Association analysis of polymorphisms in the upstream region of the human dopamine D4 receptor gene (DRD4) with schizophrenia and personality traits

The human dopamine D4 receptor (DRD4) is of major interest in molecular studies of schizophrenia and personality traits. We examined the association of schizophrenia and polymorphisms in the upstream region of the DRD4 gene (−768G>A in the negative modulator region; −521C>T, −376C>T, and −291C>T in the cell type-specific promoter region; and −616C>G between the two regions) in 208 schizophrenic patients and 210 normal controls. No significant difference in genotype and allele frequencies was observed between the two groups, indicating that these polymorphisms do not make a major contribution to the pathogenesis of schizophrenia. We also studied the association of polymorphisms in the upstream region and a 48-bp repeat polymorphism in exon III of the DRD4 gene with personality traits in 173 Japanese individuals who completed the temperament and character inventory (TCI). The −768G>A polymorphism was significantly associated with reward dependence (P = 0.044), while no significant association was observed between novelty seeking and polymorphisms in the upstream region or the exon III repeat polymorphism of the DRD4 gene. Received: August 28, 2000 / Accepted: October 25, 2000     

Novel polymorphisms in the upstream region of the human dopamine D4 receptor (DRD4) gene

We found nine novel polymorphisms in the upstream region of the human dopamine D4 receptor (DRD4) gene of Japanese by direct sequencing. These polymorphisms are −809G > A, −768G > A, −616C > G, −603T > del, −602G > del, −600G > C, −376C > T, −291C > T, and −128G > T. One known polymorphism, −521C > T, was also recognized. Six of these sites were identified as restriction fragment length polymorphisms (RFLPs). Received: June 11, 1999 / Accepted: June 23, 1999     

A hypervariable segment in the human dopamine receptor D4 (DRD4) gene

The human dopamine D₄ receptor contains a novel polymorphismwithin the putative third cytoplasmic loop of the protein. Thepolymorphism is characterized by a varying number of directimperfect 48-bp repeats in the gene. Pharmacological characterizationhas suggested that this receptor is the site through which theatypical neuroleptic clozapine exerts its antipsychotic actionand that some polymorphic variants display different pharmacologicalproperties. Further analysis of the repeat region using innovativetechnologies indicates that the alleles vary not only in thenumber of repeats (2–8 or 10 repeat units) but also inthe sequence of the repeats and the order in which they appear.In 178 unrelated chromosomes we have identified 19 differentrepeats in 25 different haplotypes coding for 18 different predictedamino acid sequences, making this one of the most variable functionalproteins currently described.     

Lack of imprinting of the human dopamine D4 receptor (DRD4) gene

The term genomic imprinting has been used to refer to the differential expression of genetic material depending on whether it has come from the male or female parent. In humans, the chromosomal region 11p15.5 has been shown to contain 2 imprinted genes (H19 and IGF2). The gene for the dopamine D4 receptor (DRD4), which is of great interest for research into neuropsychiatric disorders and psychopharmacology, is also located in this area. In the present study, we have examined the imprinting status of the DRD4 gene in brain tissue of an epileptic patient who was heterozygous for a 12 bp repeat polymorphism in exon 1 of the DRD4 gene. We show that both alleles are expressed in equivalent amounts. We therefore conclude that the DRD4 gene is not imprinted in the human brain. © 1996 Wiley-Liss, Inc.     

Exon 3 polymorphisms of dopamine D4 receptor (DRD4) gene and attention deficit hyperactivity disorder in Chinese children.

Dopamine D4 receptor (DRD4) gene is implicated in the pathogenesis of attention deficit hyperactivity disorder (ADHD). The 7-repeat allele of the variable-number-of-tandem-repeat (VNTR) polymorphism in exon 3 has been reported to be associated with ADHD. However, studies in Chinese populations have yielded conflicting results. We therefore perform another study to investigate the association between ADHD and DRD4 gene polymorphism in Chinese children in Hong Kong. In this prospective family-based and case-control study during January-June 2004, we recruited consecutive Chinese children diagnosed with ADHD by DSM-IV and sex-matched controls admitted for acute upper respiratory infection, excluding those with perinatal brain insults, mental retardation, or neurological deficits. VNTR polymorphisms of the DRD4 gene were determined by standard PCR followed by agarose gel electrophoresis. Sixty-four ADHD cases (52 boys, 12 girls), their family members, and 64 normal controls were recruited. The 4-repeat allele (84.4%) and the 4/4-repeat genotype (70.3%) were the most prevalent. Both family-based and case-control analyses showed no association between ADHD and DRD4 gene polymorphisms (transmission dysequilibrium test (TDT): P = 0.91 and P = 0.33 for the 7-repeat and 4-repeat alleles, respectively; OR for the 7-repeat allele = 2.01 (95% CI 0.07-60.4, P = 0.66), OR for the 4-repeat allele = 1.51 (95% CI 0.80-2.85, P = 0.2)). However, the longer repeat alleles had a positive trend association with ADHD (P = 0.01) in the case-control analysis. We concluded that ADHD is not associated with a particular VNTR polymorphism of the DRD4 gene. Further studies are needed to clarify the role of repeat length of the VNTR region of the DRD4 gene in the pathogenesis of ADHD.     

Association of long variants of the dopamine D4 receptor exon 3 repeat polymorphism with Parkinson's disease

The dopamine D4 receptor ( D4DR ) has a highly polymorphic region in the third exon which has been associated with novelty seeking (NS) behavior. Due to the central position of dopamine and the documented low NS in Parkinson's disease (PD), the frequency of the exon 3 variants of D4DR in 95 PD patients and 47 controls was investigated. A significantly higher frequency of exon 3 alleles with six or more repeat units was found in the PD group (p = 0.039). This provides evidence that some forms of the highly polymorphic D4DR may represent a genetic susceptibility factor for PD.     

Association of dopamine D4 receptor (DRD4) polymorphisms with attention deficit hyperactivity disorder in Indian population.

Attention deficit hyperactivity disorder (ADHD) is a childhood onset neurobehavioral disorder. Several studies worldwide have implicated a possible association between ADHD and transmission of different polymorphisms of the dopamine D4 receptor gene (DRD4) in different ethnic groups. However, this is the first report on the transmission of different polymorphisms of DRD4 in Indian subjects. Association of 5' flanking 120-bp duplication, exon 1 12-bp duplication, and exon 3 48-bp variable numbers of tandem repeats (VNTR) were analyzed in 50 ADHD cases. Haplotype-based haplotype relative risk (HHRR) analysis and transmission disequilibrium test (TDT) were carried out to ascertain the association of these polymorphisms with the disorder. Linkage disequilibria (LD) between the polymorphisms were calculated using EH+ and 2LD programs. Our preliminary data showed lack of association between ADHD and transmission of the 5' flanking 120-bp duplication and exon 1 12-bp duplication. But, the transmissions of 6 and 7 repeat alleles of exon 3 48-bp VNTR showed significant association with ADHD. We have also examined the haplotype frequencies and biased transmission of one haplotype was observed in ADHD probands. LD analysis showed very strong disequilibrium between exon 1 12-bp duplication and exon 3 48-bp VNTR. Strong LD was also observed between the 5' flanking 120-bp duplication and exon 1 12-bp duplication. The observed association between higher repeat alleles of exon 3 48-bp VNTR and Indian ADHD children is consistent with some of the earlier reports.     

Homozygosity at the dopamine D3 receptor gene in schizophrenic patients

Dopamine receptors have long been implicated in the etiology of schizophrenia. It has been reported an association of schizophrenia with homozygosity at the dopamine D3 receptor gene locus. We have investigated the distribution of a D3 receptor gene polymorphism (BalI) in 107 schizophrenic Spanish patients and 100 healthy matched controls. No statistically significant differences between the patients and control group were detected with respect to either allele frequencies or genotype distribution. However, if not corrected for multiple testing, a correlation was found between homozygosity and early age of schizophrenia (χ² = 3.1, df = 1, P = 0.03) and between A1 allele frequency and disorganized and undifferentiated schizophrenia (χ² = 3.4, df = 1, P = 0.03; χ² = 2.7, df = 1, P = 0.05, respectively). These results suggest the possibility that D3 polymorphisms may be among the physiological factors underlying schizophrenia; though not the determining factor.     

No allelic association between Parkinson's disease and dopamine D2, D3, and D4 receptor gene polymorphisms

Parkinson's disease is thought to be caused by a combination of unknown environmental, genetic, and degenerative factors. Evidence from necropsy brain samples and pharmacokinetics suggests involvement of dopamine receptors in the pathogenesis or pathophysiology of Parkinson's disease. Genetic association studies between Parkinson's disease and dopamine D2, D3 and D4 receptor gene polymorphisms were conducted. The polymorphism was examined in 71 patients with Parkinson's disease and 90 controls. There were no significant differences between two groups in allele frequencies at the D2, D3, and D4 dopamine receptor loci. Our findings do not support the hypothesis that susceptibility to Parkinson's disease is associated with the dopamine receptor polymorphisms examined. © 1994 Wiley-Liss, Inc.     

No association of the dopamine D4 receptor (DRD4) and -521 C/T promoter polymorphisms with infant attachment disorganization

In a first molecular genetic study Lakatos and colleagues found an association between attachment disorganization and the dopamine D4 receptor (DRD4) gene polymorphism, in particular in the presence of the -521 T allele in the promoter region of the DRD4 gene. Replication of their study in a sample of 132 infants did not confirm the role of the DRD4 7+ -allele and the -521C/T promoter gene in disorganized attachment. Although our sample was larger, and contained more children with CT or TT alleles, which enhanced the probability of finding the DRD4 and C/T interaction, the association was not found. Even when we combined our sample with the Lakatos sample, the interaction effect of the DRD4 and -521 C/T polymorphisms on disorganized attachment was absent.     

Association study between the dopamine D4 receptor gene and schizophrenia

The dopamine D4 receptor is of major interest in schizophrenia research due to its high affinity for the atypical neuroleptic cloza-pine and a high degree of variability in the receptor gene (DRD4). Although several genetic linkage analyses performed on schizophrenia multiplex families from different regions of the world have either excluded or failed to prove that DRD4 is a major genetic factor for the development of schizophrenia, analyses for moderate predisposing effects are still of significant interest. We performed a study examining differences in allele frequencies of 4 different DRD4 polymorphisms in schizophrenia patients and age, sex, and ethnic origin matched controls. None of these 4 polymorphisms showed evidence for genetic association with schizophrenia, although a trend towards excess of the allele with 7 repeats in the (48)_n bp exon III polymorphism was observed. Complexities in the DRD4 genetic investigation and further analytic approaches are discussed. © 1995 Wiley-Liss, Inc.     

No association of the dopamine D4 receptor (DRD4) and -521 C/T promoter polymorphisms with infant attachment disorganization

In a first molecular genetic study Lakatos and colleagues found an association between attachment disorganization and the dopamine D4 receptor (DRD4) gene polymorphism, in particular in the presence of the -521 T allele in the promoter region of the DRD4 gene. Replication of their study in a sample of 132 infants did not confirm the role of the DRD4 7⁺ -allele and the -521C/T promoter gene in disorganized attachment. Although our sample was larger, and contained more children with CT or TT alleles, which enhanced the probability of finding the DRD4 and C/T interaction, the association was not found. Even when we combined our sample with the Lakatos sample, the interaction effect of the DRD4 and -521 C/T polymorphisms on disorganized attachment was absent.     

No association of the dopamine DRD4 receptor (DRD4) gene polymorphism with attention deficit hyperactivity disorder (ADHD) in the Irish population

Attention deficit hyperactivity disorder (ADHD) is one of the most prevalent childhood-onset syndromes affecting 3%-6% of school-age children worldwide. Although the biological basis of ADHD is unknown, a dopaminergic abnormality has long been suggested. The dopamine D4 receptor gene (DRD4) has been mapped to chromosome 11p15.5 and has been implicated in predisposition to ADHD. Several independent genetic association studies have demonstrated increased frequency of the DRD4 7-repeat allele in ADHD cases compared with controls or excess transmission of the 7-repeat allele from parents to affected offspring. However, there have also been few negative studies. In this study we investigated 78 ADHD parent proband trios and 21 parent proband pairs for the transmission of the DRD4 alleles in HHRR and case control design. We found no significant differences in the frequency of the DRD4 alleles transmitted or not transmitted to ADHD cases from their parents nor when comparing case allele frequencies to ethnically matched controls. Therefore, it is unlikely that the DRD4 7-repeat allele is associated with ADHD in the Irish population.     

Anorexia nervosa, perfectionism, and dopamine D4 receptor (DRD4).

The dopamine D4 receptor (DRD4), a well-characterized, polymorphic gene, is an attractive candidate for contributing risk to disordered eating and anorexia nervosa (AN). We tested association using UNPHASED for 5 DRD4 polymorphic loci, 3 promoter region SNPs (C-521T, C-616G, A-809G), the 120 bp promoter region tandem duplication and the exon III repeat, in 202 AN trios and 418 control families. Since perfectionism characterizes AN, we tested these five loci for association with the Child and Adolescent Perfectionism Scale (CAPS) in the AN and control groups. Single locus analysis showed significant association between the 'C' C-521T allele and AN. Haplotype analysis also showed significant association, particularly a 4-locus haplotype (exon III&120 bp repeat&C-521T&A-809G). Association was also observed between DRD4 and CAPS scores both for AN and control subjects. The insulin-like growth factor 2 (IGF2) and the arginine vasopressin 1a receptor (AVPR1a), previously shown to be associated with disordered eating, were also associated with CAPS scores. Three genes associated with AN were also associated with perfectionism. Personality traits are potential endophenotypes for understanding the etiology of eating disorders and one of the several pathways to eating pathology may be mediated by the impact of DNA sequences on perfectionism.     

Adult attachment and gene polymorphisms of the dopamine D4 receptor and serotonin transporter (5-HTT)

Recently, the Dopamine D4 Receptor Gene (DRD4) and the Serotonin Transporter Gene (5-HTT) have been found to be candidate genes for infant attachment disorganization. The present study aimed to explore the relationship of these genes to adult attachment representations. The Adult Attachment Interview was used to assess attachment representations in 167 German adults. DNA from buccal cells was genotyped for the DRD4 VNTR Exon III and 5-HTT LPR polymorphisms with respect to the presence of the 7repeat allele and the short allele, respectively. DRD4 7repeat allele carriers were significantly more likely to be securely attached than those without 7repeat but only for subjects with unloving caregiver recollections. No association between the 5-HTT LPR polymorphism and adult attachment was found. These findings encourage further investigations to explore endophenotypical and mediating psychological processes between the DRD4 Gene and secure attachment patterns.     

Association between dopamine D4 receptor (DRD4) exon III polymorphism and Neuroticism in the Japanese population

The association between the dopamine D4 receptor (DRD4) exon III polymorphism and personality trait of novelty seeking (NS) has been studied intensively. In the Japanese population, the results of the previous studies did not always coincide. In the present study, we investigated the association between the polymorphism and personality traits evaluated by using the Revised NEO Personality Inventory (NEO PI-R) and State-Trait Anxiety Inventory (STAI) in 196 Japanese subjects. A meta-analysis of the present and previous Japanese studies was also conducted regarding NS. As a result, significant association was observed between the polymorphism and personality traits evaluated by using NEO PI-R as a whole (p=0.022, MANCOVA). Subsequent analyses showed a significant association between short alleles (2-4 repeats) and higher scores for Neuroticism or its subscales, Anxiety, Depression, and Vulnerability (p=0.015, 0.039, 0.021, and 0.008, respectively, uncorrected). No other significant difference in the scores for NEO PI-R was observed in the subsequent analyses. Significant association was also observed between the polymorphism and scores for STAI as a whole (p=0.004, MANCOVA). Subsequent analyses did not show significant association, although a weak trend for the relation between the genotype consisting of short alleles and Trait Anxiety was observed (p=0.10, uncorrected). The meta-analysis showed no significant association between the polymorphism and NS. Thus, the present study suggested the association between the short allele of the DRD4 exon III polymorphism and personality trait of Neuroticism in Japanese subjects.     

Investigation of dopamine receptor (DRD4) and dopamine transporter (DAT) polymorphisms for genetic linkage or association to panic disorder

Clinical and animal studies suggest a role for the neurotransmitter dopamine in anxiety states. In humans, one such condition is panic disorder, which is typified by recurrent panic attacks accompanied by anticipatory anxiety. Family, segregation, and twin studies imply a genetic component to the pathophysiology of panic disorder. In this study, we examined the genes for the D4 dopamine receptor (DRD4) and the dopamine transporter (DAT) using three common sequence polymorphisms. Two of these polymorphisms were in DRD4, a 12 base-pair insertion/deletion in exon 1 and a 48 base-pair repeat in exon 3, and the third was a 40 base-pair repeat in the 3' untranslated region of DAT. We employed a family-based design, using 622 individuals in 70 families, as well as 82 haplotype relative risk "trios". Subjects were genotyped at the polymorphic loci, and the data were analyzed for genetic association and linkage. There were no significant differences in allele frequencies or occurrence of genotypes within the triads for any of the three polymorphisms. No significant linkage between the DRD4 or DAT polymorphisms and panic disorder was observed in the multiplex families, using a variety of simulations for dominant and recessive models of inheritance. However, LOD scores of approximately 1.1 and 1.05 were observed for the DAT and DRD4 exon 1 loci, respectively. The results reported here provide little support for the role of these polymorphisms in panic disorder.     

Association between the dopamine D3 receptor gene locus (DRD3) and unipolar affective disorder

Dopamine neurotransmission has been implicated in the pathophysiology of schizophrenia and, more recently, affective disorders. Among the dopamine receptors, D3 can be considered as particularly related to affective disorders due to its neuroanatomical localization in the limbic region of the brain and its relation to the serotoninergic activity of the CNS. The possible involvement of dopamine receptor D3 in unipolar (UP) major depression was investigated by a genetic association study of the D3 receptor gene locus (DRD3) on 36 UP patients and 38 ethnically matched controls. An allelic association of DRD3 (Bal I polymorphism) and UP illness was observed, with the Gly-9 allele (allele '2', 206/98 base-pairs long) being more frequent in patients than in controls (49% vs 29%, P < 0.02). The genotypes containing this allele (1-2 and 2-2) were found in 75% of patients vs 50% of controls (P < 0.03, odds ratio = 3.00, 95% CI = 1.12-8.05). The effect of the genotype remained significant (P < 0.02) after sex and family history were controlled by a multiple linear regression analysis. These results further support the hypothesis that dopaminergic mechanisms may be implicated in the pathogenesis of affective disorder. More specifically, the '2' allele of the dopamine receptor D3 gene seems to be associated with unipolar depression and can be considered as a 'phenotypic modifier' for major psychiatric disorders.