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The transcription factor NRSF contributes to epileptogenesis by selective repression of a subset of target genes.

McClelland S, Brennan GP, Dubé C, Rajpara S, Iyer S, Richichi C, Bernard C, Baram TZ - Elife (2014)

Bottom Line: Accordingly, the repressed gene-set was rescued when NRSF binding to chromatin was blocked.Unexpectedly, genes selectively repressed by NRSF had mid-range binding frequencies to the repressor, a property that rendered them sensitive to moderate fluctuations of NRSF levels.Genes selectively regulated by NRSF during epileptogenesis coded for ion channels, receptors, and other crucial contributors to neuronal function.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Neurobiology, University of California, Irvine, Irvine, United States Department of Pediatrics, University of California, Irvine, Irvine, United States Department of Neurology, University of California, Irvine, Irvine, United States.

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Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.(A) A schematic illustrating the mechanism of action of NRSF following a seizure-induced increase and the decoy oligodeoxynucleotide (ODN) intervention strategy with the expected outcome. (B) A heat map representation of the changes in mRNA expression levels of genes that contain a putative NRSE site that were down-regulated 48 hr after KA-induced seizures. Heat map compares representative samples from two hippocampi, from each of four experimental conditions: 'controls' receiving random ODNs (n = 4); ‘controls’ receiving NRSE ODNs (n = 4); ‘KA-seizures’, rats sustaining KA induced seizure activity and receiving random ODNs (n = 3); ‘KA-seizures + NRSE-ODN’, rats sustaining KA-induced seizure activity and receiving NRSE-ODNs (n = 4). Samples and genes are plotted using hierarchical clustering using Euclidean distance and average linkage. Expression level is depicted by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (C) Independent analysis of gene expression using qPCR. Several genes that were both repressed by seizure activity and rescued by interference with NRSF function were tested (Glra2, Myo5B, Stmn2), and results analyzed using two way ANOVA. Myo5B F(1,18) = 9.35, p = 0.007; Glra2 F(1,17) = 46.89, p = 0.0001; Stmn2 F(1,18) = 1.97, p = 0.047, n = 4/group.DOI:http://dx.doi.org/10.7554/eLife.01267.005
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fig3: Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.(A) A schematic illustrating the mechanism of action of NRSF following a seizure-induced increase and the decoy oligodeoxynucleotide (ODN) intervention strategy with the expected outcome. (B) A heat map representation of the changes in mRNA expression levels of genes that contain a putative NRSE site that were down-regulated 48 hr after KA-induced seizures. Heat map compares representative samples from two hippocampi, from each of four experimental conditions: 'controls' receiving random ODNs (n = 4); ‘controls’ receiving NRSE ODNs (n = 4); ‘KA-seizures’, rats sustaining KA induced seizure activity and receiving random ODNs (n = 3); ‘KA-seizures + NRSE-ODN’, rats sustaining KA-induced seizure activity and receiving NRSE-ODNs (n = 4). Samples and genes are plotted using hierarchical clustering using Euclidean distance and average linkage. Expression level is depicted by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (C) Independent analysis of gene expression using qPCR. Several genes that were both repressed by seizure activity and rescued by interference with NRSF function were tested (Glra2, Myo5B, Stmn2), and results analyzed using two way ANOVA. Myo5B F(1,18) = 9.35, p = 0.007; Glra2 F(1,17) = 46.89, p = 0.0001; Stmn2 F(1,18) = 1.97, p = 0.047, n = 4/group.DOI:http://dx.doi.org/10.7554/eLife.01267.005

Mentions: Whereas 371 of the detected hippocampal genes contained an NRSE and were therefore potential NRSF targets, only 39 of these were repressed by KA-induced seizures when NRSF levels increased twofold to threefold. To examine if the correlation between NRSF levels and the repression of this subset (∼10%) of NRSE-containing genes was causal, we employed a decoy oligonucleotide strategy to block NRSF binding to the NRSE sequences of genomic DNA (Akhtar et al., 1991; Szklarczyk and Kaczmarek, 1995; Gilar et al., 1998; Soldati et al., 2011; Sedaghat et al., 2013). We generated oligodeoxynucleotides (ODNs) comprised of the NRSF binding sequence (NRSE), modified their backbone for stability and infused them into the brain. These ODNs acted as ‘decoys’, binding to cellular NRSF and inhibiting its ability to bind to target genes (Figure 3A) (McClelland et al., 2011a).10.7554/eLife.01267.005Figure 3.Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.


The transcription factor NRSF contributes to epileptogenesis by selective repression of a subset of target genes.

McClelland S, Brennan GP, Dubé C, Rajpara S, Iyer S, Richichi C, Bernard C, Baram TZ - Elife (2014)

Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.(A) A schematic illustrating the mechanism of action of NRSF following a seizure-induced increase and the decoy oligodeoxynucleotide (ODN) intervention strategy with the expected outcome. (B) A heat map representation of the changes in mRNA expression levels of genes that contain a putative NRSE site that were down-regulated 48 hr after KA-induced seizures. Heat map compares representative samples from two hippocampi, from each of four experimental conditions: 'controls' receiving random ODNs (n = 4); ‘controls’ receiving NRSE ODNs (n = 4); ‘KA-seizures’, rats sustaining KA induced seizure activity and receiving random ODNs (n = 3); ‘KA-seizures + NRSE-ODN’, rats sustaining KA-induced seizure activity and receiving NRSE-ODNs (n = 4). Samples and genes are plotted using hierarchical clustering using Euclidean distance and average linkage. Expression level is depicted by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (C) Independent analysis of gene expression using qPCR. Several genes that were both repressed by seizure activity and rescued by interference with NRSF function were tested (Glra2, Myo5B, Stmn2), and results analyzed using two way ANOVA. Myo5B F(1,18) = 9.35, p = 0.007; Glra2 F(1,17) = 46.89, p = 0.0001; Stmn2 F(1,18) = 1.97, p = 0.047, n = 4/group.DOI:http://dx.doi.org/10.7554/eLife.01267.005
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fig3: Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.(A) A schematic illustrating the mechanism of action of NRSF following a seizure-induced increase and the decoy oligodeoxynucleotide (ODN) intervention strategy with the expected outcome. (B) A heat map representation of the changes in mRNA expression levels of genes that contain a putative NRSE site that were down-regulated 48 hr after KA-induced seizures. Heat map compares representative samples from two hippocampi, from each of four experimental conditions: 'controls' receiving random ODNs (n = 4); ‘controls’ receiving NRSE ODNs (n = 4); ‘KA-seizures’, rats sustaining KA induced seizure activity and receiving random ODNs (n = 3); ‘KA-seizures + NRSE-ODN’, rats sustaining KA-induced seizure activity and receiving NRSE-ODNs (n = 4). Samples and genes are plotted using hierarchical clustering using Euclidean distance and average linkage. Expression level is depicted by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (C) Independent analysis of gene expression using qPCR. Several genes that were both repressed by seizure activity and rescued by interference with NRSF function were tested (Glra2, Myo5B, Stmn2), and results analyzed using two way ANOVA. Myo5B F(1,18) = 9.35, p = 0.007; Glra2 F(1,17) = 46.89, p = 0.0001; Stmn2 F(1,18) = 1.97, p = 0.047, n = 4/group.DOI:http://dx.doi.org/10.7554/eLife.01267.005
Mentions: Whereas 371 of the detected hippocampal genes contained an NRSE and were therefore potential NRSF targets, only 39 of these were repressed by KA-induced seizures when NRSF levels increased twofold to threefold. To examine if the correlation between NRSF levels and the repression of this subset (∼10%) of NRSE-containing genes was causal, we employed a decoy oligonucleotide strategy to block NRSF binding to the NRSE sequences of genomic DNA (Akhtar et al., 1991; Szklarczyk and Kaczmarek, 1995; Gilar et al., 1998; Soldati et al., 2011; Sedaghat et al., 2013). We generated oligodeoxynucleotides (ODNs) comprised of the NRSF binding sequence (NRSE), modified their backbone for stability and infused them into the brain. These ODNs acted as ‘decoys’, binding to cellular NRSF and inhibiting its ability to bind to target genes (Figure 3A) (McClelland et al., 2011a).10.7554/eLife.01267.005Figure 3.Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.

Bottom Line: Accordingly, the repressed gene-set was rescued when NRSF binding to chromatin was blocked.Unexpectedly, genes selectively repressed by NRSF had mid-range binding frequencies to the repressor, a property that rendered them sensitive to moderate fluctuations of NRSF levels.Genes selectively regulated by NRSF during epileptogenesis coded for ion channels, receptors, and other crucial contributors to neuronal function.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Neurobiology, University of California, Irvine, Irvine, United States Department of Pediatrics, University of California, Irvine, Irvine, United States Department of Neurology, University of California, Irvine, Irvine, United States.

Show MeSH
Related in: MedlinePlus