Genome-wide association data suggest ABCB1 and immune-related gene sets may be involved in adult antisocial behavior.

Adult antisocial behavior (AAB) is moderately heritable, relatively common and has adverse consequences for individuals and society. We examined the molecular genetic basis of AAB in 1379 participants from a case-control study in which the cases met criteria for alcohol dependence. We also examined whether genes of interest were expressed in human brain. AAB was measured using a count of the number of Antisocial Personality Disorder criteria endorsed under criterion A from the Diagnostic and Statistical Manual of Mental Disorders, 4th Edition (DSM-IV). Participants were genotyped on the Illumina Human 1M BeadChip. In total, all single-nucleotide polymorphisms (SNPs) accounted for 25% of the variance in AAB, although this estimate was not significant (P=0.09). Enrichment tests indicated that more significantly associated genes were over-represented in seven gene sets, and most were immune related. Our most highly associated SNP (rs4728702, P=5.77 × 10(-7)) was located in the protein-coding adenosine triphosphate-binding cassette, sub-family B (MDR/TAP), member 1 (ABCB1). In a gene-based test, ABCB1 was genome-wide significant (q=0.03). Expression analyses indicated that ABCB1 was robustly expressed in the brain. ABCB1 has been implicated in substance use, and in post hoc tests we found that variation in ABCB1 was associated with DSM-IV alcohol and cocaine dependence criterion counts. These results suggest that ABCB1 may confer risk across externalizing behaviors, and are consistent with previous suggestions that immune pathways are associated with externalizing behaviors. The results should be tempered by the fact that we did not replicate the associations for ABCB1 or the gene sets in a less-affected independent sample.

A genome-wide association study of alcohol-dependence symptom counts in extended pedigrees identifies C15orf53.

Several studies have identified genes associated with alcohol-use disorders (AUDs), but the variation in each of these genes explains only a small portion of the genetic vulnerability. The goal of the present study was to perform a genome-wide association study (GWAS) in extended families from the Collaborative Study on the Genetics of Alcoholism to identify novel genes affecting risk for alcohol dependence (AD). To maximize the power of the extended family design, we used a quantitative endophenotype, measured in all individuals: number of alcohol-dependence symptoms endorsed (symptom count (SC)). Secondary analyses were performed to determine if the single nucleotide polymorphisms (SNPs) associated with SC were also associated with the dichotomous phenotype, DSM-IV AD. This family-based GWAS identified SNPs in C15orf53 that are strongly associated with DSM-IV alcohol-dependence symptom counts (P=4.5 × 10(-8), inflation-corrected P=9.4 × 10(-7)). Results with DSM-IV AD in the regions of interest support our findings with SC, although the associations were less significant. Attempted replications of the most promising association results were conducted in two independent samples: nonoverlapping subjects from the Study of Addiction: Genes and Environment (SAGE) and the Australian Twin Family Study of AUDs (OZALC). Nominal association of C15orf53 with SC was observed in SAGE. The variant that showed strongest association with SC, rs12912251 and its highly correlated variants (D'=1, r(2) 0.95), have previously been associated with risk for bipolar disorder.

Genetic variation in the CHRNA5 gene affects mRNA levels and is associated with risk for alcohol dependence.

Alcohol dependence frequently co-occurs with cigarette smoking, another common addictive behavior. Evidence from genetic studies demonstrates that alcohol dependence and smoking cluster in families and have shared genetic vulnerability. Recently a candidate gene study in nicotine dependent cases and nondependent smoking controls reported strong associations between a missense mutation (rs16969968) in exon 5 of the CHRNA5 gene and a variant in the 3'-UTR of the CHRNA3 gene and nicotine dependence. In this study we performed a comprehensive association analysis of the CHRNA5-CHRNA3-CHRNB4 gene cluster in the Collaborative Study on the Genetics of Alcoholism (COGA) families to investigate the role of genetic variants in risk for alcohol dependence. Using the family-based association test, we observed that a different group of polymorphisms, spanning CHRNA5-CHRNA3, demonstrate association with alcohol dependence defined by Diagnostic and Statistical Manual of Mental Disorders, 4th edn (DSM-IV) criteria. Using logistic regression we replicated this finding in an independent case-control series from the family study of cocaine dependence. These variants show low linkage disequilibrium with the SNPs previously reported to be associated with nicotine dependence and therefore represent an independent observation. Functional studies in human brain reveal that the variants associated with alcohol dependence are also associated with altered steady-state levels of CHRNA5 mRNA.

Association studies of ALOX5 and bone mineral density in healthy adults.

UNLABELLED: Animal studies suggest that arachidonate 5-lipoxygenase (encoded by ALOX5) may be a genetic determinant of bone mineral density. We tested this hypothesis in a sample of healthy men and women and did not find consistent evidence for an association between variation in this gene and either lumbar spine or femoral neck BMD. INTRODUCTION: Phenotypic variation in bone mineral density (BMD) among healthy adults is influenced by both genetic and environmental factors. A recent mouse study implicated ALOX5, which encodes arachidonate 5-lipoxygenase, as a contributing factor to areal BMD (aBMD). METHODS: Fifteen single nucleotide polymorphisms (SNPs) distributed throughout ALOX5 were genotyped in three healthy groups: 1,688 European American, premenopausal sisters, 512 African American premenopausal sisters and 715 European American brothers. Statistical analyses were performed in the three groups to test for association between these SNPs and femoral neck and lumbar spine aBMD. RESULTS: Significant (p < or = 0.05) evidence of association was observed with three of the SNPs. However, despite the linkage disequilibrium between SNPs, adjacent SNPs did not provide statistical evidence of association in any of the three study groups. CONCLUSIONS: These data do not provide consistent evidence of association between genomic variation in ALOX5 and clinical variability in aBMD in healthy subjects.

Association of the kappa-opioid system with alcohol dependence.

Opioid receptors and their endogenous peptide ligands play important roles in the reward and reinforcement of drugs such as heroin, cocaine, and alcohol. The binding of dynorphins to the kappa-opioid receptor has been shown to produce aversive states, which may prevent the development of reinforcement. We genotyped SNPs throughout OPRK1, encoding the kappa-opioid receptor, and PDYN, which encodes its ligand prodynorphin, in a group of 1860 European American individuals from 219 multiplex alcohol dependent families. Family-based analyses demonstrated associations between alcohol dependence and multiple SNPs in the promoter and 3' end of PDYN, and in intron 2 of OPRK1. Haplotype analyses further supported the association of PDYN. Thus, variations in the genes encoding both the kappa-opioid receptor and its ligand, OPRK1 and PDYN, are associated with the risk for alcohol dependence; this makes biological sense as variations in either should affect signaling through the kappa-opioid system.

Polymorphisms in the bone morphogenetic protein 2 (BMP2) gene do not affect bone mineral density in white men or women.

INTRODUCTION: Peak bone mineral density (BMD) achieved during adulthood is a major determinant of osteoporotic fracture in later life. Although environmental factors affect peak BMD, it is a highly heritable trait. Recently, bone morphogenetic protein 2 (BMP2) was reported as a susceptibility gene for osteoporotic fractures and low BMD in Icelandic and Danish populations. METHODS: To determine whether polymorphisms in the BMP2 gene contribute to BMD variation in our population of healthy American whites, we tested seven single nucleotide polymorphisms (SNPs), four of which were associated with osteoporotic phenotypes in the previous study. BMD at the femoral neck and lumbar spine (L2-L4) were measured by dual energy X-ray absorptiometry (DXA) in 411 men (age 18-61) and 1,291 pre-menopausal women (age 20-50). SNP genotypes/haplotypes were tested for population-based association with BMD using analysis of variance. RESULTS: None of the polymorphisms tested reached statistical significance (all p values >0.05) for BMD at the femoral neck or lumbar spine in either gender. Two of the SNP haplotypes spanning the entire BMP2 gene were marginally associated with BMD in men (p values=0.019-0.043). However, these haplotypes would account for only a small, if any, portion of BMD variation and would not be significant after adjustment for multiple comparisons. CONCLUSIONS: These results demonstrate that genetic variations in BMP2 do not substantially contribute to BMD variation in our population of healthy American whites.

Characterization of an echinocandin B-producing strain blocked for sterigmatocystin biosynthesis reveals a translocation in the stcW gene of the aflatoxin biosynthetic pathway.

Echinocandin B (ECB), a lipopolypeptide used as a starting material for chemical manufacture of the anti-Candida agent LY303366, is produced by fermentation using a strain of Aspergillus nidulans. In addition to ECB, the wild-type strain also produces a significant level of sterigmatocystin (ST), a potent carcinogen structurally related to the aflatoxins. Characterization of a mutant designated A42355-OC-1 (OC-1), which is blocked in ST biosynthesis, was the result of a chromosomal translocation. The chromosomal regions containing the breakpoints of the translocation were isolated and DNA sequencing and PCR analysis of the chromosomal breakpoints demonstrated the translocation occurred within the stcW gene of the ST biosynthetic pathway, resulting in disruption of the open reading frame for this gene. Biochemical feeding studies indicate the involvement of this gene product in the conversion of averufin to 1-hydroxy versicolorone. This work demonstrates an effective synergy between classical strain improvement methods and molecular genetics.

Genetic modification of an echinocandin B-producing strain of Aspergillus nidulans to produce mutants blocked in sterigmatocystin biosynthesis.

The production of echinocandin B (ECB), a lipopolypeptide used for chemical manufacture of the anti-Candida agent Cilofungin, was accomplished by fermentation using a strain of Aspergillus nidulans. In addition to ECB, this fermentation also produces a significant amount of sterigmatocystin (ST), a potent carcinogen structurally related to the aflatoxins. Mutants blocked in the ST biosynthetic pathway were created by genetic modification of the polyploid production strain C747. The following steps were involved: (i) reduction of the genotype to haploid by treatment with the spindle fiber poison methyl 1-(butylcarbamoyl)-2-benzimidazole carbamate (MBC), using colony morphology, conidia size, and the ability to obtain 5-fluoro-orotic acid (5-FOA)-resistant mutants as criteria for ploidy; (ii) mutagenesis of a haploid isolate using UV irradiation; and (iii) screening of mutants for inability to produce ST by thin layer chromatography. Six mutants blocked in ST production were isolated. All six remained capable of producing ECB equivalent in quantity to the haploid strain C747-GR14. One of the mutants was shown to be the result of a chromosomal translocation.

Molecular karyotype alterations induced by transformation in Aspergillus nidulans are mitotically stable.

Clamped homogeneous electric field (CHEF)-gel electrophoresis was used to define the electrophoretic molecular karyotype of Aspergillus nidulans strain OC-1 before and after protoplast-based genetic transformation. The transforming DNA caused alterations in the molecular karyotypes in all transformants examined. Rather dramatic changes were observed in karyotypes, including apparent chromosome loss, massive size alterations, and the appearance of large chromosomes. Changes in molecular karyotype were mitotically stable.

INF56 represents a family of differentiation-specific genes from Uromyces appendiculatus.

In germlings of U. appendiculatus, a gene designated INF56 is preferentially expressed during development of the infection structures. A comparison of sequences between INF56 genomic DNA and a cDNA revealed several differences, randomly distributed throughout the coding region, which resulted in RFLPs at ASpI, HphI, NruI and ScaI sites. This observation, along with DNA-blot analysis, which revealed multiple copies of INF56 in uredospore genomic DNA, indicated that INF56 represents a multigene family. All copies of INF56 examined contain the same 67-bp intron reported previously (Xuei et al. 1992).

Characterization of INF56, a gene expressed during infection structure development of Uromyces appendiculatus.

The bean rust fungus, Uromyces appendiculatus, undergoes thigmotropic differentiation to produce infection structures. Six differentiation-specific genes have been isolated and one, INF24, has been characterized [Bhairi et al., Gene 81 (1989) 237-243]. Here, we report the structure of a second gene, INF56, which was subcloned on a 2.6-kb fragment and sequenced. The location of the 1.0-kb INF56 transcript was determined by S1 nuclease protection and primer extension. A TATA box was found 38 bp upstream and a CAAT box 130 bp upstream from the major transcription start point (tsp). The gene contains two open reading frames: ORF2 is nested within ORF1; they share a 67-bp intron. ORF1 encodes a 14.1-kDa polypeptide which has an amino acid sequence rich in Gly, Pro and Ser. It has sequence similarity to a functional domain (V2) of mammalian cytokeratin type II. ORF2 encodes a 10.1-kDa polypeptide which is rich in Pro. It shares similarity with the cell-surface recognition region of chicken fibronectin. Hybrid selection and in vitro translation of the INF56 mRNA yielded two polypeptides of 15.5 and 23 kDa, as estimated by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis. INF56 is constitutively expressed at a low level, but the abundance of its steady-state transcript is upshifted 4.5 h after spore hydration during the period that infection structures are formed.