Friedreich's ataxia: autosomal recessive disease caused by an intronic GAA triplet repeat expansion.

Friedreich's ataxia (FRDA) is an autosomal recessive, degenerative disease that involves the central and peripheral nervous systems and the heart. A gene, X25, was identified in the critical region for the FRDA locus on chromosome 9q13. This gene encodes a 210-amino acid protein, frataxin, that has homologs in distant species such as Caenorhabditis elegans and yeast. A few FRDA patients were found to have point mutations in X25, but the majority were homozygous for an unstable GAA trinucleotide expansion in the first X25 intron.

Drosophila melanogaster Models of Friedreich's Ataxia.

Friedreich's ataxia (FRDA) is a rare inherited recessive disorder affecting the central and peripheral nervous systems and other extraneural organs such as the heart and pancreas. This incapacitating condition usually manifests in childhood or adolescence, exhibits an irreversible progression that confines the patient to a wheelchair, and leads to early death. FRDA is caused by a reduced level of the nuclear-encoded mitochondrial protein frataxin due to an abnormal GAA triplet repeat expansion in the first intron of the human FXN gene. FXN is evolutionarily conserved, with orthologs in essentially all eukaryotes and some prokaryotes, leading to the development of experimental models of this disease in different organisms. These FRDA models have contributed substantially to our current knowledge of frataxin function and the pathogenesis of the disease, as well as to explorations of suitable treatments. Drosophila melanogaster, an organism that is easy to manipulate genetically, has also become important in FRDA research. This review describes the substantial contribution of Drosophila to FRDA research since the characterization of the fly frataxin ortholog more than 15 years ago. Fly models have provided a comprehensive characterization of the defects associated with frataxin deficiency and have revealed genetic modifiers of disease phenotypes. In addition, these models are now being used in the search for potential therapeutic compounds for the treatment of this severe and still incurable disease.

Mapping of Friedreich's ataxia locus by identification of recombination events in patients homozygous by descent.

The Friedreich's ataxia locus (FRDA) maps on chromosome 9q13. Genetic data, obtained from a small number of recombination events, indicated that the FRDA locus might be located centromeric to the D9S15/D9S5 linkage group, the most probable order being cen-FRDA-D9S5-D9S111-D9S15-D9S110-qter. Recently, new centromeric markers have been reported. Analysis of these markers allowed us to localize the recombination breakpoint in some of the recombinant families. However, only one proximal recombination has been found with these markers. To increase the genetic information from FRDA families, we have analyzed the centromeric markers FR1, FR2, FR7, FR8, and FR5 in patients homozygous by descent. These were ascertained because parents were consanguineous or because they were homozygous for the entire haplotype D9S15 or D9S111-D9S5-D9S411E-D9S202. Haplotype divergence for, at least, two contiguous markers was observed in two patients homozygous for the core D9S111-FR2 haplotype and in one third-degree consanguineous family homozygous for haplotype D9S411E-FR5. Interpretation of divergence as the result of ancient meiotic crossovers allowed the definition of three new recombination events which place the FRDA locus within the interval defined by markers D9S411E and FR8. A consanguineous family with first-cousin parents showed homozygosity only at D9S202 and FR2. Further investigations are needed to discern whether two different mutations are segregating in the family or whether two recombinations, one distal and one proximal, have taken place.

Evidence for a common origin of most Friedreich ataxia chromosomes in the Spanish population.

Haplotype analysis is a powerful approach to understand the spectrum of mutations accounting for a disease in a homogeneous population. We show that haplotype variation for 10 markers linked to the Friedreich ataxia locus (FRDA) argues in favor of an important mutation homogeneity in the Spanish population, and positions the FRDA locus in the region where it has been recently isolated. We also report the finding of a new single nucleotide polymorphism called FAD1. The new marker shows a very strong linkage disequilibrium with Friedreich ataxia (FA) in both the Spanish and French populations. suggesting the existence of an ancient and widespread FRDA mutations. Inclusion of FAD1 in the extended haplotype analysis has allowed to postulate that this main FRDA mutation could account for 50-90% of the disease chromosomes. The results indicate that FA, despite clinical heterogeneity, could have originated from a few initial mutations.

dfh is a Drosophila homolog of the Friedreich's ataxia disease gene.

A putative Drosophila homolog of the Friedreich's ataxia disease gene (FRDA) has been cloned and characterized; it has been named Drosophila frataxin homolog (dfh). It is located at 8C/D position on X chromosome and is spread over 1kb, a much smaller genomic region than the human gene. Its genomic organization is simple, with a single intron dividing the coding region into two exons. The predicted encoded product has 190 amino acids, being considered a frataxin-like protein on the basis of the sequence and secondary structure conservation when compared with human frataxin and related proteins from other eukaryotes. The closest match between the Drosophila and the human proteins involved a stretch of 38 amino acids at C-terminus, encoded by dfh exon 2, and exons 4 and 5a of the FRDA gene, respectively. This highly conserved region is very likely to form a functional domain with a beta sheet structure flanked by alpha-helices where the sequence is less conserved. A signal peptide for mitochondrial import has also been predicted in the Drosophila frataxin-like protein, suggesting its mitochondrial localization, as occurs for human frataxin and other frataxin-like proteins described in eukaryotes. The Drosophila gene is expressed throughout the development of this organism, with a peak of expression in 6-12h embryos, and showing a spatial ubiquitous pattern from 4h embryos to the last embryonic stage examined. The isolation of dfh will soon make available specific dfh mutants that help in understanding the pathogenesis of FRDA.

Localization of non-specific X-linked mental retardation gene (MRX73) to Xp22.2.

Clinical and molecular studies are reported on a family (MRX73) of five males with non-specific X-linked mental retardation (XLMR). A total of 33 microsatellite and RFLP markers was typed. The gene for this XLMR condition was been linked to DXS1195, with a lod score of 2.36 at theta = 0. The haplotype and multipoint linkage analyses suggest localization of the MRX73 locus to an interval of 2 cM defined by markers DXS8019 and DXS365, in Xp22.2. This interval contains the gene of Coffin-Lowry syndrome (RSK2), where a missense mutation has been associated with a form of non-specific mental retardation. Therefore, a search for RSK2 mutations was performed in the MRX73 family, but no causal mutation was found. We hypothesize that another unidentified XLMR gene is located near RSK2.

A possible association between the CCK-AR gene and persistent auditory hallucinations in schizophrenia.

Recent studies have suggested that DNA variations in the CCK-AR gene might predispose individuals to schizophrenia and particularly to auditory hallucinations (AH). The aim of this study is to assess the association between AH, using a specific scale for AH in schizophrenia (PSYRATS), and the CCK-AR polymorphism at 779 in a Spanish sample. A total of 105 DSM-IV schizophrenic patients with AH and 93 unrelated controls were studied. Twenty-two patients were considered as persistent auditory hallucinators, which showed similar clinical and demographic characteristic than patients with episodic AH, but with the exception of the PSYRATS values. The persistent AH group showed an excess of the A1 allele when was compared with episodic or control groups. Our data support the possible role of the CCK-AR gene in the development of persistent AH in schizophrenic patients.

The genetic contribution of the NO system at the glutamatergic post-synapse to schizophrenia: further evidence and meta-analysis.

NO is a pleiotropic signaling molecule and has an important role in cognition and emotion. In the brain, NO is produced by neuronal nitric oxide synthase (NOS-I, encoded by NOS1) coupled to the NMDA receptor via PDZ interactions; this protein-protein interaction is disrupted upon binding of NOS1 adapter protein (encoded by NOS1AP) to NOS-I. As both NOS1 and NOS1AP were associated with schizophrenia, we here investigated these genes in greater detail by genotyping new samples and conducting a meta-analysis of our own and published data. In doing so, we confirmed association of both genes with schizophrenia and found evidence for their interaction in increasing risk towards disease. Our strongest finding was the NOS1 promoter SNP rs41279104, yielding an odds ratio of 1.29 in the meta-analysis. As findings from heterologous cell systems have suggested that the risk allele decreases gene expression, we studied the effect of the variant on NOS1 expression in human post-mortem brain samples and found that the risk allele significantly decreases expression of NOS1 in the prefrontal cortex. Bioinformatic analyses suggest that this might be due the replacement of six transcription factor binding sites by two new binding sites as a consequence of proxy SNPs. Taken together, our data argue that genetic variance in NOS1 resulting in lower prefrontal brain expression of this gene contributes to schizophrenia liability, and that NOS1 interacts with NOS1AP in doing so. The NOS1-NOS1AP PDZ interface may thus well constitute a novel target for small molecules in at least some forms of schizophrenia.

Mesenchymal stromal-cell transplants induce oligodendrocyte progenitor migration and remyelination in a chronic demyelination model.

Demyelinating disorders such as leukodystrophies and multiple sclerosis are neurodegenerative diseases characterized by the progressive loss of myelin that may lead toward a chronic demyelination of the brain's white matter, impairing normal axonal conduction velocity and ultimately causing neurodegeneration. Current treatments modifying the pathological mechanisms are capable of ameliorating the disease; however, frequently, these therapies are not sufficient to repress the progressive demyelination into a chronic condition and permanent loss of function. To this end, we analyzed the effect that bone marrow-derived mesenchymal stromal cell (BM-MSC) grafts exert in a chronically demyelinated mouse brain. As a result, oligodendrocyte progenitors were recruited surrounding the graft due to the expression of various trophic signals by the grafted MSCs. Although there was no significant reaction in the non-grafted side, in the grafted regions oligodendrocyte progenitors were detected. These progenitors were derived from the nearby tissue as well as from the neurogenic niches, including the subependymal zone and dentate gyrus. Once near the graft site, the cells matured to myelinating oligodendrocytes. Finally, electrophysiological studies demonstrated that axonal conduction velocity was significantly increased in the grafted side of the fimbria. In conclusion, we demonstrate here that in chronic demyelinated white matter, BM-MSC transplantation activates oligodendrocyte progenitors and induces remyelination in the tissue surrounding the stem cell graft.

Association between neonatal temperament, SLC6A4, DRD4 and a functional polymorphism located in TFAP2B.

Genetic studies on human personality have provided little satisfactory results to date mainly because of the complexity of this trait. Neonatal temperament using observational measures is an alternative phenotype to approach genetics to human behavior. An association study was conducted on 117 Caucasian newborns. Their temperament was evaluated using the Neonatal Behavior Assessment Scale 48 h after birth. Thirteen polymorphisms in the SLC6A4, DRD4 and TFAP2B genes were genotyped. Linear regression was performed to analyze data, and Bonferroni correction was applied. To check the functional effect of the TFAP2B Indel Intron 2 polymorphism, reporter gene luciferase assays using a mouse cortical neural progenitor cell line and quantitative polymerase chain reaction (qPCR) studies in human post-mortem brain samples were performed. A significant association was found between 5-HTTLPR, 5-HTTLPR + rs25531 and TFAP2B Indel Intron 2 with Range of State cluster as well as an interaction between rs25531 and TFAP2B Indel Intron 2 with Range of State. DRD4 variable number tandem repeat exon 3 was associated with orientation. A 30% increase in the luciferase levels of the TFAP2B 5-repeat alleles compared with the 6-repeat alleles (P-value = 0.03) was found using the pGL3 promoter vector. The qPCR experiments showed the same trend as the in vitro studies, although no significant results were obtained. This study supports a role of the SLC6A4, DRD4 and TFAP2B genes in the temperament, including a gene-gene interaction between SLC6A4 and TFAP2B. It also provides evidence about an effect of the TFAP2B polymorphism in TFAP2B gene transcription.

Puff activity after heat shock in two species of the Drosophila obscura group.

When individuals of Drosophila guanche are submitted to heat shock, five new puffs are induced. These puffs usually do not appear during normal development. Comparing these results with those obtained in Drosophila subobscura, also belonging to the obscura group, differences between the induced puffing pattern of both species have been found.

Tirant is a new member of the gypsy family of retrotransposons in Drosophila melanogaster.

In this paper, we propose a consensus sequence for a putative complete Tirant retrotransposon. Several defective copies, as well as relevant sequences available in databases have been analyzed. The putative complete Tirant element is 8533 bp long, and presents all the structural features of a retrovirus-like transposable element of the gypsy family. It contains three ORFs (open reading frames) that encode putative products resembling the retroviral Gag, Pol, and Env proteins. Southern blot analyses show that complete and defective Tirant elements are widespread in Drosophila melanogaster. The different hybridization patterns observed in several natural populations of this species suggest that Tirant is an active element.

The banding pattern of polytene chromosomes of Drosophila guanche compared with that of D. subobscura.

A detailed map of the salivary gland chromosomes of Drosophila guanche is presented and compared to the standard gene arrangements of D. subobscura. Generally, the polytene chromosome banding patterns of the two species show a high degree of homology. Only Segment I of the sex chromosome (Chromosome A) shows marked differences. The banding pattern proposed for this segment in D. guanche could have originated from a cluster of overlapping inversions including A1 arrangement.

Differences in gene activity in a Drosophila species cluster belonging to the Obscura group.

The polytene chromosome puffing pattern of Drosophila madeirensis was established and compared with those of the related species D. subobscura and D. guanche. A total of 145 loci, active in some of the 12 developmental stages analysed, were described, 38 of which were found to form the puffing pattern characteristic to this species. Taking into account the number of puffs as well as the mean puff expression, D. madeirensis shows a similar activity level to D. guanche, both species being less active than D. subobscura. The low gene activity of D. madeirensis and D. guanche was explained as a consequence of their ecological characteristics.

Heat shock proteins in three related Drosophila species belonging to the obscura group.

The effect of heat shock on protein synthesis in three related Drosophila species belonging to the obscura group was analyzed on SDS-acrylamide gels. Four major heat shock proteins (hsps) were found in these species, in which synthesis reaches a maximum at 34 degrees C. Although the higher molecular weight proteins are conserved, differences in size were found for the small hsps in these species. By means of in situ hybridization using D. melanogaster probes for the small hsp genes, it was inferred that the small hsp genes of the obscura group species are clustered at the 27A locus in all three species.

Gene activity of polytene chromosomes in Drosophila species of the obscura group.

The polytene chromosome puffing patterns of Drosophila guanche were established and compared with those of Drosophila subobscura. A total of 150 loci, active in some of the 17 developmental stages studied, were described and 23 of them were found to form the characteristic puffing pattern of D. guanche. Taking into account the number of puffs as well as the gene activity of each chromosome and the total gene activity, D. guanche seems to be less active than D. subobscura. Although both species show a degree of homology in their puffing patterns lower than that found for sibling species, the degree of homology is stronger than that between species belonging to the same group but to different subgroups. Thus, D. guanche and D. subobscura must be considered as phylogenetically closely related species, belonging to the same subgroup.

Genetic analysis of heat shock response in three Drosophila species of the obscura group.

Heat shock response was investigated in three species of the obscura group of the Drosophila genus (D. subobscura, D. guanche, and D. madeirensis) by chromosome cytology analysis and [3H]uridine labeling. A set of eight puffs (2C, 15DE, 18C, 27A, 31CD, 85AB, 89A, and 94A) were induced after heat treatments in each of the three species; 18C, 27A, 89A, and 94A were the most heavily labeled in the autoradiograms after the induced conditions. From the in situ results using the major heat shock genes of D. melanogaster as a probe, it was inferred that the 18C, 94A, 89A, and 27A loci of the three obscura group species are homologous to D. melanogaster loci, which contain, HSP82, HSP70, HSP68, and HSPs encoding for the small heat shock proteins, respectively. When this organization was compared with that of D. melanogaster, fewer evolutionary changes, mainly gene duplications, were found to have occurred in the obscura group species than in the D. melanogaster group. In the three species analyzed in this work, as well as in the other Drosophila species studied, the heat shock genes are distributed on D and E Muller's elements, behaving as single copy genes that do not move around the genome.

Phylogenetic relationships between Drosophila subobscura, D. guanche and D. madeirensis based on Southern analysis of heat shock genes.

A Southern analysis of genomic DNA using Drosophila melanogaster probes for the major heat shock protein genes (Hsp82, Hsp70, Hsps encoding small proteins) was made to study the phylogenetic relationships between three Drosophila species belonging to the obscura group (D. subobscura, D. guanche, and D. madeirensis). The phylogenetic trees showed that D. madeirensis and D. subobscura are the most closely related species in the triad, while D. guanche is the most distantly related one. As in other Drosophila species, Hsp82 is a single copy gene in D. subobscura, D. guanche, and D. madeirensis, while Hsp70 and Hsps, which encode small proteins, are genic families. At least four sequences homologous to D. melanogaster Hsp70 were found in the obscura group species. These species have sequences which showed similarity with the four small Hsps of D. melanogaster.

Tirant: a new retrotransposon-like element in Drosophila melanogaster.

In this paper we report a new retrotransposon-like element of Drosophila melanogaster called Tirant. This sequence is moderately repeated in the genome of this species and it has been found to be widely dispersed throughout its distribution area. From Southern blot and in situ analyses, this sequence appears to be mobile in D. melanogaster, since its chromosome location and the hybridization patterns vary among the different strains analyzed. In this way, partial sequencing of Tirant ends suggests that it is a retrotransposon, since it is flanked by two LTRs. The presence of sequences homologous to Tirant has been also investigated in 28 species of the genus Drosophila by means of Southern analyses. These sequences were only detected in species from melanogaster and obscura groups. These data suggest that ancestral sequences of Tirant appeared after the Sophophora radiation and before the divergence of those groups.