Exome Sequencing of Extended Families with Alzheimer's Disease Identifies Novel Genes Implicated in Cell Immunity and Neuronal Function.

OBJECTIVE: Alzheimer's disease (AD) is a neurodegenerative disorder for which more than 20 genetic loci have been implicated to date. However, studies demonstrate not all genetic factors have been identified. Therefore, in this study we seek to identify additional rare variants and novel genes potentially contributing to AD. METHODS: Whole exome sequencing was performed on 23 multi-generational families with an average of eight affected subjects. Exome sequencing was filtered for rare, nonsynonymous and loss-of-function variants. Alterations predicted to have a functional consequence and located within either a previously reported AD gene, a linkage peak (LOD>2), or clustering in the same gene across multiple families, were prioritized. RESULTS: Rare variants were found in known AD risk genes including AKAP9, CD33, CR1, EPHA1, INPP5D, NME8, PSEN1, SORL1, TREM2 and UNC5C. Three families had five variants of interest in linkage regions with LOD>2. Genes with segregating alterations in these peaks include CD163L1 and CLECL1, two genes that have both been implicated in immunity, CTNNA1, which encodes a catenin in the cerebral cortex and MIEF1, a gene that may induce mitochondrial dysfunction and has the potential to damage neurons. Four genes were identified with alterations in more than one family include PLEKHG5, a gene that causes Charcot-Marie-Tooth disease and THBS2, which promotes synaptogenesis. CONCLUSION: Utilizing large families with a heavy burden of disease allowed for the identification of rare variants co-segregating with disease. Variants were identified in both known AD risk genes and in novel genes.

Targeted sequencing of ABCA7 identifies splicing, stop-gain and intronic risk variants for Alzheimer disease.

Several variants in the gene ABCA7 have been identified as potential causal variants for late-onset Alzheimer's disease (LOAD). In order to replicate these findings, and search for novel causal variants, we performed targeted sequencing of this gene in cohorts of non-Hispanic White (NHW) and African-American (AA) LOAD cases and controls. We sequenced the gene ABCA7 in 291 NHW LOAD cases and 103 controls. Variants were prioritized for rare, damaging variants and previously reported variants associated with LOAD, and were follow-up genotyped in 4076 NHW and 1157 AA cases and controls. We confirm three previously associated ABCA7 risk variants and extend two of these associations to other populations, an intronic variant in NHW (P=3.0×10-3) (originally reported in a Belgian population), and a splice variant originally associated in the Icelandic population, which was significantly associated in the NHW cohort (P=1.2×10-6) and nominally associated in the AA cohort (P=0.017). We also identify a 3'-UTR splice variant that segregates in four siblings of one family and is nominally associated with LOAD (P=0.040). Multiple variants in ABCA7 contribute to LOAD risk.

Variants in several genomic regions associated with asperger disorder.

Asperger disorder (ASP) is one of the autism spectrum disorders (ASD) and is differentiated from autism largely on the absence of clinically significant cognitive and language delays. Analysis of a homogenous subset of families with ASP may help to address the corresponding effect of genetic heterogeneity on identifying ASD genetic risk factors. To examine the hypothesis that common variation is important in ASD, we performed a genome-wide association study (GWAS) in 124 ASP families in a discovery data set and 110 ASP families in a validation data set. We prioritized the top 100 association results from both cohorts by employing a ranking strategy. Novel regions on 5q21.1 (P = 9.7 × 10(-7) ) and 15q22.1-q22.2 (P = 7.3 × 10(-6) ) were our most significant findings in the combined data set. Three chromosomal regions showing association, 3p14.2 (P = 3.6 × 10(-6) ), 3q25-26 (P = 6.0 × 10(-5) ) and 3p23 (P = 3.3 × 10(-4) ) overlapped linkage regions reported in Finnish ASP families, and eight association regions overlapped ASD linkage areas. Our findings suggest that ASP shares both ASD-related genetic risk factors, as well as has genetic risk factors unique to the ASP phenotype.

Association and gene-gene interaction of SLC6A4 and ITGB3 in autism.

Autism is a heritable neurodevelopmental disorder with substantial genetic heterogeneity. Studies point to possible links between autism and two serotonin related genes: SLC6A4 and ITGB3 with a sex-specific genetic effect and interaction between the genes. Despite positive findings, inconsistent results have complicated interpretation. This study seeks to validate and clarify previous findings in an independent dataset taking into account sex, family-history (FH) and gene-gene effects. Family-based association analysis was performed within each gene. Gene-gene interactions were tested using extended multifactor dimensionality reduction (EMDR) and MDR-phenomics (MDR-P) using sex of affecteds and FH as covariates. No significant associations with individual SNPs were found in the datasets stratified by sex, but associations did emerge when we stratified by family history. While not significant in the overall dataset, nominally significant association was identified at RS2066713 (P = 0.006) within SLC6A4 in family-history negative (FH-) families, at RS2066713 (P = 0.038) in family-history positive (FH+) families but with the opposite risk allele as in the FH- families. For ITGB3, nominally significant association was identified at RS3809865 overall (P = 0.040) and within FH+ families (P = 0.031). However, none of the associations survived the multiple testing correction. MDR-P confirmed gene-gene effects using sex of affecteds (P = 0.023) and family history (P = 0.014, survived the multiple testing corrections) as covariates. Our results indicate the extensive heterogeneity within these two genes among families. The potential interaction between SLC6A4 and ITGB3 may be clarified using family history as an indicator of genetic architecture, illustrating the importance of covariates as markers of heterogeneity in genetic analyses.

Association of SLC11A1 with tuberculosis and interactions with NOS2A and TLR2 in African-Americans and Caucasians.

SETTING: Host defense factors may influence the development of active tuberculosis (TB). OBJECTIVE: To test variants in solute carrier family 11A, member 1 (SLC11A1), for an association with TB. METHODS: A mixed case-control study of TB cases, relatives or close contact controls, consisting of 474 African-Americans (243 families) and 381 Caucasians (192 families), examined 13 SLC11A1 polymorphisms for association with pulmonary TB using generalized estimating equations adjusting for age and sex. RESULTS: Two associations were observed in Caucasians (rs3731863, P = 0.03, and rs17221959, P = 0.04) and one in African-Americans (rs3731865, P = 0.05). Multilocus analyses between polymorphisms in SLC11A1 and 11 TB candidate genes detected interactions between SLC11A1 and inducible nitric oxide synthase (NOS2A) in Caucasians (rs3731863 [SLC11A1] x rs8073782 [NOS2A], P = 0.009; rs3731863 [SLC11A1] x rs17722851 [NOS2A], P = 0.007) and toll-like receptor 2 (TLR2) in African-Americans (rs3731865 [SLC11A1] x rs1816702, P = 0.005). CONCLUSIONS: No association was detected with 5'(GT)(n) promoter polymorphism previously associated with lower SLC11A1 expression, rs17235409 (D543N), or rs17235416 (3' TGTG insertion/deletion polymorphism). SLC11A1 polymorphism rs3731865 was associated with TB in African-Americans, consistent with previous findings in West Africans. These results suggest that variants in SLC11A1 increase susceptibility to pulmonary TB and interact with other variants that differ by race.

Examination of tetrahydrobiopterin pathway genes in autism.

Autism is a complex disorder with a high degree of heritability and significant phenotypic and genotypic heterogeneity. Although candidate gene studies and genome-wide screens have failed to identify major causal loci associated with autism, numerous studies have proposed association with several variations in genes in the dopaminergic and serotonergic pathways. Because tetrahydrobiopterin (BH4) is the essential cofactor in the synthesis of these two neurotransmitters, we genotyped 25 SNPs in nine genes of the BH4 pathway in a total of 403 families. Significant nominal association was detected in the gene for 6-pyruvoyl-tetrahydropterin synthase, PTS (chromosome 11), with P = 0.009; this result was not restricted to an affected male-only subset. Multilocus interaction was detected in the BH4 pathway alone, but not across the serotonin, dopamine and BH4 pathways.

Examination of association of genes in the serotonin system to autism.

Autism is characterized as one of the pervasive developmental disorders, a spectrum of often severe behavioral and cognitive disturbances of early development. The high heritability of autism has driven multiple efforts to identify genetic variation that increases autism susceptibility. Numerous studies have suggested that variation in peripheral and central metabolism of serotonin (5-hydroxytryptamine) may play a role in the pathophysiology of autism. We screened 403 autism families for 45 single nucleotide polymorphisms in ten serotonin pathway candidate genes. Although genome-wide linkage scans in autism have provided support for linkage to various loci located within the serotonin pathway, our study does not provide strong evidence for linkage to any specific gene within the pathway. The most significant association (p = 0.0002; p = 0.02 after correcting for multiple comparisons) was found at rs1150220 (HTR3A) located on chromosome 11 ( approximately 113 Mb). To test specifically for multilocus effects, multifactor dimensionality reduction was employed, and a significant two-way interaction (p value = 0.01) was found between rs10830962, near MTNR1B (chromosome11; 92,338,075 bp), and rs1007631, near SLC7A5 (chromosome16; 86,413,596 bp). These data suggest that variation within genes on the serotonin pathway, particularly HTR3A, may have modest effects on autism risk.

Novel congenital myopathy locus identified in Native American Indians at 12q13.13-14.1.

BACKGROUND: Native American myopathy (NAM) is an autosomal recessive congenital myopathy first reported in the Lumbee Indian people. Features of NAM include congenital weakness, cleft palate, ptosis, short stature, and susceptibility to malignant hyperthermia provoked by anesthesia. METHOD: We identified five individuals with NAM from the Lumbee population, and hypothesized that these affected individuals have disease alleles shared identical-by-descent inherited from common ancestry. To identify a NAM disease locus, homozygosity mapping methods were employed on a genomewide 10K single-nucleotide polymorphism (SNP) screen. To confirm regions of homozygosity identified in the SNP screen, microsatellite repeat markers were genotyped within those homozygous segments. RESULTS: The SNP data demonstrated five regions of shared homozygosity in individuals with NAM. The additional genotyping data narrowed the region to one common segment of homozygosity spanning D12S398 to rs3842936 mapping to 12q13.13-14.1. Notably, loss of heterozygosity estimates from the SNP data also detected this same 12q region in the affected individuals. CONCLUSION: This study reports the first gene mapping of Native American myopathy (NAM) using single-nucleotide polymorphism-based homozygosity mapping in only a few affected individuals from simplex families and identified a novel NAM locus. Identifying the genetic basis of NAM may suggest new genetic etiologies for other more common conditions such as congenital myopathy and malignant hyperthermia.

Linkage and association study of late-onset Alzheimer disease families linked to 9p21.3.

A chromosomal locus for late-onset Alzheimer disease (LOAD) has previously been mapped to 9p21.3. The most significant results were reported in a sample of autopsy-confirmed families. Linkage to this locus has been independently confirmed in AD families from a consanguineous Israeli-Arab community. In the present study we analyzed an expanded clinical sample of 674 late-onset AD families, independently ascertained by three different consortia. Sample subsets were stratified by site and autopsy-confirmation. Linkage analysis of a dense array of SNPs across the chromosomal locus revealed the most significant results in the 166 autopsy-confirmed families of the NIMH sample. Peak HLOD scores of 4.95 at D9S741 and 2.81 at the nearby SNP rs2772677 were obtained in a dominant model. The linked region included the cyclin-dependent kinase inhibitor 2A gene (CDKN2A), which has been suggested as an AD candidate gene. By re-sequencing all exons in the vicinity of CDKN2A in 48 AD cases, we identified and genotyped four novel SNPs, including a non-synonymous, a synonymous, and two variations located in untranslated RNA sequences. Family-based allelic and genotypic association analysis yielded significant results in CDKN2A (rs11515: PDT p = 0.003, genotype-PDT p = 0.014). We conclude that CDKN2A is a promising new candidate gene potentially contributing to AD susceptibility on chromosome 9p.

Examination of association to autism of common genetic variationin genes related to dopamine.

Autism is a severe neurodevelopmental disorder characterized by a triad of complications. Autistic individuals display significant disturbances in language and reciprocal social interactions, combined with repetitive and stereotypic behaviors. Prevalence studies suggest that autism is more common than originally believed, with recent estimates citing a rate of one in 150. Although multiple genetic linkage and association studies have yielded multiple suggestive genes or chromosomal regions, a specific risk locus has yet to be identified and widely confirmed. Because many etiologies have been suggested for this complex syndrome, we hypothesize that one of the difficulties in identifying autism genes is that multiple genetic variants may be required to significantly increase the risk of developing autism. Thus, we took the alternative approach of examining 14 prominent dopamine pathway candidate genes for detailed study by genotyping 28 single nucleotide polymorphisms. Although we did observe a nominally significant association for rs2239535 (P=0.008) on chromosome 20, single-locus analysis did not reveal any results as significant after correction for multiple comparisons. No significant interaction was identified when Multifactor Dimensionality Reduction was employed to test specifically for multilocus effects. Although genome-wide linkage scans in autism have provided support for linkage to various loci along the dopamine pathway, our study does not provide strong evidence of linkage or association to any specific gene or combination of genes within the pathway. These results demonstrate that common genetic variation within the tested genes located within this pathway at most play a minor to moderate role in overall autism pathogenesis.

Genotype-phenotype study in an FSHD family with a proximal deletion encompassing p13E-11 and D4Z4.

BACKGROUND: In the majority of facioscapulohumeral muscular dystrophy (FSHD) cases, the molecular basis of the disease is due to loss of subtelomeric D4Z4 repeat units at 4q35. Occasionally, an apparent absence of the contracted D4Z4 repeat is associated with FSHD. One explanation for this finding is a deletion in the region proximal to the D4Z4 repeat array that encompasses the p13E-11 (D4F104S1) probe-binding site used in the DNA diagnosis. The frequency of such proximally extended deletions is unknown, and to date, few patients have been described due to the difficulties in the molecular identification of such cases. METHODS: We describe a family (DUK 2531) in which a contracted D4Z4 allele and a large proximal deletion of approximately 75 kb are segregating to 11 individuals. This is the largest deletion identified to date. Family DUK 2531 was initially thought to have normal D4Z4 fragment size and therefore unlinked to the 4q35 region (FSHD1B). RESULTS: Further molecular analysis of DUK 2531 reveals the presence of 10 repeat units (33 kb). The extended deletion includes the probe p13E-11 and B31 binding sites, the inverted repeat D4S2463, and genes FRG2 and TUBB4Q. CONCLUSION: Despite the length of the proximal deletion in this family, the range and severity of the clinical manifestations are typical for the disorder. Because such deletions can lead to misinterpretation in the diagnostic setting, this suggests the need for additional diagnostic tests in facioscapulohumeral muscular dystrophy.

Dissecting the locus heterogeneity of autism: significant linkage to chromosome 12q14.

Autism is a common neurodevelopmental disorder with a significant genetic component and locus heterogeneity. To date, 12 microsatellite genome screens have been performed using various data sets of sib-pair families (parents and affected children) resulting in numerous regions of potential linkage across the genome. However, no universal region or consistent candidate gene from these regions has emerged. The use of large, extended pedigrees is a recognized powerful approach to identify significant linkage results, as these families potentially contain more potential linkage information than sib-pair families. A genome-wide linkage analysis was performed on 26 extended autism families (65 affected, 184 total individuals). Each family had two to four affected individuals comprised of either avuncular or cousin pairs. For analysis, we used a high-density single-nucleotide polymorphism genotyping assay, the Affymetrix GeneChip Human Mapping 10K array. Two-point analysis gave peak heterogeneity limit of detection (HLOD) of 2.82 at rs2877739 on chromosome 14q. Suggestive linkage evidence (HLOD>2) from a two-point analysis was also found on chromosomes 1q, 2q, 5q, 6p,11q and 12q. Chromosome 12q was the only region showing significant linkage evidence by multipoint analysis with a peak HLOD=3.02 at rs1445442. In addition, this linkage evidence was enhanced significantly in the families with only male affected (multipoint HLOD=4.51), suggesting a significant gender-specific effect in the etiology of autism. Chromosome-wide haplotype analyses on chromosome 12 localized the potential autism gene to a 4 cM region shared among the affected individuals across linked families. This novel linkage peak on chromosome 12q further supports the hypothesis of substantial locus heterogeneity in autism.

Exploring the association of glyceraldehyde-3-phosphate dehydrogenase gene and Alzheimer disease.

BACKGROUND: Previous linkage studies have shown that chromosome 12 harbors susceptibility genes for late-onset Alzheimer disease (LOAD). However, association studies of several candidate genes on this chromosome region have produced ambiguous results. A recent study reported the association between the glyceraldehyde-3-phosphate dehydrogenase (GAPD) gene on chromosome 12p and the risk of LOAD. METHODS: The authors conducted family-based and case-control association studies in two independent LOAD data sets on 12 single-nucleotide polymorphisms (SNPs) in the GAPD gene and its paralogs. RESULTS: No association was found of the GAPD gene with LOAD in the family-based data set, but marginal evidence of association was seen in the later-onset subgroup when age at onset was stratified. The SNP rs2029721 in one GAPD pseudogene was also found to be associated with risk for LOAD in the unrelated case-control data set (p = 0.003). CONCLUSIONS: The GAPD gene and its pseudogene may play a role in the development of late-onset Alzheimer disease. However, the effect, if any, is likely to be limited.

An analysis paradigm for investigating multi-locus effects in complex disease: examination of three GABA receptor subunit genes on 15q11-q13 as risk factors for autistic disorder.

Gene-gene interactions are likely involved in many complex genetic disorders and new statistical approaches for detecting such interactions are needed. We propose a multi-analytic paradigm, relying on convergence of evidence across multiple analysis tools. Our paradigm tests for main and interactive effects, through allele, genotype and haplotype association. We applied our paradigm to genotype data from three GABAA receptor subunit genes (GABRB3, GABRA5, and GABRG3) on chromosome 15 in 470 Caucasian autism families. Previously implicated in autism, we hypothesized these genes interact to contribute to risk. We detected no evidence of main effects by allelic (PDT, FBAT) or genotypic (genotype-PDT) association at individual markers. However, three two-marker haplotypes in GABRG3 were significant (HBAT). We detected no significant multi-locus associations using genotype-PDT analysis or the EMDR data reduction program. However, consistent with the haplotype findings, the best single locus EMDR model selected a GABRG3 marker. Further, the best pairwise genotype-PDT result involved GABRB3 and GABRG3, and all multi-locus EMDR models also selected GABRB3 and GABRG3 markers. GABA receptor subunit genes do not significantly interact to contribute to autism risk in our overall data set. However, the consistency of results across analyses suggests that we have defined a useful framework for evaluating gene-gene interactions.

Lack of association between UBQLN1 and Alzheimer disease.

Alzheimer disease (AD) is heterogeneous and complex with a strong genetic diathesis. It is the most common cause of dementia affecting the elderly. Linkage studies [Kehoe et al., 1999; Hum Mol Genet 8: 237-245]; [Pericak-Vance et al., 2000; Exp Gerontol 35: 1343-1352]; [Myers et al., 2002; Am J Med Genet 114: 235-244]; [Blacker et al., 2003; Hum Mol Genet 12: 23-32] identified chromosome 9q as a region containing a possible AD candidate gene. Functional protein studies [Mah et al., 2000; J Cell Biol 151: 847-862]; [Ko et al., 2002; J Biol Chem 277: 35386-35392] identified the UBQLN1 gene on chromosome 9q that encodes ubiquilin as a likely candidate for a role in late-onset AD pathogenesis. A recent family-based study by [Bertram et al., 2005; N Engl J 352: 884-894] reported genetic association and expression evidence for a putative AD risk allele of an intronic single nucleotide polymorphism (SNP) within the UBQLN1 gene. In this study, we comprehensively assessed whether any of seven polymorphisms located across the UBQLN1 gene are associated with AD in another large family-based data set and an independent case-control data set. We found no significant association of AD risk with any of the seven SNPs genotyped (including those SNPs previously reported by Bertram et al.) in either the family-based or case-control data set. Age-specific analyses and analyses conditional on Apolipoprotein E (ApoE) genotype and sex also revealed no significant associations to AD risk in either data set. Using age at onset (AAO) as a quantitative trait revealed a modest age modifying association; however, the results were inconsistent between the data sets. Our results suggest that UBQLN1 variants do not increase risk for AD in these data.

Covariate analysis of late-onset Alzheimer disease refines the chromosome 12 locus.

Alzheimer disease (AD) is a progressive neurodegenerative disorder of later life with a complex etiology and a strong genetic component. Several genomic screens have suggested that a region between chromosome 12p13 and 12q22 contains at least one additional locus underlying the susceptibility of AD. However, localization of this locus has been difficult. We performed a 5 cM microsatellite marker screen across 74 cM on chromosome 12 with 15 markers in 585 multiplex families consisting of 994 affected sibpairs and 213 other affected relative pairs. Analyses across the entire data set did not reveal significant evidence of linkage. However, suggestive linkage was observed in several subsets. In the 91 families where no affected individuals carry an ApoE varepsilon4 allele, an HLOD score of 1.55 was generated at D12S1042. We further examined the linkage data considering the proposed linkages to chromosome 9 (D9S741) and chromosome 10 (alpha-catenin gene). There was a modest (P=0.20) increase in the LOD score for D12S368 (MLOD=1.70) when using the D9S741 LOD scores as a covariate and a highly significant (P<0.001) increase in the MLOD score (4.19) for D12S1701 in autopsy-confirmed families (n=228) when using alpha-catenin LOD scores as a covariate. In both cases, families with no evidence of linkage to D9S741 or alpha-catenin demonstrated most of the evidence of linkage to chromosome 12, suggesting locus heterogeneity. Taken together, our data suggest that the 16 cM region between D12S1042 and D12S368 should be the subject of further detailed genomic efforts for the disease.

Interaction between the alpha-T catenin gene (VR22) and APOE in Alzheimer's disease.

BACKGROUND: APOE is the only gene that has been consistently replicated as a risk factor for late onset Alzheimer's disease. Several recent studies have identified linkage to chromosome 10 for both risk and age of onset, suggesting that this region harbours genes that influence the development of the disease. A recent study reported association between single nucleotide polymorphisms (SNPs) in the VR22 gene (CTNNA3) on chromosome 10 and plasma levels of Abeta42, an endophenotype related to Alzheimer's disease. OBJECTIVE: To assess whether polymorphisms in the VR22 gene are associated with Alzheimer's disease in a large sample of Alzheimer's disease families and an independent set of unrelated cases and controls. RESULTS: Several SNPs showed association in either the family based or case-control analyses (p<0.05). The most consistent findings were with SNP6, for which there was significant evidence of association in both the families and the unrelated cases and controls. Furthermore, there was evidence of significant interaction between APOE-4 and two of the VR22 SNPs, with the strongest evidence of association being concentrated in individuals carrying APOE-4. CONCLUSIONS: This study suggests that VR22 or a nearby gene influences susceptibility to Alzheimer's disease, and the effect is dependent on APOE status.

Identification of significant association and gene-gene interaction of GABA receptor subunit genes in autism.

Autism is a common neurodevelopmental disorder with a significant genetic component. Existing research suggests that multiple genes contribute to autism and that epigenetic effects or gene-gene interactions are likely contributors to autism risk. However, these effects have not yet been identified. Gamma-aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the adult brain, has been implicated in autism etiology. Fourteen known autosomal GABA receptor subunit genes were studied to look for the genes associated with autism and their possible interactions. Single-nucleotide polymorphisms (SNPs) were screened in the following genes: GABRG1, GABRA2, GABRA4, and GABRB1 on chromosome 4p12; GABRB2, GABRA6, GABRA1, GABRG2, and GABRP on 5q34-q35.1; GABRR1 and GABRR2 on 6q15; and GABRA5, GABRB3, and GABRG3 on 15q12. Intronic and/or silent mutation SNPs within each gene were analyzed in 470 white families with autism. Initially, SNPs were used in a family-based study for allelic association analysis--with the pedigree disequilibrium test and the family-based association test--and for genotypic and haplotypic association analysis--with the genotype-pedigree disequilibrium test (geno-PDT), the association in the presence of linkage (APL) test, and the haplotype family-based association test. Next, with the use of five refined independent marker sets, extended multifactor-dimensionality reduction (EMDR) analysis was employed to identify the models with locus joint effects, and interaction was further verified by conditional logistic regression. Significant allelic association was found for markers RS1912960 (in GABRA4; P = .01) and HCV9866022 (in GABRR2; P = .04). The geno-PDT found significant genotypic association for HCV8262334 (in GABRA2), RS1912960 and RS2280073 (in GABRA4), and RS2617503 and RS12187676 (in GABRB2). Consistent with the allelic and genotypic association results, EMDR confirmed the main effect at RS1912960 (in GABRA4). EMDR also identified a significant two-locus gene-gene effect model involving RS1912960 in GABRA4 and RS2351299 in GABRB1. Further support for this two-locus model came from both the multilocus geno-PDT and the APL test, which indicated a common genotype and haplotype combination positively associated with disease. Finally, these results were also consistent with the results from the conditional logistic regression, which confirmed the interaction between GABRA4 and GABRB1 (odds ratio = 2.9 for interaction term; P = .002). Through the convergence of all analyses, we conclude that GABRA4 is involved in the etiology of autism and potentially increases autism risk through interaction with GABRB1. These results support the hypothesis that GABA receptor subunit genes are involved in autism, most likely via complex gene-gene interactions.

Whole genomewide linkage screen for neural tube defects reveals regions of interest on chromosomes 7 and 10.

Neural tube defects (NTDs) are the second most common birth defects (1 in 1000 live births) in the world. Periconceptional maternal folate supplementation reduces NTD risk by 50-70%; however, studies of folate related and other developmental genes in humans have failed to definitively identify a major causal gene for NTD. The aetiology of NTDs remains unknown and both genetic and environmental factors are implicated. We present findings from a microsatellite based screen of 44 multiplex pedigrees ascertained through the NTD Collaborative Group. For the linkage analysis, we defined our phenotype narrowly by considering individuals with a lumbosacral level myelomeningocele as affected, then we expanded the phenotype to include all types of NTDs. Two point parametric analyses were performed using VITESSE and HOMOG. Multipoint parametric and nonparametric analyses were performed using ALLEGRO. Initial results identified chromosomes 7 and 10, both with maximum parametric multipoint lod scores (Mlod) >2.0. Chromosome 7 produced the highest score in the 24 cM interval between D7S3056 and D7S3051 (parametric Mlod 2.45; nonparametric Mlod 1.89). Further investigation demonstrated that results on chromosome 7 were being primarily driven by a single large pedigree (parametric Mlod 2.40). When this family was removed from analysis, chromosome 10 was the most interesting region, with a peak Mlod of 2.25 at D10S1731. Based on mouse human synteny, two candidate genes (Meox2, Twist1) were identified on chromosome 7. A review of public databases revealed three biologically plausible candidates (FGFR2, GFRA1, Pax2) on chromosome 10. The results from this screen provide valuable positional data for prioritisation of candidate gene assessment in future studies of NTDs.

The genes encoding for D4Z4 binding proteins HMGB2, YY1, NCL, and MYOD1 are excluded as candidate genes for FSHD1B.

Facioscapulohumeral muscular dystrophy is a disease of skeletal muscle, with symptoms including both facial and shoulder girdle weakness and progression to involve the pelvic girdle and extremities in the majority of cases. For most cases of FSHD, the molecular basis of the disease can be identified as a partial deletion of the D4Z4 repeat array on the end of the long arm of chromosome 4. However, in up to 5% of FSHD families there is no linkage to 4q35. These cases are designated as FSHD1B. Proteins have been identified that bind to the D4Z4 repeats of chromosome 4q35. The genes encoding D4Z4 binding proteins YY1, HMGB2, NCL, and MYOD1 were investigated as candidate genes for FSHD1B. Coding sequences and promoter region were analyzed for HMBG2 and no sequence variations were detected. For YY1, all five exons were analyzed and a polymorphism was detected in both the unaffected and affected populations. In nucleolin (NCL), several SNPs were identified, including a SNP causing the non-synonymous change P515H; however, all polymorphisms either occurred in control samples or were previously reported. A novel polymorphism was also detected in MYOD1, but did not represent a disease-specific variation. These results suggest that HMBG2, YY1, NCL, and MYOD1 are unlikely to represent the genes responsible for FSHD in these families.