Dlx homeobox genes play a crucial role in the migration and differentiation of the subpallial precursor cells that give rise to various subtypes of gamma-aminobutyric acid (GABA)-expressing neurons of the forebrain, including local-circuit cortical interneurons. Aberrant development of GABAergic interneurons has been linked to several neurodevelopmental disorders, including epilepsy, schizophrenia, Rett syndrome and autism. Here, we report in mice that a single-nucleotide polymorphism (SNP) found in an autistic proband falls within a functional protein binding site in an ultraconserved cis-regulatory element. This element, I56i, is involved in regulating Dlx5/Dlx6 homeobox gene expression in the developing forebrain. We show that the SNP results in reduced I56i activity, predominantly in the medial and caudal ganglionic eminences and in streams of neurons tangentially migrating to the cortex. Reduced activity is also observed in GABAergic interneurons of the adult somatosensory cortex. The SNP affects the affinity of Dlx proteins for their binding site in vitro and reduces the transcriptional activation of the enhancer by Dlx proteins. Affinity purification using I56i sequences led to the identification of a novel regulator of Dlx gene expression, general transcription factor 2 I (Gtf2i), which is among the genes most often deleted in Williams-Beuren syndrome, a neurodevelopmental disorder. This study illustrates the clear functional consequences of a single nucleotide variation in an ultraconserved non-coding sequence in the context of developmental abnormalities associated with disease.
"Since the GxE approach identifies differentially expressed genes that are candidates for mediating the interaction, we then sought to uncover underlying single nucleotide variants (SNVs) in cis-regulatory elements within the proximal promoters of these candidate genes. SNVs within the promoters are excellent candidates for regulating expression of the nearby altered genes, as regulatory SNVs can alter the location and function of enhancers and promoters (De Gobbi et al., 2006) that change transcriptional levels (Munkhtulga et al., 2010) in specific cell types (Poitras et al., 2010) and lead to allele-specific changes in expression (Azad et al., 2013). SNV studies narrowed the list to 16 genes that showed both a GxE interaction based on gene expression studies and an SNV driven to fixation in opposite directions in the proximal promoter. "
[Show abstract][Hide abstract] ABSTRACT: Maternal alcohol consumption inflicts a multitude of phenotypic consequences that range from undetectable changes to severe dysmorphology. Using tightly controlled murine studies that deliver precise amounts of alcohol at discrete developmental stages, our group and other labs demonstrated in prior studies that the C57BL/6 and DBA/2 inbred mouse strains display differential susceptibility to the teratogenic effects of alcohol. Since the phenotypic diversity extends beyond the amount, dosage and timing of alcohol exposure, it is likely that an individual's genetic background contributes to the phenotypic spectrum. To identify the genomic signatures associated with these observed differences in alcohol-induced dysmorphology, we conducted a microarray-based transcriptome study that also interrogated the genomic signatures between these two lines based on genetic background and alcohol exposure. This approach is called a gene x environment (GxE) analysis; one example of a GxE interaction would be a gene whose expression level increases in C57BL/6, but decreases in DBA/2 embryos, following alcohol exposure. We identified 35 candidate genes exhibiting GxE interactions. To identify cis-acting factors that mediated these interactions, we interrogated the proximal promoters of these 35 candidates and found 241 single nucleotide variants (SNVs) in 16 promoters. Further investigation indicated that 186 SNVs (15 promoters) are predicted to alter transcription factor binding. In addition, 62 SNVs created, removed or altered the placement of a CpG dinucleotide in 13 of the proximal promoters, 53 of which overlapped putative transcription factor binding sites. These 53 SNVs are also our top candidates for future studies aimed at examining the effects of alcohol on epigenetic gene regulation.
Frontiers in Genetics 06/2014; 5:173. DOI:10.3389/fgene.2014.00173
"It is plausible that the disruption or deletion of UCEs may have a strong deleterious impact on survival in complex natural environments but would have very little effect on fitness under controlled lab conditions as observed for non-coding RNA BC1 . Indeed, an SNP in an ultraconserved regulatory sequence is linked with Dlx5/Dlx6 expression in the forebrain . Another paper reported an enrichment of UCEs in chromosomal rearrangements, especially pathogenic deletions, identified in 200 people with idiopathic neurodevelopmental disorders . "
[Show abstract][Hide abstract] ABSTRACT: Metazoan genomes contain many ultra-conserved elements (UCEs), long sequences identical between distant species. In this study we identified UCEs in drosophilid and vertebrate species with a similar level of phylogenetic divergence measured at protein-coding regions, and demonstrated that both the length and number of UCEs are larger in vertebrates. The proportion of non-exonic UCEs declines in distant drosophilids whilst an opposite trend was observed in vertebrates. We generated a set of 2,126 Sophophora UCEs by merging elements identified in several drosophila species and compared these to the eutherian UCEs identified in placental mammals. In contrast to vertebrates, the Sophophora UCEs are depleted around transcription start sites. Analysis of 52,954 P-element, piggyBac and Minos insertions in the D. melanogaster genome revealed depletion of the P-element and piggyBac insertions in and around the Sophophora UCEs. We examined eleven fly strains with transposon insertions into the intergenic UCEs and identified associated phenotypes in five strains. Four insertions behave as recessive lethals, and in one case we observed a suppression of the marker gene within the transgene, presumably by silenced chromatin around the integration site. To confirm the lethality is caused by integration of transposons we performed a phenotype rescue experiment for two stocks and demonstrated that the excision of the transposons from the intergenic UCEs restores viability. Sequencing of DNA after the transposon excision in one fly strain with the restored viability revealed a 47 bp insertion at the original transposon integration site suggesting that the nature of the mutation is important for the appearance of the phenotype. Our results suggest that the UCEs in flies and vertebrates have both common and distinct features, and demonstrate that a significant proportion of intergenic drosophila UCEs are sensitive to disruption.
PLoS ONE 12/2013; 8(12):e82362. DOI:10.1371/journal.pone.0082362 · 3.23 Impact Factor
"Proteins coded by the genes from GTF2I family play a role in modification of chromatin structure and hence gene regulation (Wen et al. 2003; Tussie-Luna et al. 2002). Such effects were described for transcription of immunoglobulin heavy-chain gene (Ren et al. 2011), beta-globin gene (Crusselle-Davis et al. 2007), Dlx5/Dlx6 genes in the forebrain (Poitras et al. 2010), troponin I (Tay et al. 2003), some HIV-1 genes (Malcolm et al. 2007, 2008), Rous sarcoma virus genes (Mobley and Sealy 2000), etc. It was found that proteins from this family affect concentration of Ca ions in cytoplasm (Roy 2006; Caraveo et al. 2006). "
[Show abstract][Hide abstract] ABSTRACT: Three genes GTF2IRD1, GTF2I, and GTF2IRD2, which encode members of the GTF2I (or TFII-I) family of so-called general transcription factors, were discovered and studied during the last two decades. Chromosome location and similarity of exon-intron structures suggest that the family evolved by duplications. The initial duplication of ancestral proto-GTF2IRD1 gene likely occurred in early vertebrates prior to origin of cartilaginous fish and led to formation of GTF2I (>450 MYA), which was later lost in bony fish but successfully evolved in the land vertebrates. The second duplication event, which created GTF2IRD2, occurred prior to major radiation events of eutherian mammalian evolution (>100 MYA). During recent steps of primate evolution there was another duplication which led to formation of GTF2IRD2B (<4 MYA). Two latest duplications were coupled with inversions. Genes belonging to the family have several highly conservative repeats which are implicated in DNA binding. Phylogenetic analysis of the repeats revealed a pattern of intragenic duplications, deletions and substitutions which led to diversification of the genes and proteins. Distribution of statistically rare atypical substitutions (p ≤ 0.01) sheds some light on structural differentiation of repeats and hence evolution of the genes. The atypical substitutions are often located on secondary structures joining α-helices and affect 3D arrangement of the protein globule. Such substitutions are commonly traced at the early stages of evolution in Tetrapoda, Amniota, and Mammalia.
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