Evidence for Three Novel QTLs for Adiposity on Chromosome 2 With Epistatic Interactions: The NHLBI Family Heart Study

We sought to identify quantitative trait loci (QTLs) by genome‐wide linkage analysis for BMI and waist circumference (WC) exploring various strategies to address heterogeneity including covariate adjustments and complex models based on epistatic components of variance. Because cholesterol‐lowering drugs and diabetes medications may affect adiposity and risk of coronary heart disease, we excluded subjects medicated for hypercholesterolemia and hyperglycemia. The evidence of linkage increased on 2p25 (BMI: lod = 1.59 vs. 2.43, WC: lod = 1.32 vs. 2.26). Because environmental and/or genetic components could mask the effect of a specific locus, we investigated further whether a QTL could influence adiposity independently of lipid pathway and dietary habits. Strong evidence of linkage on 2p25 (BMI: lod = 4.31; WC: lod = 4.23) was found using Willet's dietary factors and lipid profile together with age and sex in adjustment. It suggests that lipid profile and dietary habits are confounding factors for detecting a 2p25 QTL for adiposity. Because evidence of linkage has been previously detected for BMI on 7q34 and 13q14 in National Heart, Lung, and Blood Institute Family Heart Study (NHLBI FHS), and for diabetes on 15q13, we investigated epistasis between chromosome 2 and these loci. Significant epistatic interactions were found between QTLs 2p25 and 7q34, 2q37 and 7q34, 2q31 and 13q14, and 2q31–q36 and 15q13. These results suggest multiple pathways and factors involving genetic and environmental effects influencing adiposity. By taking some of these known factors into account, we clarified our linkage evidence of a QTL on 2p25 influencing BMI and WC. The 2p25, 2q24–q31, and 2q36–q37 showed evidence of epistatic interaction with 7q34, 13q14, and 15q13.     

A QTL on 12q Influencing an Inflammation Marker and Obesity in White Women: The NHLBI Family Heart Study

It has been recognized that obese individuals are intrinsically in a state of chronic inflammation, as indicated by positive correlations between serum levels of C‐reactive protein (CRP) and various anthropometric measures of obesity. To explore the hypothesis that a gene(s) may underlie this relationship, we conducted bivariate linkage analyses of BMI and CRP in white and African‐American (AA) families of the National Heart, Lung, and Blood Institute (NHLBI) Family Heart Study (FHS). Variance components linkage analysis as implemented in SOLAR was performed in 1,825 whites (840 men and 985 women) and 548 AAs (199 men and 351 women). CRP exhibited significant genetic correlations with BMI in women (0.54 ± 0.10 for white and 0.53 ± 0.14 for AA) and the combined samples (0.37 ± 0.09 for white and 0.56 ± 0.13 for AA), but not in men. We detected a maximum bivariate lod score of 3.86 on chromosome 12q24.2–24.3 at 139 cM and a suggestive linkage signal (lod = 2.19) on chromosome 19p13.1 (44 cM) in white women. Both bivariate peaks were substantially higher than their respective univariate lods at the same locus for each trait. No significant lod scores were detected in AAs. Our results indicate that chromosome 12q may harbor quantitative trait loci (QTLs) jointly regulating BMI and CRP in white women.     

A YAC Contig of Approximately 3 Mb from Human Chromosome 5q31 → q33

The human chromosome 5q31-q33 region contains an interesting cluster of growth factor and receptor genes. In addition, several genetic disease loci have been localized within this region, but have not as yet been isolated as molecular clones. These include those loci involved in autosomal dominant limb-girdle muscular dystrophy, diastrophic dysplasia. Treacher Collins syndrome, and myeloid disorders associated with the 5q- syndrome. A yeast artificial chromosome (YAC) contig of this region would assist in the further localization and isolation of these genes. We have used YACs isolated from the Washington University and Centre d'Etude du Polymorphisme Humain YAC libraries, including YACs from the large insert (mega) YAC library to build a contig greater than 3 Mb in size. An STS content strategy coupled with limited walking from YAC ends was used to isolate 22 overlapping YACs with as much as sixfold coverage. A total of 20 STSs, derived from genes, anonymous sequences, and vector Alu-PCR or inverse PCR products, were used to compile this contig. The order of loci, centromere-GRL-D5S207-D5S70-D5S545-D5S546-D5S547-D5S68-D5S548-D5S210-D5S549-D5S686-ADRB2-D5S559-CSF1R-D5S551-RPS14-D5S519-SPARC-telomere, was derived from the overlapping clones. This contig and clones derived from it will be useful substrates in selecting candidate cDNAs for the disease loci in this interval.     

Neuropeptide Y Receptor Genes on Human Chromosome 4q31–q32 Map to Conserved Linkage Groups on Mouse Chromosomes 3 and 8

Npy1randNpy2r,the genes encoding mouse type 1 and type 2 neuropeptide Y receptors, have been mapped by interspecific backcross analysis. Previous studies have localized the human genes encoding these receptors to chromosome 4q31–q32. We have now assignedNpy1randNpy2rto conserved linkage groups on mouse Chr 8 and Chr 3, respectively, which correspond to the distal region of human chromosome 4q. Using yeast artificial chromosomes, we have estimated the distance between the human genes to be approximately 6 cM. Although ancient tandem duplication events may account for some closely spaced G-protein-coupled receptor genes, the large genetic distance between the human type 1 and type 2 neuropeptide Y receptor genes raises questions about whether this mechanism accounts for their proximity.     

Genome-wide linkage analysis replicates susceptibility locus for fasting plasma triglycerides: NHLBI Family Heart Study

Recent reports implicate chromosomal regions linked to inter-individual variation in plasma triglycerides. We conducted genome-wide scans to replicate these linkages and/or identify other loci influencing plasma triglycerides in the NHLBI Family Heart Study (FHS). Data were obtained for 501 three-generational families. Genotyping was done by the Utah Molecular Genetics Laboratory and NHLBI Mammalian Genotyping Service; markers from both were placed on one genetic map. Analysis was done using multipoint variance components linkage. Fasting plasma triglycerides were log-transformed and age-, sex-, and field center-adjusted; suggestive linkage evidence was found on chromosome 8 (LOD=2.80 at 89 cM, marker D8S1141). Further adjustment for waist girth, BMI, diabetes, hypertension, and lipid-lowering drugs suggested linkage regions on chromosomes 6 (LOD=2.29 at 79 cM, marker D6S295) and 15 (LOD=1.85 at 43 cM, marker D15S659). Since HDL is correlated with triglycerides and because it was linked to this region on chromosome 15 in FHS, we created a composite triglyceride–HDL phenotype. The combined phenotype LOD score was 3.0 at the same marker on chromosome 15. Chromosome 15 likely harbors a susceptibility locus with an influence on triglycerides and HDL. Regions on chromosomes 6 and 8 may also contain loci contributing to inter-individual variation in plasma triglycerides.     

New Genes on Human Chromosome 7: Bioinformation Analysis of a Cluster of Genes from the POLR2J Family

Four independent genes encoding various variants of the hRPB11 subunit of Homo sapiens RNA polymerase II were revealed in human chromosome 7. Three genes (POLR2 J1, POLR2 J2, and POLR2 J3) form a cluster of total length of 214 530 bp in the genetic locus 7q22.1 on the long arm of chromosome 7 (contig NT_007933). The fourth gene (POLR2 J4, 31 040 bp) was localized in the cytogenetic locus 7p13 of the short arm of chromosome 7 (contig NT_007819). An analysis enabled us to refine dissimilar experimental data on the mapping of the hRPB11 subunit gene on chromosome 7. In particular, the presence of three sites of its localization according to data on hybridization with fluorescent-labeled probes (the FISH method) was explained. It was established that, upon the expression of the four human POLR2 J genes, at least 14 types of mature mRNAs encoding somewhat differing hRPB11 isoforms can be synthesized. Eleven of these mRNAs were revealed (as full-length copies or clearly identifiable fragments) in the available databases of expressed sequence tags and cDNAs. The most probable scheme of origination of the multiple genes of the POLR2 J family as a result of three consecutive segmented duplications increasing in size was proposed and substantiated. On the basis of the scheme, some assumptions on the pathways of evolution of separate human genes and the mechanisms of generation of protein diversity in higher eukaryotes were made.     

Cloning of the Human Monocarboxylate Transporter MCT3 Gene: Localization to Chromosome 22q12.3–q13.2

Lactate transport across cell membranes is mediated by a family of proton-coupled monocarboxylate transporters (MCTs). The retinal pigment epithelium (RPE) expresses a unique member of this family, MCT3. A portion of the human MCT3 gene was cloned by polymerase chain reaction using primers designed from rat RPE MCT3 cDNA sequence. The human genomic sequence was used to design primers to clone human MCT3 cDNA and to identify a bacterial artificial chromosome clone containing the human MCT3 gene. The human MCT3 cDNA contained a 1512-nucleotide open reading frame with a deduced amino sequence 85% identical to rat MCT3. Comparison of the cDNA and genomic sequences revealed that the MCT3 gene was composed of five exons distributed over 5 kb of DNA. The exon–intron borders were conserved between the human and the chicken MCT3 genes. Using radiation hybrid mapping, the MCT3 gene was mapped to chromosome 22 between markers WI11639 and SGC30687. A search of chromosome 22 in the Sanger Centre database confirmed the location of the human MCT3 gene at 22q12.3–q13.2.     

A Cluster of Three Novel Ca Channel γ Subunit Genes on Chromosome 19q13.4: Evolution and Expression Profile of the γ Subunit Gene Family

The CACNG1 gene on chromosome 17q24 encodes an integral membrane protein that was originally isolated as the regulatory γ subunit of voltage-dependent Ca²⁺ channels from skeletal muscle. The existence of an extended family of γ subunits was subsequently demonstrated upon identification of CACNG2 (22q13), CACNG3 (16p12–p13), and CACNG4 and CACNG5 (17q24). In this study, we describe a cluster of three novel γ subunit genes, CACNG6, CACNG7, and CACNG8, located in a tandem array on 19q13.4. Phylogenetic analysis indicates that this array is paralogous to the cluster containing CACNG1, CACNG5, and CACNG4, respectively, on chromosome 17q24. We developed sensitive RT-PCR assays and examined the expression profile of each member of the γ subunit gene family, CACNG1–CACNG8. Analysis of 24 human tissues plus 3 dissected brain regions revealed that CACNG1 through CACNG8 are all coexpressed in fetal and adult brain and differentially transcribed among a wide variety of other tissues. The expression of distinct complements of γ subunit isoforms in different cell types may be an important mechanism for regulating Ca²⁺ channel function.     

Autistic Disorder and Chromosome 15q11–q13: Construction and Analysis of a BAC/PAC Contig

Autistic disorder (AD) is a neurodevelopmental disorder that affects approximately 2–10/10,000 individuals. Chromosome 15q11–q13 has been implicated in the genetic etiology of AD based on (1) cytogenetic abnormalities; (2) increased recombination frequency in this region in AD versus non-AD families; (3) suggested linkage with markers D15S156, D15S219, and D15S217; and (4) evidence for significant association with polymorphisms in the γ-aminobutyric acid receptor subunit B3 gene (GABRB3). To isolate the putative 15q11–q13 candidate AD gene, a genomic contig and physical map of the approximately 1.2-Mb region from the GABA receptor gene cluster to the OCA2 locus was generated. Twenty-one bacterial artificial chromosome (BAC) clones, 32 P1-derived artificial chromosome (PAC) clones, and 2 P1 clones have been isolated using the markers D15S540, GABRB3, GABRA5, GABRG3, D15S822, and D15S217, as well as 34 novel markers developed from the end sequences of BAC/PAC clones. In contrast to previous findings, the markers D15S822 and D15S975 have been localized within the GABRG3 gene, which we have shown to be approximately 250 kb in size. NotI and numerous EagI restriction enzyme cut sites were identified in this region. The BAC/PAC genomic contig can be utilized for the study of genomic structure and the identification and characterization of genes and their methylation status in this autism candidate gene region on human chromosome 15q11–q13.     

Characterization of Three Novel Human Cadherin Genes (CDH7, CDH19, and CDH20) Clustered on Chromosome 18q22–q23 and with High Homology to Chicken Cadherin-7

Full-length coding sequences of two novel human cadherin cDNAs were obtained by sequence analysis of several EST clones and 5′ and 3′ rapid amplification of cDNA ends (RACE) products. Exons for a third cDNA sequence were identified in a public-domain human genomic sequence, and the coding sequence was completed by 3′ RACE. One of the sequences (CDH7L1, HGMW-approved gene symbol CDH7) is so similar to chicken cadherin-7 gene that we consider it to be the human orthologue. In contrast, the published partial sequence of human cadherin-7 is identical to our second cadherin sequence (CDH7L2), for which we propose CDH19 as the new name. The third sequence (CDH7L3, HGMW-approved gene symbol CDH20) is almost identical to the mouse “cadherin-7” cDNA. According to phylogenetic analysis, this mouse cadherin-7 and its here presented human homologue are most likely the orthologues of Xenopus F-cadherin. These novel human genes, CDH7, CDH19, and CDH20, are localized on chromosome 18q22–q23, distal of both the gene CDH2 (18q11) encoding N-cadherin and the locus of the six desmosomal cadherin genes (18q12). Based on genetic linkage maps, this genomic region is close to the region to which Paget's disease was linked. Interestingly, the expression patterns of these three closely related cadherins are strikingly different.     

A novel obesity locus on chromosome 4q: the Strong Heart Family Study

Objective: Obesity is a growing and important public health problem in Western countries and worldwide. There is ample evidence that both environmental and genetic factors influence the risk of developing obesity. Although a number of genes influencing obesity and obesity‐related measures have been localized, it is clear that others remain to be identified. The rate of obesity is particularly high in American Indian populations. This study reports the results of a genome‐wide scan for loci influencing BMI and weight in 963 individuals in 58 families from three American Indian populations in Arizona, Oklahoma, and North and South Dakota participating in the Strong Heart Family Study. Research Methods and Procedures: Short tandem repeat markers were genotyped, resulting in a marker map with an average spacing of 10 centimorgans. Standard multipoint variance component linkage methods were used. Results: Significant evidence of linkage was observed in the overall sample, including all three study sites, for a locus on chromosome 4q35 [logarithm of the odds (LOD) = 5.17 for weight, 5.08 for BMI]. Analyses of the three study sites individually showed that the greatest linkage support for the chromosome 4 locus came from Arizona (LOD = 2.6 for BMI), but that LOD scores for weight were >1 in all three samples. Suggestive linkage signals (LOD >2) were also observed on chromosomes 5, 7, 8, and 10. Discussion: The chromosome 4 locus detected in this scan is in a region lacking any obvious positional candidate genes with known functions related to obesity. This locus may represent a novel obesity gene.