Human Dopamine D2 Receptor Gene Research Articles

Heteromerization between α2A adrenoceptors and different polymorphic variants of the dopamine D4 receptor determines pharmacological and functional differences. Implications for impulsive-control disorders

Polymorphic alleles of the human dopamine D4 receptor gene (DRD4) have been consistently associated with individual differences in personality traits and neuropsychiatric disorders, particularly between the gene encoding dopamine D4.7 receptor variant and attention deficit hyperactivity disorder (ADHD). The α2A adrenoceptor gene has also been associated with ADHD. In fact, drugs targeting the α2A adrenoceptor (α2AR), such as guanfacine, are commonly used in ADHD treatment. In view of the involvement of dopamine D4 receptor (D4R) and α2AR in ADHD and impulsivity, their concurrent localization in cortical pyramidal neurons and the demonstrated ability of D4R to form functional heteromers with other G protein-coupled receptors, in this study we evaluate whether the α2AR forms functional heteromers with D4R and weather these heteromers show different properties depending on the D4R variant involved. Using cortical brain slices from hD4.7R knock-in and wild-type mice, here, we demonstrate that α2AR and D4R heteromerize and constitute a significant functional population of cortical α2AR and D4R. Moreover, in cortical slices from wild-type mice and in cells transfected with α2AR and D4.4R, we detect a negative crosstalk within the heteromer. This negative crosstalk is lost in cortex from hD4.7R knock-in mice and in cells expressing the D4.7R polymorphic variant. We also show a lack of efficacy of D4R ligands to promote G protein activation and signaling only within the α2AR-D4.7R heteromer. Taken together, our results suggest that α2AR-D4R heteromers play a pivotal role in catecholaminergic signaling in the brain cortex and are likely targets for ADHD pharmacotherapy.

Genetic polymorphism of dopamine receptor D4 (DRD4) gene in ten Indian rhesus macaques (Macaca mulatta mulatta)

Advances in molecular genetical research have revealed a new platform for studying the configuration and activity correlations of dopamine receptor D4 (DRD4) gene in the non-human primates and human. Among the other five dopamine receptor genes, dopamine receptor D4 (DRD4) gene has variable number of tandem repeats which are polymorphic and located in the third cytoplasmic loop of the receptor. These polymorphism are expressed in the repeat numbers of 48-base pair (bp) sequence coding for 16 amino acids. This study is designed to observe the genetic polymorphism of dopamine receptor D4 gene within the ten Indian rhesus macaques (Macaca mulatta mulatta). This research is also planned to compare the genetic sequences among the Indian rhesus macaque DRD4 genetic sequences of our recent study, rhesus macaque DRD4 gene reference sequence provided by ENSEMBL genome browser and human DRD4 gene reference sequence provided by National Center for Biotechnology Information (NCBI), USA. In our study, only DRD4 5-repeat allele variant is observed in all the ten selected Indian rhesus macaques. Sequencing data of DRD4 gene of these rhesus macaques indicate that rhesus macaques exhibit 48-bp tandem repeats in the DRD4 receptor gene which is similar to humans and there is intraspecies variability within the rhesus macaque DRD4 genetic sequences.

Genetic variants in dopamine receptors influence on heterodimerization in the context of antipsychotic drug action.

Human dopamine D2 receptor (D2R) gene has polymorphic variants, three of them alter its amino acid sequence: Val96Ala, Pro310Ser and Ser311Cys. Their functional role never became the object of extensive studies, even though there are some evidence that they correlate with schizophrenia. The present work reviews data indicating that these mutations play a role in dimer formation with dopamine D1 receptor (D1R), with the strongest effect observed for Ser311Cys variant. Similarly, the affinity for antipsychotic drugs of this genetic variant depends on whether it is expressed together with D1R or not. Better understanding of altered ability of genetic variants of D2R to form dimers with D1R, as well as of altered affinity for antipsychotic drugs, depending on the absence or presence of the second dopamine receptor is of great importance-since these two receptors are not always co-expressed in the same cell. It may well be that targeting new compounds toward the D1R-D2R dimers, which the most probably form under conditions of excessive dopamine release, will result in antipsychotic drugs devoid of serious side effects.

34.2 DOPAMINE D4 RECEPTORS IN ANIMAL MODELS OF NEURODEVELOPMENTAL DISORDERS

Previous research suggests that the exon 3 variable number of tandem repeat (VNTR) polymorphism of the human dopamine D4 receptor gene (DRD4) contributes to ADHD and other psychiatric phenotypes. Animal studies may show how differences in D4 expression and function lead to these phenotypes and whether environmental factors influence behavioral outcomes.

Genetic variants of dopamine D2 receptor impact heterodimerization with dopamine D1 receptor.

The human dopamine D2 receptor gene has three polymorphic variants that alter its amino acid sequence: alanine substitution by valine in position 96 (V96A), proline substitution by serine in position 310 (P310S) and serine substitution by cysteine in position 311 (S311C). Their functional role has never been the object of extensive studies, even though there is some evidence that their occurrence correlates with schizophrenia. The HEK293 cell line was transfected with dopamine D1 and D2 receptors (or genetic variants of the D2 receptor), coupled to fluorescent proteins which allowed us to measure the extent of dimerization of these receptors, using a highly advanced biophysical approach (FLIM-FRET). Additionally, Fluoro-4 AM was used to examine changes in the level of calcium release after ligand stimulation of cells expressing different combinations of dopamine receptors. Using FLIM-FRET experiments we have shown that in HEK 293 expressing dopamine receptors, polymorphic mutations in the D2 receptor play a role in dimmer formation with the dopamine D1 receptor. The association level of dopamine receptors is affected by ligand administration, with variable effects depending on polymorphic variant of the D2 dopamine receptor. We have found that the level of heteromer formation is reflected by calcium ion release after ligand stimulation and have observed variations of this effect dependent on the polymorphic variant and the ligand. The data presented in this paper support the hypothesis on the role of calcium signaling regulated by the D1-D2 heteromer which may be of relevance for schizophrenia etiology.

Effects of l-Tyrosine on working memory and inhibitory control are determined by DRD2 genotypes: A randomized controlled trial

l-Tyrosine (TYR), the precursor of dopamine (DA), has been shown to enhance facets of cognitive control in situations with high cognitive demands. However some previous outcomes were mixed: some studies reported significant improvements, while other did not. Given that TYR increases DA level in the brain, we investigated, in a double-blind, randomized, placebo-controlled design, whether the C957T genotypes of a functional synonymous polymorphism in the human dopamine D2 receptor (DRD2) gene (rs6277) contribute to individual differences in the reactivity to TYR administration and whether this factor predicts the magnitude of TYR-induced performance differences on inhibiting behavioral responses in a stop-signal task and working memory (WM) updating in a N-back task. Our findings show that T/T homozygotes (i.e., individuals potentially associated with lower striatal DA level) showed larger beneficial effects of TYR supplementation than C/C homozygotes (i.e., individuals potentially associated with higher striatal DA level), suggesting that genetically determined differences in DA function may explain inter-individual differences in response to TYR supplementation. These findings reinforce the idea that genetic predisposition modulates the effect of TYR in its role as cognitive enhancer.

MiR-217 mediates the protective effects of the dopamine D2 receptor on fibrosis in human renal proximal tubule cells.

Lack or downregulation of the dopamine D2 receptor (D2R) increases the vulnerability to renal inflammation independent of blood pressure in mice. Common single nucleotide polymorphisms (SNPs) rs6276, 6277, and 1800497 in the human D2R gene are associated with decreased receptor expression/function and hypertension. Human renal proximal tubule cells from subjects carrying these SNPs have decreased D2R expression and increased expression of profibrotic factors and production of extracellular matrix proteins. We tested the hypothesis that the D2R mediates these effects by regulating micro-RNA expression. In cells carrying D2R SNPs, micro-RNAs (miRs)-217, miR-224, miR-335, and miR-1265 were downregulated, whereas miR-1290 was upregulated >4-fold compared with those carrying D2R wild-type alleles. However, only miR-217 was directly regulated by D2R expression. In cells carrying D2R wild-type, miR-217 inhibitor increased the expression of transforming growth factor (TGF)-β1, matrix metalloproteinase 3, fibronectin 1, and collagen 1a, whereas miR-217 mimic had the opposite effect. In cells carrying D2R SNPs, miR-217 mimic also decreased the expression of TGFβ1 and its targets. Wnt5a, a miR-217 target, was increased in cells carrying D2R SNPs and decreased by miR-217 mimic but increased by miR-217 inhibitor in both cell types. In cells carrying D2R wild-type, Wnt5a treatment increased TGFβ1 while silencing Ror2, a Wnt5a receptor, decreased TGFβ1 and blunted the Wnt5a-induced increase in cells carrying D2R wild-type. Our results show that renal proximal tubule cells from subjects carrying D2R SNPs resulting in D2R downregulation have increased TGFβ1 that is mediated by decreased regulation of the miR-217-Wnt5a-Ror2 pathway.

Abstract 257: miR-217 Mediates the Protective Effects of the Dopamine D2 Receptor on Renal Inflammation and Fibrosis.

Some common single nucleotide polymorphisms (SNPs; rs 6276, 6277, and 1800497) in the human dopamine D2 receptor (DRD2) gene are associated with decreased receptor expression and function, as well as high blood pressure. We have reported that human renal proximal tubule cells (hRPTCs) from subjects carrying D2R SNPs had decreased D2R expression and function and increased expression of the pro-fibrotic factor TGFβ1 and its signaling targets Smad3 and Snail1. These cells also showed induction of epithelial mesenchymal transition and production of extracellular matrix proteins fibronectin-1 (FN-1), Collagen 1a (Col 1a), and vimentin. To determine the mechanisms underlying the effects of the D2R SNPs, we studied the expression of several microRNAs (miRs) that are associated with the regulation of chemokines, anti-fibrosis, extracellular matrix remodeling, and cell adhesion through post-transcriptional repression of their target mRNAs. The expression of 4 miRs (miR-217, miR-224, miR-335, miR-1265) was downregulated (-10.3 to -6.0 fold) and that of one miRNA (miR-1290; 5.3 fold) was upregulated in hRPTCs with D2R SNPs. The expression of miR-217 and miR-224 is regulated by the D2R; the expression is decreased by D2R downregulation in cells not carrying D2R SNPs (WT) and normalized by treating cells carrying D2R SNPs with a Drd2 plasmid. Treatment of WT cells with a miR-217 mimic significantly decreased the mRNA expression of TGFβ1 (43±2%), MMP3 (65±4%), and FN-1 (30±4%) while treatment with a miR-217 inhibitor increased TGFβ1 (94± 9%), MMP3 (96±5%), and FN-1 (212±30%). miR-217 mimic treatment of D2R SNPs cells also significantly decreased the expression of TGFβ1, MMP3, and FN-1. However, treatment of these cells with a miR-217 inhibitor had non-significant effects probably because miR-217 level was already reduced. In contrast treatment of either WT or D2R SNPs cells with miR-224 mimic or inhibitor had no significant effects on the expression of TGFβ1, MMP3, or FN-1. These results indicate that the protective effects of D2R on inflammation and fibrosis are at least, in part, mediated by regulation of miR-217 expression. Dysregulation of miR-217 expression may be responsible for the profibrotic phenotype of hRPTCs in individuals with these D2R SNPs.

Functional characterization of rare variants in human dopamine receptor D4 gene by genotype–phenotype correlations

Next generation sequencing technologies have facilitated a notable shift from common disease common variant hypothesis to common disease rare variant, as also witnessed in recent literature on schizophrenia. Dopamine receptor D4 (DRD4), a G-protein-coupled receptor is associated with psychiatric disorders and has high affinity for atypical antipsychotic clozapine. We investigated the functional role of rare genetic variants in DRD4 which may have implications for translational medicine. CHO-K1 cells independently expressing four rare non-synonymous variants of DRD4 namely R237L, A281P, S284G located in the third cytosolic loop and V194G, located in the fifth transmembrane domain were generated. Their genotype–phenotype correlations were evaluated using [3H]spiperone binding, G-protein activation and molecular dynamics-simulation studies. A281P and S284G were functionally similar to wildtype (WT). With R237L, potency of dopamine and quinpirole reduced ∼sixfold and threefold respectively compared to WT; [3H]spiperone binding studies showed a reduction in total number of binding sites (∼40%) but not binding affinity, in silico docking studies revealed that binding of both dopamine and spiperone to R237L was structurally similar to WT. Of note, V194G variant failed to inhibit forskolin-stimulated adenylate cyclase activity and phosphorylate extracellular signal-regulated kinase; showed significant reduction in binding affinity (Kd=2.16nM) and total number of binding sites (∼66%) compared to WT in [3H]spiperone binding studies; and ligand docking studies showed that binding of dopamine and spiperone is superficial due to probable structural alteration. Transmembrane variant V194G in DRD4.4 results in functional alteration warranting continuing functional analysis of rare variants.

An evolutionary conserved region (ECR) in the human dopamine receptor D4 gene supports reporter gene expression in primary cultures derived from the rat cortex

BackgroundDetecting functional variants contributing to diversity of behaviour is crucial for dissecting genetics of complex behaviours. At a molecular level, characterisation of variation in exons has been studied as they are easily identified in the current genome annotation although the functional consequences are less well understood; however, it has been difficult to prioritise regions of non-coding DNA in which genetic variation could also have significant functional consequences. Comparison of multiple vertebrate genomes has allowed the identification of non-coding evolutionary conserved regions (ECRs), in which the degree of conservation can be comparable with exonic regions suggesting functional significance.ResultsWe identified ECRs at the dopamine receptor D4 gene locus, an important gene for human behaviours. The most conserved non-coding ECR (D4ECR1) supported high reporter gene expression in primary cultures derived from neonate rat frontal cortex. Computer aided analysis of the sequence of the D4ECR1 indicated the potential transcription factors that could modulate its function. D4ECR1 contained multiple consensus sequences for binding the transcription factor Sp1, a factor previously implicated in DRD4 expression. Co-transfection experiments demonstrated that overexpression of Sp1 significantly decreased the activity of the D4ECR1 in vitro.ConclusionBioinformatic analysis complemented by functional analysis of the DRD4 gene locus has identified a) a strong enhancer that functions in neurons and b) a transcription factor that may modulate the function of that enhancer.

Effect of model choice in genetic association studies: DRD4 exon III VNTR and cigarette use in young adults

A major concern with the vast literature associating the highly polymorphic 48 bp VNTR in exon III of the human dopamine receptor D4 gene (DRD4) with various behavioral phenotypes is the lack of concordance between studies. Part of the problem arises from the absence of a universally accepted scheme for pooling the large number of low frequency genotypes into appropriate categories. Here, we investigated the effect of different pooling strategies and genetic models on the reported association between DRD4-exIII-VNTR polymorphism and cigarette smoking. Genotyping was performed on a large randomly selected community-based sample of 2,274 individuals aged 20-24 years. Participants were grouped into sub-samples based on their genotypes to test specific genetic models. Multiple regression analyses were used to assess the relationship between DRD4-exIII-VNTR genotype and cigarette smoking measures while controlling for confounders. While smoking status and age at start of smoking were not associated with the genotype, a significantly (P = 0.006) higher rate of cigarette consumption was observed among carriers of the 7-repeat (7r) allele. Thus, 7r carriers were not more likely to be smokers but if they did smoke they consumed significantly more cigarettes per day than 4r carriers. Unlike previous studies this association was observed only when comparing carriers of the 7r with the 4r but not the other repeat alleles. Our study demonstrates the need for caution when grouping functionally different DRD4-exIII-VNTR alleles in association studies. It particularly highlights the requirement for better functional characterization of the DRD4-exIII-VNTR alleles for interpreting results from association studies.

Associations between Dopamine D4 Receptor Gene Variation with Both Infidelity and Sexual Promiscuity

BackgroundHuman sexual behavior is highly variable both within and between populations. While sex-related characteristics and sexual behavior are central to evolutionary theory (sexual selection), little is known about the genetic bases of individual variation in sexual behavior. The variable number tandem repeats (VNTR) polymorphism in exon III of the human dopamine D4 receptor gene (DRD4) has been correlated with an array of behavioral phenotypes and may be predicatively responsible for variation in motivating some sexual behaviors, particularly promiscuity and infidelity.Methodology/Principal FindingsWe administered an anonymous survey on personal history of sexual behavior and intimate relationships to 181 young adults. We also collected buccal wash samples and genotyped the DRD4 VNTR. Here we show that individuals with at least one 7-repeat allele (7R+) report a greater categorical rate of promiscuous sexual behavior (i.e., having ever had a “one-night stand”) and report a more than 50% increase in instances of sexual infidelity.Conclusions/SignificanceDRD4 VNTR genotype varies considerably within and among populations and has been subject to relatively recent, local selective pressures. Individual differences in sexual behavior are likely partially mediated by individual genetic variation in genes coding for motivation and reward in the brain. Conceptualizing these findings in terms of r/K selection theory suggests a mechanism for selective pressure for and against the 7R+ genotype that may explain the considerable global allelic variation for this polymorphism.

Genetic diagnostics of functional variants of the human dopamine D2 receptor gene

The importance of dopamine D2 receptors (DRD2) for central nervous dopaminergic signalling makes variants in the DRD2 gene potential modulators of the risk or course of various behavioural, psychiatric or neurologic diseases (e.g. addiction, schizophrenia, Parkinson's disease). We developed Pyrosequencing genetic screening assays for single nucleotide polymorphisms spanning the whole range of the DRD2 gene locus up to the functionally related ankyrin repeat and kinase domain containing 1 gene (ANKK1) located at approximately 10 kb downstream of DRD2. Assays for 11 genetic variants with reported functional association were developed in DNA samples from 300 unrelated healthy Caucasians and validated by independent conventional sequencing. In all DNA samples the DRD2/ANKK1 genetic variants were identified correctly as verified by the control samples. The observed frequencies of homozygous, heterozygous and noncarriers of the minor alleles were in agreement with the Hardy-Weinberg equilibrium. Observed minor allele frequencies were DRD2 rs12364283T>C: 6.5%, rs1799978A>G: 4.8%, rs1799732C del: 14.2%, rs4648317C>T: 12.8%, rs1079597G>A: 13.8%, rs1076560G>T: 14.5%, rs1800496C>T: 0.2%, rs1801028C>G: 3.0%, rs6275C>T: 32.7%, rs6277C>T: 53.0% and ANKK1 rs1800497C>T: 17.5%. The presently developed Pyrosequencing assays are provided to facilitate further research toward personalized approaches to pathophysiological conditions involving behavioural, psychiatric and neurologic disorders including addiction, schizophrenia and Parkinson's disease.

C957T polymorphism of dopamine D2 receptor gene affects striatal DRD2 in vivo availability by changing the receptor affinity

The C957T polymorphism of the human dopamine D2 receptor gene (DRD2) regulates DRD2 availability in striatum in vivo. Specifically, the T allele predicts high DRD2 availability in healthy volunteers (T/T>T/C>C/C). However, this finding was unexpected as in vitro the T allele is associated with a decrease in DRD2 mRNA stability and synthesis of the receptor through a putative alteration in the receptor mRNA folding. To elucidate further how changes in DRD2 density (B(max)) and affinity (K(D)) contribute to the differences in DRD2 availability between the C957T genotypes, we studied these parameters separately in a sample of 45 healthy volunteers. The subjects had two PET scans with [(11)C]raclopride (high and low specific radioactivity scans) for the estimation of B(max) and K(D), and were genotyped for the C957T. Moreover, the role of the related and previously studied functional TaqIA polymorphism of ankyrin repeat and kinase domain containing 1 (ANKK1) gene was reassessed for comparative purposes. The results indicate that the C957T increased binding potential by decreasing DRD2 K(D) (C/C>C/T>T/T), while B(max) was not significantly altered. These preliminary findings indicate that the C957T genotype-dependent changes in DRD2 availability are driven by alterations in receptor affinity and putatively in striatal dopamine levels. This mechanism seems to differ from that observed previously for the ANKK1 gene TaqIA polymorphism, where the minor allele (A1) affects DRD2 availability predominantly by changing B(max). The hypothesis that the two SNPs may have independent effects on dopamine neurotransmission needs to be further tested.

Genetic contributions to avoidance-based decisions: striatal D2 receptor polymorphisms

Individuals differ in their tendencies to seek positive decision outcomes or to avoid negative ones. At the neurobiological level, our model suggests that phasic changes in dopamine support learning to reinforce good decisions via striatal D1 receptors, and to avoid maladaptive choices via striatal D2 receptors. Accordingly, in a previous study individual differences in positive and negative learning were strongly modulated by two genetic polymorphisms factors related to striatal D1 and D2 function, respectively. Nevertheless, whereas the role for dopamine in positive learning is relatively well accepted, that in learning to avoid negative outcomes is more controversial. Here we further explore D2-receptor-related genetic contributions to probabilistic avoidance in humans, in light of recent data showing that particular DRD2 polymorphisms are associated with functional modulation of receptor expression [Zhang Y, Bertolino A, Fazio L, Blasi G, Rampino A, Romano R, Lee M-LT, Xiao T, Papp A, Wang D, Sadée W (2007) Polymorphisms in human dopamine d2 receptor gene affect gene expression, splicing, and neuronal activity during working memory. Proc Natl Acad Sci U S A 104(51):20552–20557]. We find that a promoter polymorphism rs12364283 associated with transcription and D2 receptor density was strongly and selectively predictive of avoidance-based decisions. Two further polymorphisms (rs2283265 and rs1076560) associated with relatively reduced presynaptic relative to postsynaptic D2 receptor expression were predictive of relative impairments in negative compared to positive decisions. These previously undocumented effects of DRD2 polymorphisms were largely independent of those we reported previously for the C957T polymorphism (rs6277) associated with striatal D2 density. In contrast, effects of the commonly studied Taq1A polymorphism on reinforcement-based decisions were due to indirect association with C957T. Taken together these findings suggest multiple D2-dependent genetic mechanisms contributing to avoidance. We discuss these effects in the context of neurocomputational models of reinforcement leaning in the basal ganglia.

C957T polymorphism of the human dopamine D2 receptor gene predicts extrastriatal dopamine receptor availability in vivo

The C957T (rs6277) single nucleotide polymorphism (SNP) of the human dopamine D2 receptor (DRD2) gene ( DRD2) affects DRD2 mRNA stability and has been shown to predict striatal DRD2 availability ( B max/ K D) in vivo in man. Specifically, the C/C genotype is associated with low striatal DRD2 availability (C/C<C/T<T/T). It is not known, however, whether this pattern of genetic regulation of DRD2 expression also applies to low density DRD2 populations in extrastriatal regions. We analyzed extrastriatal DRD2 availability (indexed by binding potential, BP ND) measured in 38 healthy male volunteers with 3D-PET and the high-affinity DRD2 radioligand [ 11C]FLB457. The subjects were genotyped for the C957T as well as for two other widely studied DRD2 SNPs, the TaqIA (rs1800497) and the −141C Ins/Del (rs1799732). Statistical analyses showed that the C957T C/C genotype was associated with high extrastriatal DRD2 BP ND throughout the cortex and the thalamus (C/C>C/T>T/T). Also the TaqIA A1 allele carriers ( p = 0.101) tended to have higher extrastriatal DRD2 BP ND compared to non-carriers whereas the −141C Ins/Del genotype did not influence extrastriatal DRD2 BP ND. Our findings indicate that the DRD2 SNPs regulate DRD2 availability in the human cortex and in the thalamus in vivo. However, the regulation pattern is different from that observed previously for striatal DRD2 availability in vivo, which may reflect distinct functional roles of dopamine and DRD2 in the cortex versus the striatum. The results provide useful information for the interpretation of genetic studies exploring the role of the DRD2 in normal physiology as well as in psychiatric and neurological diseases.

Polymorphisms in human dopamine D2 receptor gene affect gene expression, splicing, and neuronal activity during working memory

Subcortical dopamine D2 receptor (DRD2) signaling is implicated in cognitive processes and brain disorders, but the effect of DRD2 variants remains ambiguous. We measured allelic mRNA expression in postmortem human striatum and prefrontal cortex and then performed single nucleotide polymorphism (SNP) scans of the DRD2 locus. A previously uncharacterized promoter SNP (rs12364283) located in a conserved suppressor region was associated with enhanced DRD2 expression, whereas previously studied DRD2 variants failed to affect expression. Moreover, two frequent intronic SNPs (rs2283265 and rs1076560) decreased expression of DRD2 short splice variant (expressed mainly presynaptically) relative to DRD2 long (postsynaptic), a finding reproduced in vitro by using minigene constructs. Being in strong linkage disequilibrium with each other, both intronic SNPs (but not rs12364283) were also associated with greater activity of striatum and prefrontal cortex measured with fMRI during working memory and with reduced performance in working memory and attentional control tasks in healthy humans. Our results identify regulatory DRD2 polymorphisms that modify mRNA expression and splicing and working memory pathways.

Association of polymorphisms in the dopamine D4 receptor gene and the activity‐impulsivity endophenotype in dogs

A variable number of tandem repeats (VNTR) polymorphism in exon 3 of the human dopamine D4 receptor gene (DRD4) has been associated with attention deficit hyperactivity disorder (ADHD). Rodents possess no analogous repeat sequence, whereas a similar tandem repeat polymorphism of the DRD4 gene was identified in dogs, horses and chimpanzees. Here, we present a genetic association study of the DRD4 VNTR and the activity-impulsivity dimension of the recently validated dog-ADHD Rating Scale. To avoid false positives arising from population stratification, a single breed of dogs (German shepherd) was studied. Two DRD4 alleles (referred to as 2 and 3a) were detected in this breed, and genotype frequencies were in Hardy-Weinberg equilibrium. For modelling distinct environmental conditions, 'pet' and 'police' German shepherds were characterized. Police German shepherds possessing at least one 3a allele showed significantly higher scores in the activity-impulsivity dimension of the dog-ADHD Rating Scale than dogs without this allele (P = 0.0180). This difference was not significant in pet German shepherds. To the best of our knowledge, this is the first report of an association between a candidate gene and a behaviour trait in dogs, and it reinforces the functional role of DRD4 exon 3 polymorphism.

Genetic structure of the dopamine receptor D4 gene ( DRD4) and lack of association with schizophrenia in Japanese patients

In order to investigate the contribution of genetic variation in the human dopamine receptor D4 gene ( DRD4) to the risk of developing schizophrenia, we carried out a genetic analysis of 27 polymorphisms in 216 schizophrenic patients and 243 healthy controls from the Kyushu region of Japan. Twenty-two single nucleotide polymorphisms (SNPs) and five insertion/deletion polymorphisms were analyzed in this study, including four novel SNPs and a novel mononucleotide repeat. Linkage disequilibrium (LD) and haplotype analyses reveal weak LD across the DRD4 gene. In univariate analysis female individuals with allele −521C had a higher risk for schizophrenia. However, this finding was not significant after correction for multiple hypothesis testing. No other polymorphisms or haplotypes differed between schizophrenic patients and controls. Likewise, multivariate analyses did not reveal any statistically significant associations.

No direct effect of the -521 C/T polymorphism in the human dopamine D4 receptor gene promoter on transcriptional activity

BackgroundThe human dopamine D4 receptor (DRD4) gene has been studied extensively as a candidate gene for certain psychological traits and several behavioural and psychiatric disorders. Both the 5' regulatory region and the coding sequence contain a number of polymorphisms. The promoter variants have received particular attention in the past few years due to their possible role in the regulation of gene transcription. Previously, the -521C/T SNP was shown to influence promoter activity. The aim of this study is to perform an in-depth analysis of this effect in the context of various neural cell lines.ResultsEndogenous mRNA expression of the DRD4 gene was demonstrated in two neuroblastoma (SK-N-F1, IMR32) and one retinoblastoma cell line (Y79) by RT-PCR. In addition, very low DRD4 mRNA levels were also detected in HeLa cells. The transcriptional activity of a series of 5' promoter deletion mutants was determined by transient transfection of luciferase reporter constructs. The activity profile of these promoter fragments was similar in each of the cell lines tested. The highest luciferase reporter activity was obtained with a construct containing promoter sequences between nucleotides -668 to -389, while a putative silencer region was localised spanning from nucleotide -1571 to -800. Surprisingly, the -521 C/T polymorphism had no significant effect on transcriptional activity of the reporter construct with the highest activity (-668 to -389) in any of the three cell lines tested.ConclusionOur results do not confirm previous data assigning different transcriptional activities to the -521 C/T alleles of the human DRD4 promoter. Furthermore, these findings highlight the need for further characterization of the 5' regulatory region of the DRD4 gene and identification of additional functional promoter polymorphic sites, especially in the context of haplotype.