DNA mismatch repair protects the genome from oxygen-induced replicative mutagenesis.

DNA mismatch repair (MMR) corrects mismatched DNA bases arising from multiple sources including polymerase errors and base damage. By detecting spontaneous mutagenesis using whole genome sequencing of cultured MMR deficient human cell lines, we show that a primary role of MMR is the repair of oxygen-induced mismatches. We found an approximately twofold higher mutation rate in MSH6 deficient DLD-1 cells or MHL1 deficient HCT116 cells exposed to atmospheric conditions as opposed to mild hypoxia, which correlated with oxidant levels measured using electron paramagnetic resonance spectroscopy. The oxygen-induced mutations were dominated by T to C base substitutions and single T deletions found primarily on the lagging strand. A broad sequence context preference, dependence on replication timing and a lack of transcriptional strand bias further suggested that oxygen-induced mutations arise from polymerase errors rather than oxidative base damage. We defined separate low and high oxygen-specific MMR deficiency mutation signatures common to the two cell lines and showed that the effect of oxygen is observable in MMR deficient cancer genomes, where it best correlates with the contribution of mutation signature SBS21. Our results imply that MMR corrects oxygen-induced genomic mismatches introduced by a replicative process in proliferating cells.

Characterisation of the spectrum and genetic dependence of collateral mutations induced by translesion DNA synthesis.

Translesion DNA synthesis (TLS) is a fundamental damage bypass pathway that utilises specialised polymerases with relaxed template specificity to achieve replication through damaged DNA. Misinsertions by low fidelity TLS polymerases may introduce additional mutations on undamaged DNA near the original lesion site, which we termed collateral mutations. In this study, we used whole genome sequencing datasets of chicken DT40 and several human cell lines to obtain evidence for collateral mutagenesis in higher eukaryotes. We found that cisplatin and UVC radiation frequently induce close mutation pairs within 25 base pairs that consist of an adduct-associated primary and a downstream collateral mutation, and genetically linked their formation to TLS activity involving PCNA ubiquitylation and polymerase κ. PCNA ubiquitylation was also indispensable for close mutation pairs observed amongst spontaneously arising base substitutions in cell lines with disrupted homologous recombination. Collateral mutation pairs were also found in melanoma genomes with evidence of UV exposure. We showed that collateral mutations frequently copy the upstream base, and extracted a base substitution signature that describes collateral mutagenesis in the presented dataset regardless of the primary mutagenic process. Using this mutation signature, we showed that collateral mutagenesis creates approximately 10-20% of non-paired substitutions as well, underscoring the importance of the process.

Factor Structure, Psychometric Properties, and Validation of the Hungarian Version of the Experiences in Close Relationships Revised (ECR-R-HU) Questionnaire in a Nationally Representative Community Sample.

The Experiences in Close Relationships - Revised (ECR-R) is a widely used self-report instrument to assess adult romantic attachment. The present study aimed at examining the factor structure, reliability, construct validity, and temporal stability of the Hungarian version of the ECR-R (ECR-R-HU) in a nationally representative community sample (N = 958). The original avoidance and anxiety dimensions of the ECR-R could only be identified, when reversed-item method factors and residual correlations were included in the confirmatory factor analysis (CFA). The Avoidance and Anxiety subscales of the ECR-R-HU showed high reliabilities and adequate temporal stability over 4 months. The subscales were not significantly associated with respondents' age, gender, and residence type, while being engaged in a romantic relationship was related to significantly lower scores on both subscales. Correlations with measures of family functioning problems, perceived stress, depressed mood, and well-being were significant and in the expected directions. These results confirm the ECR-R-HU as a reliable and valid assessment tool.

BRCA1 deficiency specific base substitution mutagenesis is dependent on translesion synthesis and regulated by 53BP1.

Defects in BRCA1, BRCA2 and other genes of the homology-dependent DNA repair (HR) pathway cause an elevated rate of mutagenesis, eliciting specific mutation patterns including COSMIC signature SBS3. Using genome sequencing of knock-out cell lines we show that Y family translesion synthesis (TLS) polymerases contribute to the spontaneous generation of base substitution and short insertion/deletion mutations in BRCA1 deficient cells, and that TLS on DNA adducts is increased in BRCA1 and BRCA2 mutants. The inactivation of 53BP1 in BRCA1 mutant cells markedly reduces TLS-specific mutagenesis, and rescues the deficiency of template switch-mediated gene conversions in the immunoglobulin V locus of BRCA1 mutant chicken DT40 cells. 53BP1 also promotes TLS in human cellular extracts in vitro. Our results show that HR deficiency-specific mutagenesis is largely caused by TLS, and suggest a function for 53BP1 in regulating the choice between TLS and error-free template switching in replicative DNA damage bypass.

Two main mutational processes operate in the absence of DNA mismatch repair.

The analysis of tumour genome sequences has demonstrated high rates of base substitution mutagenesis upon the inactivation of DNA mismatch repair (MMR), and the resulting somatic mutations in MMR deficient tumours appear to significantly enhance the response to immune therapy. A handful of different algorithmically derived base substitution mutation signatures have been attributed to MMR deficiency in tumour somatic mutation datasets. In contrast, mutation data obtained from whole genome sequences of isogenic wild type and MMR deficient cell lines in this study, as well as from published sources, show a more uniform experimental mutation spectrum of MMR deficiency. In order to resolve this discrepancy, we reanalysed mutation data from MMR deficient tumour whole exome and whole genome sequences. We derived two base substitution signatures using non-negative matrix factorisation, which together adequately describe mutagenesis in all tumour and cell line samples. The two new signatures broadly resemble COSMIC signatures 6 and 20, but perform better than existing COSMIC signatures at identifying MMR deficient tumours in mutation signature deconstruction. We show that the contribution of the two identified signatures, one of which is dominated by C to T mutations at CpG sites, is biased by the different sequence composition of the exome and the whole genome. We further show that the identity of the inactivated MMR gene, the tissue type, the mutational burden or the patient's age does not influence the mutation spectrum, but that a tendency for a greater contribution by the CpG mutational process is observed in tumours as compared to cultured cells. Our analysis suggest that two separable mutational processes operate in the genomes of MMR deficient cells.

Differential Genetic Effect of the Norepinephrine Transporter Promoter Polymorphisms on Attention Problems in Clinical and Non-clinical Samples.

Among the monoaminergic modulatory neurotransmitters, norepinephrine is involved in task orienting, hence noradrenergic genetic variants have been studied in connection to attentional processes. The role of this catecholamine system is also highlighted by the selective norepinephrine transporter blocking atomoxetine, which has proved to be effective in the pharmacological treatment of Attention Deficit Hyperactivity Disorder (ADHD). In the present genetic association study three single nucleotide polymorphisms (rs28386840, rs2242446, rs3785143 SNPs) were analyzed from the 5' region of the norepinephrine transporter (NET, SLC6A2) gene, which have been linked to ADHD previously. Attention problems scores of the mother-rated Child Behavior Checklist (CBCL) were used in separate analyses of 88 preschoolers (59.1% male, 6 years of age) recruited from the general population and 120 child psychiatry patients with ADHD diagnosis (85.8% male, age: 9.8 ± 2.9). The NET SNPs showed associations with attention problems, but the direction was different in the two groups. Regarding the promoter variant rs28386840, which showed the most consistent association, the T-allele-carrier patients with ADHD had lower CBCL attention problems scores compared to patients with AA genotype (p = 0.023), whereas T-allele-carriers in the community sample had more attention problems (p = 0.042). Based on previous reports of lower NE levels in ADHD children and the inverted-U shape effect of NE on cognitive functions, we propose that rs28386840 (-3081) T-allele, which is associated with lower NET expression (and potentially higher synaptic NE level) would support attention processes among ADHD patients (similarly as atomoxetine increases NE levels), whereas it would hinder cortical functions in healthy children.

A genetic study based on PCNA-ubiquitin fusions reveals no requirement for PCNA polyubiquitylation in DNA damage tolerance.

Post-translational modifications of Proliferating Cell Nuclear Antigen (PCNA) play a key role in regulating the bypass of DNA lesions during DNA replication. PCNA can be monoubiquitylated at lysine 164 by the RAD6-RAD18 ubiquitin ligase complex. Through this modification, PCNA can interact with low fidelity Y family DNA polymerases to promote translesion synthesis. Monoubiquitylated PCNA can be polyubiquitylated on lysine 63 of ubiquitin by a further ubiquitin-conjugating complex. This modification promotes a template switching bypass process in yeast, while its role in higher eukaryotes is less clear. We investigated the function of PCNA ubiquitylation using a PCNAK164R mutant DT40 chicken B lymphoblastoma cell line, which is hypersensitive to DNA damaging agents such as methyl methanesulfonate (MMS), cisplatin or ultraviolet radiation (UV) due to the loss of PCNA modifications. In the PCNAK164R mutant we also detected cell cycle arrest following UV treatment, a reduced rate of damage bypass through translesion DNA synthesis on synthetic UV photoproducts, and an increased rate of genomic mutagenesis following MMS treatment. PCNA-ubiquitin fusion proteins have been reported to mimic endogenous PCNA ubiquitylation. We found that the stable expression of a PCNAK164R-ubiquitin fusion protein fully or partially rescued the observed defects of the PCNAK164R mutant. The expression of a PCNAK164R-ubiquitinK63R fusion protein, on which the formation of lysine 63-linked polyubiquitin chains is not possible, similarly rescued the cell cycle arrest, DNA damage sensitivity, reduction of translesion synthesis and increase of MMS-induced genomic mutagenesis. Template switching bypass was not affected by the genetic elimination of PCNA polyubiquitylation, but it was reduced in the absence of the recombination proteins BRCA1 or XRCC3. Our study found no requirement for PCNA polyubiquitylation to protect cells from replication-stalling DNA damage.

Genome Sequence Analysis of Mycoplasma sp. HU2014, Isolated from Tissue Culture.

The draft genome sequence of a novel Mycoplasma strain, designated Mycoplasma sp. HU2014, has been determined. The genome comprises 1,084,927 nucleotides and was obtained from a mycoplasma-infected culture of chicken DT40 cells. Phylogenetic analysis places this taxon in a group comprising the closely related species Mycoplasma yeatsii and Mycoplasma cottewii.

[Validation of the Hungarian version of the Strengths and Difficulties Questionnaire in an adolescent clinical population]. / A Képességek és Nehézségek Kérdoív (SDQ-Magy) validálása serdülokorú klinikai populációban.

INTRODUCTION: The short Strengths and Difficulties Questionnaire (SDQ) available in parent, teacher and self-report versions, is used world-wide for assessing and screening childhood behavior and mental problems, as part of clinical assessments, therapy outcome evaluations, and research tool. The aim of the present study was to extend the use of Hungarian version to a clinical sample, to examine the clinical cut-off values suggested previously on the basis of a normative sample, and to test the questionnaire's sensitivity in differentiating between different psychiatric disorders. METHOD: The parent and self-report versions of the five scale SDQ-Magy questionnaire was filled in by 716 parents and their children admitted to Vadaskert Child Psychiatry and Outpatient Clinic. Clinical (ICD) diagnoses were determined during psychiatric examination. RESULTS: With a few exceptions, internal consistencies of the scales were satisfactory (0,55-0,79), the parent version showing greater reliability compared to the self-report version. Children's age, gender, and parents' level of education had some effect on the scale scores. The questionnaire's Total problem and symptom scale scores were very effective in discriminating between the control and the clinical sample. In the clinical sample, parents rated their children's behavior and mental problems as more severe. Profiles of scale scores distinguished the wider internalizing, externalizing, and co-morbid diagnostic categories, as well as the eight specific diagnoses. Based on the normal-abnormal cut-off values proposed earlier (Turi et al., 2011), the great majority of clinical cases were screened by the questionnaire. CONCLUSIONS: The Hungarian version of the SDQ proved applicable in a clinical sample. Psychometric properties, variances due to age, gender and informant were consistent with international experiences. SDQ profiles related to clinical diagnoses and their difference from the control group show the sensitivity and discriminative power of the questionnaire, while the screening ability based on clinical cut-offs also supports the clinical use of the questionnaire.

[Psychometric parameters of the Hungarian version of Mothers' Object Relations Scales - Short Form (H-MORS-SF) in a large sample]. / A röviditett Szulö-Csecsemö Kapcsolat Skála magyar valtozátnak (H-MORS-SF) pszichometriai mutatói nagy mintán.

INTRODUCTION: The Mothers' Object Relations Scale (MORS) was developed by John M. Oates (Open University, Milton Keynes, UK) in the late 1990s. The MORS is an appropriate instrument for gathering parental perceptions about the child and the parent-infant relationship. The questionnaire and its short form were improved further and validated in British and Hungarian samples in the beginning of the 2000s and the questionnaire was used in several applied studies in the UK where its predictive validity was further confirmed. METHODS: The development and validation phases were based on small samples. The diverse social-demographic characteristics of the For Healthy Offspring project, allowed for further testing the reliability and validity of the Hungarian short-form in a large (n=1164) sample. RESULTS: High internal consistency was found in the original and the imputed data obtained from parents of 0-3-year old children for both of H-MORS-SF dimensions: Invasion and Warmth. The scales had interpretable and systematic cross-correlations with measures of infant temperament (IBQ-R, ECBQ) and mental state (DS1K) of both parents. These results confirm and exceed the previous results based on small samples. CONCLUSION: Given the convincing psychometric indicators and its fast and simple usage, the H-MORS-SF can be considered as an effective preventive screening test for monitoring the developing parent-infant relationship, therefore we suggest its use for professionals working in developmental psychology, child health and social fields.

[Further examination of the Strengths and Difficulties Questionnaire (SDQ-Magy) in a community sample of young adolescents]. / A Képessgék es Nehézségek Kérdoiv (SDQ-Magy) tovabbi vizsgalata nem-klinikai mintan, fiatal serdulok köreben.

INTRODUCTION: The Strengths and Difficulties Questionnaire (SDQ-Magy) is a brief instrument suitable for assessing and screening childhood behavior and mental problems, available in parent and teacher, and from 11 years of age, self-report versions. The aim of the present study was to extend our previous investigation in a community sample to an older age group, to examine its psychometric properties, and to assess cross-informant agreements and differences, effects of gender and age, as well as to determine cut-off points between normal and abnormal scores within the community sample. METHODS: Parent, teacher, and self-report questionnaire data were collected on 286 pupils of 12-17 years of age. RESULTS: With a few exceptions, internal consistencies of the scales were satisfactory, the teacher-reports showing the highest reliability. Regarding scale means, there were few significant differences from the British normative data. Correlations among problem scales were weak to moderate, and only self and parent reports showed moderate interrater agreements. Boys scored higher on Conduct problems and Hyperactivity scales, girls were reported to show more Prosocial behaviors and Emotional symptoms. Children's age affected teacher reports only, showing an increase of problems with age. We propose cut-off values for separating normal and clinical ranges. CONCLUSIONS: Psychometric properties and inter-correlations of the SDQ scales, as well as gender differences are consistent with the previous Hungarian and international studies. Rater differences found on certain scales suggest contextual effects on problematic behaviors, discrepancies between informants, and their potentially importance for researchers and mental health professionals.

[Screening childhood behavior problems using short questionnaires II.: The Hungarian version of the SWAN-scale (Strength and Weakness of ADHD-symptoms and Normal-behavior) for screening attention deficit/hyperactivity disorder]. / Gyermekkori viselkedési prblémák felismerésének lehetoségei rövid kérdoívekkel II: A SWAN (Strength and Weakness of ADHD-symptoms and Normal-behavior) skála magyar változata a figyelemhiányos/hiperaktivitás zavar szurésére.

INTRODUCTION: The SWAN (Strength and Weakness of ADHD-symptoms and Normal-behavior) Questionnaire is a short instrument suitable for screening attention deficit/hyperactivity disorder. Its completion by parents or teachers requires a few minutes. Positive re-wording of attention- and activity-related behaviors and the extended 7- point rating scale anchored to average behavior make the instrument especially suitable for normal populations. Here, we report the Hungarian version of SWAN and compare its scales with relevant scales of the Child Behavior Checklist (CBCL) and the Strengths and Difficulties Questionnaire (SDQ). METHOD: Questionnaire data were collected from parents of 156 six-year-old children in a community sample. Of the 156 children 89 were participants of the longitudinal Budapest Infant-Parent Study (BIPS). RESULTS: Internal consistency of the Hungarian SWAN scales was excellent (Cronbach alfa: 0.87-0.93), correlations with relevant CBCL and SDQ scales were as expected (0.40-0.49 and 0.67-0.74) showing that the short questionnaire was suitable for detection of attention and hyperactivity problems. SWAN scores in our sample were normally distributed and scale means were also similar to foreign studies. CONCLUSIONS: These initial Hungarian data confirm international experience with the SWAN. Psychometric indices, distribution of scale scores in the sample and across the sexes were consistent with foreign studies. Following collection of normative data, the Hungarian version of SWAN might be suitable for assessing mental health of children and adolescents, and for screening problem cases. Due to the wider range and the normal distribution of scores, SWAN provides a more suitable phenotype for genetic studies, than symptom scales.

Environmental and genetic influences on early attachment.

Attachment theory predicts and subsequent empirical research has amply demonstrated that individual variations in patterns of early attachment behaviour are primarily influenced by differences in sensitive responsiveness of caregivers. However, meta-analyses have shown that parenting behaviour accounts for about one third of the variance in attachment security or disorganisation. The exclusively environmental explanation has been challenged by results demonstrating some, albeit inconclusive, evidence of the effect of infant temperament. In this paper, after reviewing briefly the well-demonstrated familial and wider environmental influences, the evidence is reviewed for genetic and gene-environment interaction effects on developing early attachment relationships. Studies investigating the interaction of genes of monoamine neurotransmission with parenting environment in the course of early relationship development suggest that children's differential susceptibility to the rearing environment depends partly on genetic differences. In addition to the overview of environmental and genetic contributions to infant attachment, and especially to disorganised attachment relevant to mental health issues, the few existing studies of gene-attachment interaction effects on development of childhood behavioural problems are also reviewed. A short account of the most important methodological problems to be overcome in molecular genetic studies of psychological and psychiatric phenotypes is also given. Finally, animal research focusing on brain-structural aspects related to early care and the new, conceptually important direction of studying environmental programming of early development through epigenetic modification of gene functioning is examined in brief.

Age-related differences in distraction and reorientation in an auditory task.

Behavioral and event-related potential measures of distraction and reorientation were obtained from children (6 years), young (19-24 years) and elderly adults (62-82 years) in an auditory distraction-paradigm. Participants performed a go/nogo duration discrimination task on a sequence of short and long (50-50%) tones. In children, reaction times were longer and discrimination (d') scores were lower than in adults. Occasionally (15%), the pitch of the presented tones was changed. The task-irrelevant feature variation resulted in longer reaction times and lower d' scores with no significant differences between the three groups. Task-irrelevant changes affected the N1 amplitude and elicited the mismatch negativity, N2b, P3 and reorienting negativity (RON) sequence of event-related brain potentials. In children, the P3 latency was the same as in young adults. However the RON component was delayed by about 100ms. In the elderly, P3 and RON were uniformly delayed by about 80ms compared to young adults. This pattern of results provides evidence that distraction influences different processing stages in the three groups. Restoration of the task-optimal attention set was delayed in children, whereas in the elderly, the triggering of involuntary attention-switching required longer time.

[Screening childhood behavior problems using short questionnaires I.: the Hungarian version of the Strengths and Difficulties Questionnaire]. / Gyermekkori viselkedési problémák felismerésének lehetoégei rövid kérdoívekkel I: a Strengths and Difficulties Questionnaire magyar változata.

INTRODUCTION: The Strengths and Difficulties Questionnaire (SDQ) is a short questionnaire suitable for screening childhood behavior problems. Completing the questionnaire requires 5 minutes of parents' or teachers' time. The scales of the original version showed good agreement with relevant scales of the much longer Child Behavior Checklist (CBCL). In this study, we report the use of the Hungarian version of the SDQ and results of a comparison between the scales of the SDQ and the CBCL. METHODS: Questionnaire data were collected from the parents of 156 six-year-old children from a community sample. Of the 156 children, 89 were participants of the longitudinal Budapest Infant-Parent Study (BIPS). RESULTS: Internal consistency of the Hungarian SDQ scales were moderate-satisfactory (0.43-0.70), correlations between the relevant SDQ and CBCL scales were as expected (0.41-0.65), showing that the short questionnaire was equally suitable for the detection of problems. In our six-year-old age group, the mean of the total problem scores (11.0) was much higher than the means measured in other Western European countries and North-America. However, in developing countries, such as Brazil or China, levels were higher and closer to the Hungarian mean score. CONCLUSIONS: These initial Hungarian data confirm international experience with the SDQ, i.e., psychometric indices and the distribution of scale scores across the sexes are consistent with the results of foreign studies. On the other hand, a cautious generalization of our results indicates a higher level of problems in Hungary than in Western Europe and other developed countries. We think that following further collection of normative data the Hungarian version of the SDQ, which will take only a few minutes to complete, will be suitable for assessing the mental health of children and adolescents, and for the quick screening of problematic cases.

Dopaminergic candidate genes in Tourette syndrome: association between tic severity and 3' UTR polymorphism of the dopamine transporter gene.

Multiple evidence suggests an involvement of the dopamine neurotransmitter system in Tourette syndrome (TS). Therefore, dopaminergic candidate genes are in the center of genetic association analyses of TS. In this study, 103 TS patients and their parents have been characterized for different dopamine-related polymorphisms including the 48 bp variable number of tandem repeats (VNTR) of the dopamine D4 receptor (DRD4) gene, the 40 bp VNTR of the dopamine transporter (DAT1, SLC6A3) gene and the Val158Met polymorphism of the catechol-O-methyltransferase (COMT) gene. In addition, the 120 bp duplication and three single nucleotide polymorphisms (SNPs) were assessed in the promoter region of the DRD4 gene. The -616G allele and the 2-G-A-C haplotype (i.e., the 2-repeat form of the 120 bp sequence approximately -616G approximately -615A approximately -521C combination) were preferentially transmitted, however, these results did not remain significant after correction for multiple testing. Case-control analyses have also been carried out, resulting in negative findings. On the other hand, using a dimensional approach, the DAT1 40 bp VNTR showed an association with the peak tic-severity as measured by the Yale Global Tic Severity Scale. Patients with at least one copy of the 9-repeat allele had significantly more severe symptoms than individuals with the homozygous 10/10 genotype (P = 0.002). In summary, allele frequencies did not differ between cases and controls, but DAT1 genotype accounted for variations of tic severity within the TS group.

Infant genotype may moderate sensitivity to maternal affective communications: attachment disorganization, quality of care, and the DRD4 polymorphism.

Disorganized attachment is an early predictor of the development of psychopathology in childhood and adolescence. Lyons-Ruth et al. (1999) developed the AMBIANCE coding scheme to assess disrupted communication between mother and infant, and reported the link between maternal behavior and disorganized attachment. The Hungarian group found an association between a polymorphism of the DRD4 gene and disorganized attachment (Lakatos et al., 2000; 2002; Gervai et al., 2005). The present collaborative work investigated the interplay between genetic and caregiving contributions to disorganized attachment. 138 mother-infant dyads, 96 from a Hungarian low-social-risk sample and 42 from a US high-social-risk sample, were assessed for infant disorganized attachment behavior, for DRD4 gene polymorphisms, and for disrupted forms of maternal affective communication with the infant. In accord with literature reports, we found a robust main effect of maternal AMBIANCE scores on infant disorganization. However, this relation held only for the majority of infants who carried the short form of the DRD4 allele. Among carriers of the 7-repeat DRD4 allele, there was no relation between quality of maternal communication and infant disorganization. This interaction effect was independent of degree of social risk and maternal DRD4 genotype.

[Evaluation of major life events among Hungarian mothers rearing small children]. / Jelentös életesemények megítélése kisgyermekeket nevelö magyar anyák körében.

In this study we present LCU (life change unit) and quality rank orders of 92 significant life events as perceived by 184 Hungarian mothers caring for young children. For data collection, a modified Hungarian version of the Social Readjustment Rating Questionnaire (Miller and Rahe, 1997) was used. As with the American results, we could find no significant demographic effects in our sample on the LCU and quality ranking of the events. Although the Hungarian and the American LCU rank orders of 1997 showed high correlation, the LCU mean of the 63 common items was significantly lower for Hungarian mothers than for American ones. In addition, the stressfulness of some specific life events was perceived differently in the two cultures. The rank order of several life events obtained in an earlier Hungarian study (Tringer and Veér, 1977) allowed us to make a chronological comparison. The correlation of the rank orders of the 32 common items used in the two Hungarian data sets almost 30 years apart was somewhat lower than the association of the present Hungarian rank orders and American ones of 1997. In addition, some life events were ranked quite differently in the two Hungarian studies. Thus, perception of significant life events might differ by culture and by time and this observation should be taken into account in the design of research projects and mainly in their evaluation.

Association between dopamine D4 receptor (DRD4) gene polymorphisms and novelty-elicited auditory event-related potentials in preschool children.

We investigated associations of the exon III repeat and the -521 C/T polymorphisms of the DRD4 gene with novelty-elicited auditory ERP components and behavioral resistance to distraction in 57 healthy, typically developing 6-year-old children. Dopamine-related gene polymorphisms have previously been linked to processes directing focused attention. We did not find associations between the 7-repeat allele or the T.7 haplotype and the early ERP responses suggesting that DRD4 polymorphisms did not affect the detection of novelty. However, the same polymorphisms affected the late negative components (LN1 and LN2). Late negativities elicited by deviant and novel sounds have been regarded as reflecting reorientation after distraction or additional processing of new information. Children carrying the T.7 haplotype had significantly smaller LN1 and LN2 amplitudes. The presence of the T.7 haplotype also significantly enhanced behavioral resistance to distraction. We suggest that less distraction in T.7 carriers led to less reorienting activity (reflected by the LN components). We also speculate that activation of less sensitive and fewer D4 receptors (as with the T.7 haplotype) is less effective in modulating GABAergic inhibitory signaling, which in turn is reflected in smaller LN amplitudes.

Linkage analysis and molecular haplotyping of the dopamine D4 receptor gene promoter region.

OBJECTIVES: Polymorphic regions of the dopamine D4 receptor gene and its promoter region are in the focus of psychogenetic association studies. Besides the accurate phenotype characterization, highly reliable genotyping methods are also of outstanding importance in these works. METHODS: DNA samples of 598 healthy unrelated Caucasian individuals were used to validate the described molecular haplotyping methods and to determine the allele, genotype and haplotype frequencies and the linkage disequilibrium between the polymorphisms of the dopamine D4 receptor promoter region. RESULTS: We described a double genotyping system for the -521CT and -616CG polymorphisms, using a polymerase chain reaction restriction fragment length polymorphism or an allele-specific amplification. Allele and genotype frequencies of the novel -615AG single-nucleotide polymorphism are also determined (-615G=13.21%). For molecular haplotyping of the three single-nucleotide polymorphisms and a 120-bp duplication polymorphism, the allele-specific amplification was combined with restriction digestion. The results of the elaborated haplotyping methods were validated by molecular haplotyping of cloned fragments. CONCLUSIONS: The developed methods have been arranged into an 'economic' protocol that might be extended for higher reliability with a double haplotyping ('full mode'). Despite the close proximity of these sites, only a moderate linkage was found between the -615AG and -616CG (Delta(2)=0.162), between the -616AG and -521CT (Delta(2)=0.0221) and between the -615AG and -521CT single-nucleotide polymorphisms (Delta(2)=0.0346). The 120-bp duplication was shown to be in linkage equilibrium with any of the three single-nucleotide polymorphisms. Applications of these results should accelerate psychogenetic association studies of the dopamine D4 receptor gene.