Functional PAX-6 gene-linked polymorphic region: potential association with paranoid schizophrenia.

BACKGROUND: Early differentiation of the nervous system and adult CNS neuroplasticity is modulated by PAX-6. We have shown previously that a highly polymorphic, functional AC/AG repeat in the 5' regulatory region of the gene showed significantly increased promoter activity, if containing > or = 29 repeats, and that the heterozygous genotype (< or = 28/> or = 29) revealed increased mRNA PAX-6 levels in human brain tissue compared to the homozygous short variant. METHODS: In a case-control study of 655 unrelated individuals, allele frequencies and genotype distributions of the functional PAX-6 promoter polymorphism were investigated comprising patients with DSM-IV schizophrenia, patients with affective disorders, and population controls. RESULTS: No allelic or genotypic association of the PAX-6 promoter polymorphism to affective disorder or to schizophrenia as one disease entity was observed. After subtyping schizophrenia into paranoid and nonparanoid forms, potential evidence was found for a genotypic association of the high-activity variant with the paranoid subtype of schizophrenia (p = .02). The estimated odds ratio was 1.7 (95% CI .98 to 2.95) for those heterozygous and 1.4 (95% CI .82 to 2.42) for those heterozygous or homozygous for the high-activity variant compared to the homozygous low-activity variant. CONCLUSIONS: Our finding indicates that early developmental genes may be involved in the etiopathogenesis of schizophrenia subtypes via variable transcriptional regulation in the developing and adult human brain.

Phenomenon of life span instability in Drosophila melanogaster: II. Change in rhythm of natural variations of life span after single exposure to gamma-irradiation.

The dynamics of life span (LS) have been studied in successive generations of postirradiation and control groups of Drosophila melanogaster, strain D-32, after a single exposure to Co60 gamma-quantum irradiation. It has been shown using mathematical procedures that in all postirradiation generations, with one exception, survival curves retain their canonical shape. This is indicative of the unchangeable nature of LS distribution. The mean LS of the progeny of irradiated parents either coincides with control values or can be higher or lower. Moreover, single irradiation results in an altered time-scanning of LS variations in successive generations as compared with controls. The possible origin of LS instability is discussed.

Phenomenon of life span instability in Drosophila melanogaster: I. Nonrandom origin of life span variations in successive generations.

The dynamics of life span (LS) and fecundity in Drosophila melanogaster, strain D-32, were analyzed in a series of successive generations. Highly reliable variations in both fitness components were found. On initial inspection the variations would be characterized as random or irregular wherein mean values differed up to threefold. The variance in longevity is greater in females than in males. By use of mathematical procedures we have shown a scale regularity in LS distributions for all generations. Such regularity, in spite of considerable differences in absolute values (mean and maximum LS), suggested that the origin of LS instability is nonrandom.

Association analysis of the functional monoamine oxidase A gene promoter polymorphism in psychiatric disorders.

Functional characterization studies revealed that transcriptional activity of the human monoamine oxidase A (MAOA) gene is modulated by a polymorphic repetitive sequence located approximately 1.2 kb upstream of the ATG codon. To investigate the possible influence of the allelic variants of the MAOA gene-linked polymorphic region (MAOA-LPR) on the genetic predisposition to psychiatric disorders, we have performed a case-control association study. 174 patients with affective disorders and 258 patients with schizophrenia according to DSM-IV, as well as 229 population controls were tested. Statistical analysis showed no significant differences in allele or genotype frequencies between control and patient groups. Our results suggest that there is no association between MAOA-LPR genotype and susceptibility to recurrent major depression, bipolar disorder, and schizophrenia in our population.

Regulation of PAX-6 gene transcription: alternate promoter usage in human brain.

We have isolated and characterized the 5'-flanking regulatory region of the human PAX-6 gene. Mapping of transcription initiation sites revealed the existence of an additional non-coding 5' exon, exon 1A. Functional analyses indicated that PAX-6 transcription is regulated by two distinct promoters, A and B, resulting in alternative transcription of exon 1A or 1B and joint transcription of exons 2 to 13. While a single initiation site was identified for exon 1A, transcription of exon 1B appears to be initiated from more than one site downstream of the promoter B-associated TATA motif. Multiple potential binding sites for transcription factors were found in the regions of promoter A and B. Although a 1.1-kb fragment of promoter A and a 1.5 kb fragment of promoter B, which had been fused to a reporter gene and transiently expressed in cell lines, displayed constitutive promoter activity, transcription of PAX-6 driven by promoter B was considerably higher than by promoter A in various regions of human postmortem brain. Transcript PAX-6B was primarily expressed in cerebellar cortex, whereas relatively low concentrations were detected in other brain areas. Functional dissection by serial deletions revealed several clusters of both activating elements and cell-selective silencers within the regulatory regions upstream of exon 1A and 1B. Coexpression of the promoter B constructs with a vector expressing PAX-6 modulated promoter B activity, thus indicating autoregulation by PAX-6 transcription. In conclusion, our findings suggest that PAX-6 transcription is regulated by alternate usage of promoter A and B, and that in adult human brain expression of PAX-6 is primarily controlled by promoter B. Alternate promoter usage and differential PAX-6 transcription are likely to play a critical role in brain development and neuroplasticity.

Serotonin transporter function is modulated by brain-derived neurotrophic factor (BDNF) but not nerve growth factor (NGF).

The serotonin transporter (5-HTT) regulates serotonergic neurotransmission by determining the magnitude and duration of serotonergic responses. We have recently described a polymorphism in the 5-HTT gene promoter (5-HTTLPR) which influences the function of the 5-HTT and is associated with several psychiatric disorders. Immortalized B lymphocytes express the 5-HTT, and a B lymphocyte line has been shown to express the receptor for brain-derived neurotrophic factor, trkB. Since brain-derived neurotrophic factor (BDNF) is a specific growth and differentiation factor for serotonergic neurons, we assessed whether BDNF is able to modulate 5-HTT function in B lymphoblasts. Nerve growth factor (NGF), another neurotrophin which acts via the trkA receptor, was also studied. Eight immortalized B lymphoblast lines were generated and genotyped for the 5-HTTLPR. After treatment with BDNF or NGF, 5-HT uptake and proliferation of the cell lines were assessed. Two of the B cell lines showed a dose-dependent reduction of 5-HT uptake after exposure to BDNF. Both of these cell lines were homozygous for the long allele of the 5-HTTLPR. NGF did not influence 5-HT uptake or cellular proliferation in any of the cell lines. Thus, BDNF but not NGF may influence 5-HT uptake in some B lymphocytes. The fact that regulation of the 5-HTT was observed preferentially in cells of the long/long genotype indicates that presence of a short allele confers reduced regulatory capacity on the 5-HTT. In conclusion, B lymphoblasts represent a practical model for functional regulation of the 5-HTT by neurotrophins in serotonergic neurons.

Enhancement of serotonin transporter function by tumor necrosis factor alpha but not by interleukin-6.

Serotonin (5-HT) is a prime candidate for studies of the interaction between the nervous and immune systems, since it is both an important neurotransmitter and released at high concentrations at sites of inflammation. Serotonergic neurotransmission is regulated by the 5-HT transporter (5-HTT), which determines the magnitude and duration of serotonergic responses. Since tumor necrosis factor alpha (TNF-alpha) and interleukin-6 are two inflammatory mediators that are central to the initiation of inflammation, we studied the impact of these cytokines on the 5-HTT. As model system we used a cell line which constitutively expresses the 5-HTT, namely the choriocarcinoma cell line JAR. We found that TNF-alpha enhances 5-HT uptake, with a doubling of the maximal velocity of uptake. Interleukin-6, on the other hand, had no effect. We thus show for the first time that the cytokine TNF-alpha modulates 5-HTT function. Furthermore, we propose a molecular mechanism for this effect. Since both 5-HT and TNF-alpha are elevated at sites of inflammation, TNF-alpha may act to renormalize 5-HT levels by way of its effect on the 5-HTT. This is especially important for the central nervous system, where the TNF-alpha effect shown here can aid in preventing disturbances of serotonergic neurotransmission.

Short CAG repeats within the hSKCa3 gene associated with schizophrenia: results of a family-based study.

In a family-based association study we investigated transmission of a multiallelic CAG repeat in a novel neuronal potassium channel gene, hSKCa3, in 59 parent/ offspring trios. In contrast to recent reports of an association of moderately large repeats with schizophrenia in case-control studies, our findings indicate that short CAG repeats (< or=19 repeats) are transmitted at an increased frequency to schizophrenic offspring (p=0.014), particularly among familial cases (p=0.007). No evidence for a parent-of-origin effect was found. Multiallelic TDT procedure showed no association of individual CAG repeats to schizophrenia. Further studies using family-based designs should clarify whether hSKCa3 is a susceptibility factor to schizophrenia or co-segregates with a major disease gene in tight linkage.

Association analysis of a regulatory promoter polymorphism of the PAX-6 gene with idiopathic generalized epilepsy.

The PAX-6 gene is a member of the paired-box-containing (PAX) gene family, encoding a transcriptional activator, that plays an important role in the development of the central nervous system. The present association study tested the hypothesis that length variation of a novel regulatory dinucleotide repeat polymorphism in the promoter region of the PAX-6 gene (PAX-6 gene-linked polymorphic region, PAX-6LPR) confers susceptibility to the epileptogenesis of common subtypes of idiopathic generalized epilepsy (IGE). The repeat length of the regulatory dinucleotide repeat polymorphism was assessed in 354 German control subjects and 125 German IGE patients, comprising 70 patients with juvenile myoclonic epilepsy (JME) and 55 patients with an idiopathic absence epilepsy (IAE). The allelic distribution of the PAX-6LPR did not deviate significantly between the controls and the IGE patients (Wilcoxon Rank-Sum test: P > 0.76), or both subgroups of either JME patients (P > 0.78) or IAE patients (P > 0.87). Our results do not provide evidence that length variation of the polymorphic dinucleotide sequence in the PAX-6LPR contributes a frequent and relevant effect to the pathogenesis of common subtypes of IGE.

Association of a regulatory polymorphism in the promoter region of the monoamine oxidase A gene with antisocial alcoholism.

We analyzed a novel functional 30-bp repeat polymorphism in the promoter region of the X-chromosomal monoamine oxidase A gene (MAOA) to test whether length variation of the repeat polymorphism contributes to variation in the individual vulnerability to antisocial behavior and liability to alcohol dependence. The repeat number (3-5) of the MAOA polymorphism was assessed in 488 male subjects of German descent, a sample comprising 185 psychiatrically screened control subjects and 303 alcohol-dependent subjects including 59 alcoholics with antisocial personality disorder. The frequency of the low-activity 3-repeat allele was significantly increased in 59 antisocial alcoholics compared to 185 control subjects (51 vs. 35%; P = 0.031) and to 244 alcoholics without antisocial personality disorder (51 vs. 32%; P = 0.008), respectively. We found no significant difference in the frequency of the 3-repeat allele between 244 alcoholics without an antisocial personality disorder and the control subjects. Our findings suggest that the low-activity 3-repeat allele of the MAOA promoter polymorphism confers increased susceptibility to antisocial behavior rather than alcohol dependence per se in alcohol-dependent males.

Polymorphic MAO-A and 5-HT-transporter genes: analysis of interactions in panic disorder.

Recurrent panic attacks, anticipatory anxiety and phobic avoidance characterise panic disorder. The influence of genetic factors on liability to the disease has been the object of several linkage and association studies and appears to relate to an oligo- or polygenic rather than a monogenic mode of inheritance. Recently, an excess of high activity monoamine oxidase A (MAO-A) gene promoter alleles was found in female patients with panic disorder. An analysis of possible synergistic effects of the MAO-A gene promoter variant and the short serotonin transporter (5-HTT) gene promoter variant in panic disorder was performed in a German and an Italian sample (combined panic disorder n = 144, combined controls n = 175). There was no significant difference in odds ratios, suggesting that the observed increase of genetic liability by the long MAO-A gene promoter allele is not modified by the 5-HTT gene promoter polymorphism.

[Role of the homologous translocation, located on the Y-chromosome, in mutagenesis of the double super-unstable system in Drosophila melanogaster males]. / Rol' gomologichnoi translokatsii, raspolozhennoi na Y-khromosome, v mutageneze dvoinoi supernestabil'noi sistemy y samtsov Drosophila melanogaster.

Main features of mutagenesis induced in the double super-unstable system in Drosophila melanogaster males and females was studied earlier. Very different y- and sc-alleles arose under the mutagenesis. These are differed one from another by the expression of yellow and scute loci. However, it was shown that the female's mutagenesis very differed from the male's one. The structure of a homologous chromosome has played an important role in the mutagenesis in Drosophila females. We found here that the homologous translocation in Y chromosome led to double decrease of the frequency of the mutagenesis in males. It is proposed that interconnections between two homologous insertions are essential in the recombinational processes by which alleles change. The nature and mechanism of the sub-lethal allele's formation in the presence of translocation in Y chromosome is discussed.

[A new type of locus-specific instability in Drosophila melanogaster not dependent on P-M hybrid dysgenesis]. / Novyi tip lokusspetsifichnoi nestabil'nosti u Drosophila melanogaster, ne zavisiashchii ot P-M gibridnogo disgeneza.

In analysis of instability at loci yellow and scute, induced by introduction of the active P-element transposase into the genome of line y2nsscme, alleles of these loci with transposase-independent instability were selected. The mutation rate of these alleles reached tens of percent. Two types of transposase-independent instability were described: (1) instability limited to the scute locus and (2) instability at both yellow and scute loci.

[Effect of mei mutations on the processes occurring in the double super-unstable system at the yellow and scute loci in Drosophila melanogaster]. / Vliianie mei-mutatsii na protsessy, proiskhodiashchie v dvoinoi supernestabil'noi sisteme v lokusakh yellow i scute u Drosophila melanogaster.

The influence of meiotic mutations on the mutation changes in the double super-unstable system in the yellow and scute loci of Drosophila melanogaster was studied. The mei-41D5 and mei-218 mutations changed the spectrum and frequency of mutagenesis in males of the y2nsscme strain, in contrast to the postulate that meiotic mutations do not interfere with male recombination in D. melanogaster. These mutations also changed the frequency and spectrum of mutagenesis in females. In particular, they inhibited mutagenesis at early stages of ovogenesis. Meiotic conversion did not change specifically by mei mutations. At the same time, the mei-41D5 mutation increased all recombination processes in meiosis. The results obtained indicated the involvement of genetic recombination in mutation changes occurring in the double super-unstable system. Therefore, the latter may be successfully used in studies of the role of different genes and their products in recombination.

[[A dual system of superinstability at the yellow and scute loci in Drosophila melanogaster]. / Dvoinaia sistem supernestabil'nosti v lokusakh yellow i scute Drosophila melanogaster.

We have characterized the phenomenon of super-unstability in yellow and scute loci of Drosophila melanogaster. A few derivatives with different combinations of yellow and scute phenotypes appeared after dysgenic cross between potentially super-unstable stock y2nsscme and P[ry+, (delta 2-3)] (99B). Essentially, the double alterations of yellow and scute phenotypes constitute more than 40% of all derivatives. Most frequently the mutations in both yellow and scute loci change coordinately giving rise to the y+nssc+ allele. Lethal derivatives were not observed. The spectrum and the frequency of the y2nsscme mutagenesis in females differ considerably from the analogous in males. Possibly, the neutral homologous chromosome in females changes contacts between two insertions, and because of that mutagenesis changes too. Thus, all alterations in the double super-unstable system seem to be connected with the recombination between two unstable insertions A genetic exchange may by initiated by a double-strand breaks induced by the transposase of the ends of insertions.

[Molecular analysis of integration sites of the drosophila retrotransposon burdock]. / Molekuliarnyi analiz mest integratsii retrotranspozona drozofily repeinik (burdock).

Data on molecular analysis of the insertion sites of nine random copies of burdock retrotransposon are presented. The 12-bp consensus sequence of the insertion sites, YNNUTUTUYAYA (Y-pyrimidine; U-purine), was determined. Homology between the burdock sequence and ribosomal genes was revealed. Three copies of this element were located within the region of ribosomal repeats: one copy in the 18S RNA gene, and two copies in the same intergenic spacer region, in the so-called Alu-repeats of Drosophila, in different copies of ribosomal genes.

[Generation of Kruppel phenocopies by injecting into Drosophila embryos RNA complementary to mRNA in parallel orientation]. / Poluchenie fenokopii Kruppel pri in''ektsiiakh v émbriony drozofily RNK, komplementarnoi mRNK v parallel'nom napravlenii.

RNA preparations synthesized in vitro were used to study the influence of RNA interference on the Kruppel gene activity in Drosophila embryos. RNA complementary in parallel orientation to the mRNA fragment proved to induce the development of Kruppel phenocopies. The data obtained indicate that mechanisms of specific regulation of gene activity exist in Drosophila cells, which are sensitive to the formation of both parallel and antiparallel RNA-RNA duplexes that include mRNA of the corresponding gene.