorin 3A has been shown to be upregulated in the cerebellum of individuals with schizophrenia. These observations, together with the genetic results, make PLXNA2 a likely candidate for the 1q32 schizophrenia susceptibility locus.

Genomewide significant linkage to migrainous headache on chromosome 5q21

Familial typical migraine is a common, complex disorder that shows strong familial aggregation. Using latent-class analysis (LCA), we identified subgroups of people with migraine/severe headache in a community sample of 12,245 Australian twins (60% female), drawn from two cohorts of individuals aged 23-90 years who completed an interview based on International Headache Society criteria. We report results from genomewide linkage analyses involving 756 twin families containing a total of 790 independent sib pairs (130 affected concordant, 324 discordant, and 336 unaffected concordant for LCA-derived migraine). Quantitative-trait linkage analysis produced evidence of significant linkage on chromosome 5q21 and suggestive linkage on chromosomes 8, 10, and 13. In addition, we replicated previously reported typical-migraine susceptibility loci on chromosomes 6p12.2-p21.1 and 1q21-q23, the latter being within 3 cM of the rare autosomal dominant familial hemiplegic migraine gene (ATP1A2), a finding which potentially implicates ATP1A2 in familial typical migraine for the first time. Linkage analyses of individual migraine symptoms for our six most interesting chromosomes provide tantalizing hints of the phenotypic and genetic complexity of migraine. Specifically, the chromosome 1 locus is most associated with phonophobia; the chromosome 5 peak is predominantly associated with pulsating headache; the chromosome 6 locus is associated with activity-prohibiting headache and photophobia; the chromosome 8 locus is associated with nausea/vomiting and moderate/severe headache; the chromosome 10 peak is most associated with phonophobia and photophobia; and the chromosome 13 peak is completely due to association with photophobia. These results will prove to be invaluable in the design and analysis of future linkage and linkage disequilibrium studies of migraine.

Separate and interacting effects within the catechol-O-methyltransferase (COMT) are associated with schizophrenia

Several lines of evidence have implicated the catechol-O-methyltransferase (COMT) gene as a candidate for schizophrenia (SZ) susceptibility, not only because it encodes a key dopamine catabolic enzyme but also because it maps to the velocardiofacial syndrome region of chromosome 22q11 which has long been associated with SZ predisposition. The interest in COMT as a candidate SZ risk factor has led to numerous case-control and family-based studies, with the majority placing emphasis on examining a functional Val/Met polymorphism within this enzyme. Unfortunately, these studies have continually produced conflicting results. To assess the genetic contribution of other COMT variants to SZ susceptibility, we investigated three single-nucleotide polymorphisms (SNPs) (rs737865, rs4633, rs165599) in addition to the Val/Met variant (rs4680) in a highly selected sample of Australian Caucasian families containing 107 patients with SZ. The Val/Met and rs4633 variants showed nominally significant associations with SZ (P<0.05), although neither of the individual SNPs remained significant after adjusting for multiple testing (most significant P=0.1174). However, haplotype analyses showed strong evidence of an association; the most significant being the three-marker haplotype rs737865-rs4680-rs165599 (global P=0.0022), which spans more than 26 kb. Importantly, conditional analyses indicated the presence of two separate and interacting effects within this haplotype, irrespective of gender. In addition, our results indicate the Val/Met polymorphism is not disease-causing and is simply in strong linkage disequilibrium with a causative effect, which interacts with another as yet unidentified variant approximately 20 kb away. These results may help explain the inconsistent results reported on the Val/Met polymorphism and have important implications for future investigations into the role of COMT in SZ susceptibility.

Use of pulsed field gel electrophoresis to characterize BCR gene involvement in CML patients lacking M-BCR rearrangement.

We studied the pattern of BCR involvement in 52 patients with chronic myeloid leukemia by Southern blotting. Of 33 Philadelphia (Ph)-positive patients, 30 had evidence of M-BCR rearrangement, two cases were difficult to interpret, and one clearly lacked evidence of M-BCR rearrangement. Of 19 Ph-negative patients, nine showed M-BCR rearrangement, nine showed no rearrangement, and one result was uncertain. We selected for more detailed study eight patients (three Ph-positive and five Ph-negative). Two of the Ph-positive patients, whose Southern blots were difficult to interpret, had rearranged bands when the BCR gene was studied by pulsed field gel electrophoresis (PFGE). Results of PFGE studies and in situ hybridization to metaphase chromosomes in the third Ph-positive patient, whose DNA clearly lacked M-BCR rearrangement on Southern analysis, were consistent with a breakpoint on chromosome 22 located 3' of all known exons of the BCR gene. However, mRNA studied with the polymerase chain reaction showed evidence of a classical b2-a2 linkage. The findings in this patient may be explained by an unusual genomic breakpoint downstream of the BCR gene associated with long range splicing that excluded all of the 3' BCR exons. Of the five patients with Ph-negative M-BCR non-rearranged CML studied by PFGE for BCR gene rearrangement, none had evidence of rearranged bands. We conclude that PFGE is a valuable adjunct to standard molecular techniques for the study of atypical cases of CML. Occasional patients with Ph-positive CML have breakpoints outside M-BCR. The BCR gene is probably not involved in patients with Ph-negative, M-BCR non-rearranged CML.

Interstitial insertion of varying amounts of ABL-containing genetic material into chromosome 22 in Ph-negative CML.

We studied the cells from three selected patients with Ph-chromosome-negative chronic myeloid leukemia (CML) by Southern blotting, polymerase chain reaction, and in situ hybridization of informative probes to metaphase chromosomes. All three patients had rearrangement of M-BCR sequences in the BCR gene and expression of one or other of the mRNA species characteristic of Ph-positive CML. Leukemic metaphases studied after trypsin-Giemsa banding were indistinguishable from normal. The ABL probe localized both to chromosome 9 and 22 in each case. A probe containing 3' M-BCR sequences localized only to chromosome 22, and not to chromosome 9 as would be expected in Ph-positive CML. Two new probes that recognize different polymorphic regions distal to the ABL gene on chromosome 9 in normal subjects localized exclusively to chromosome 9 in two patients and to both chromosomes 9 and 22 in one patient. These results show that Ph-negative CML with BCR rearrangement is associated with insertion of a variable quantity of chromosome 9 derived material into chromosome 22q11; there is no evidence for reciprocal translocation of material from chromosome 22 to chromosome 9.

Dual-colour HER2/chromosome 17 chromogenic in situ hybridisation enables accurate assessment of HER2 genomic status in ovarian tumours

Ovarian cancer is a leading cause of gynaecological cancer-related morbidity and mortality. There has been increasing interest in the potential utility of anti-human epidermal growth factor receptor 2 (anti-HER2) agents in the treatment of this disease, with the attendant need to identify suitable predictive biomarkers of response to treatment.

Dual-colour HER2/chromosome 17 chromogenic in situ hybridisation assay enables accurate assessment of HER2 genomic status in gastric cancer and has potential utility in HER2 testing of biopsy samples

Determination of HER2 protein expression by immunohistochemistry (IHC) and genomic status by fluorescent in situ hybridisation (FISH) are important in identifying a subset of high HER2-expressing gastric cancers that might respond to trastuzumab. Although FISH is considered the standard for determination of HER2 genomic status, brightfield ISH is being increasingly recognised as a viable alternative. Also, the impact of HER2 protein expression/genomic heterogeneity on the accuracy of HER2 testing has not been well studied in the context of gastric biopsy samples.

Exclusion of linkage between schizophrenia and the D2 dopamine receptor gene region of chromosome 11q in 112 Irish multiplex families

A leading theory hypothesizes that schizophrenia arises from dysregulation of the dopamine system in certain brain regions. As this dysregulation could arise from abnormal expression of D2 dopamine receptors, the D2 receptor gene (DRD2) on chromosome 11q is a candidate locus for schizophrenia. We tested whether allelic variation at DRD2 and five surrounding loci cosegregated with schizophrenia in 112 small- to moderate-size Irish families containing two or more members affected with schizophrenia or schizoaffective disorder, defined by DSM-III-R. Evidence of linkage was assessed using varying definitions of illness and modes of transmission. Assuming genetic homogeneity, linkage between schizophrenia and large regions of 11q around DRD2 could be strongly excluded. Assuming genetic heterogeneity, variation at the DRD2 locus could be rejected as a major risk factor for schizophrenia in more than 50% of these families for all models tested and in as few as 25% of the families for certain models. The DRD2 linkage in fewer than 25% of these families could not be excluded under any of the models tested. Our results suggest that the major component of genetic susceptibility to schizophrenia is not due to allelic variation at the DRD2 locus or other genes in the surrounding chromosomal region.

Evidence for a schizophrenia vulnerability locus on chromosome 8p in the Irish Study of High-Density Schizophrenia Families

This study was an attempt to replicate evidence for a vulnerability locus for schizophrenia and associated disorders in the 8p22-21 region reported by Pulver and colleagues.

Support for a possible schizophrenia vulnerability locus in region 5q22-31 in Irish families

In our genome scan for schizophrenia genes in 265 Irish pedigrees, marker D5S818 in 5q22 produced the second best result of the first 223 markers tested (P = 0.002). We then tested an additional 13 markers and the evidence suggests the presence of a vulnerability locus for schizophrenia in region 5q22-31. This region appears to be distinct from those chromosome 5 regions studied in two prior reports, but the same as that producing positive results in the report by Wildenauer and colleagues found elsewhere in this issue. The largest pairwise heterogeneity LOD (H-LOD) score was found with marker D5S393 (max 3.04, P = 0.0005), assuming a narrow phenotypic category, and a genetic model with intermediate heterozygotic liability. In marked contrast to the H-LOD scores from our sample with markers from the regions of interest on chromosomes 6p and 8p, expanding the disease definition to include schizophrenia spectrum or nonspectrum disorders produced substantially smaller scores, with a number of markers failing to yield positive values at any recombination fraction. Using multipoint H-LODS, the strongest evidence for linkage occurs under the narrow phenotypic definition and recessive genetic model, with a peak at marker D5S804 (max 3.35, P = 0.0002). Multipoint nonparametric linkage analysis produced a peak in the same location (max z = 2.84, P = 0.002) with the narrow phenotypic definition. This putative vulnerability locus appears to be segregating in 10-25% of the families studied, but this estimate is tentative. Comparison of individual family multipoint H-LOD scores at the regions of interest on chromosomes 6p, 8p and 5q showed that only a minority of families yield high lod scores in two or three regions.

No major schizophrenia locus detected on chromosome 1q in a large multicenter sample

Reports of substantial evidence for genetic linkage of schizophrenia to chromosome 1q were evaluated by genotyping 16 DNA markers across 107 centimorgans of this chromosome in a multicenter sample of 779 informative schizophrenia pedigrees. No significant evidence was observed for such linkage, nor for heterogeneity in allele sharing among the eight individual samples. Separate analyses of European-origin families, recessive models of inheritance, and families with larger numbers of affected cases also failed to produce significant evidence for linkage. If schizophrenia susceptibility genes are present on chromosome 1q, their population-wide genetic effects are likely to be small.

No evidence for linkage or association of neuregulin-1 (NRG1) with disease in the Irish study of high-density schizophrenia families (ISHDSF)

The neuregulin-1 gene (NRG1) at chromosome 8p21-22 has been implicated as a schizophrenia susceptibility gene in Icelandic, Scottish, Irish and mixed UK populations. The shared ancestry between these populations led us to investigate the NRG1 polymorphisms and appropriate marker haplotypes for linkage and/or association to schizophrenia in the Irish study of high-density schizophrenia families (ISHDSF). Neither single-point nor multi-point linkage analysis of NRG1 markers gave evidence for linkage independent of our pre-existing findings telomeric on 8p. Analysis of linkage disequilibrium (LD) across the 252 kb interval encompassing the 7 marker core Icelandic/Scottish NRG1 haplotype revealed two separate regions of modest LD, comprising markers SNP8NRG255133, SNP8NRG249130 and SNP8NRG243177 (telomeric) and microsatellites 478B14-428, 420M9-1395, D8S1810 and 420M9-116I12 (centromeric). From single marker analysis by TRANSMIT and FBAT we found no evidence for association with schizophrenia for any marker. Haplotype analysis for the three SNPs in LD region 1 and, separately, the four microsatellites in LD region 2 (analyzed in overlapping 2-marker windows), showed no evidence for overtransmission of specific haplotypes to affected individuals. We therefore conclude that if NRG1 does contain susceptibility alleles for schizophrenia, they impact quite weakly on risk in the ISHDSF.

Relationship between a high-risk haplotype in the DTNBP1 (dysbindin) gene and clinical features of schizophrenia

The purpose of this study was to determine whether a haplotype in the dystrobrevin binding protein 1 (DTNBP1) gene previously associated with schizophrenia not only increases the susceptibility to psychotic illness but also to a more or less clinically specific form of psychotic illness.

Association between the 5q31.1 gene neurogenin1 and schizophrenia

Multiple lines of evidence suggest that schizophrenia results from aberrant neurodevelopment. The neurogenin1 gene (neurog1) consists of a single 1,666 bp exon that encodes a basic helix-loop-helix (bHLH) transcription factor that causes neuronal differentiation and induces cortical and glutamatergic differentiation programs. Because of its function and its location in 5q31.1, which has been linked to schizophrenia in multiple samples, we tested it for association with the disorder. We sequenced neurog1 in 25 affected subjects from the Irish Study of High-Density Schizophrenia Families. We observed a 5'-UTR SNP at position -60, already present in databases as rs8192558, and tested it along with rs2344485, rs8192559, and rs2344484. Narrow, intermediate, and broad diagnostic definitions were used. The major alleles of rs8192558 and rs2344484 were over-transmitted to affected subjects using both Pedigree Disequilibrium Test (PDT) (0.01 <or = P <or = 0.06) and FBAT (0.02 <or = P <or = 0.07). A haplotype consisting of the major alleles of all four SNPs was significantly over-transmitted in FBAT to the broad definition (P = 0.049), with trend significance to the narrow and intermediate definitions, and with trend significance in PDT. In confirmatory tests using 657 cases and 411 controls, this haplotype was slightly but not significantly over-represented in cases (81% vs. 77%, P = 0.21). These results, along with a priori evidence for the involvement of neurog1 in neurodevelopment, suggest that variants in neurog1 might have a small effect on susceptibility to schizophrenia. This gene should be tested in additional and larger samples.