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Shawn dissertation NRSF
1. Dissertation Defense Shawn McClelland NRSF mediated epigenetic processes in neuroplasticity Committee Members: Dr. Tallie Z. Baram Dr. Martin Smith Dr. Marcelo Wood
2. What makes us who we are? Genes Changes induced by the environment
4. Why focus on Epilepsy*? - Third most common chronic brain disorder - The most common chronic brain disorder in children & young adults - >1% of the population - Enormous personal, social & fiscal impact * Generation of spontaneous seizures by the brain
5. Epilepsy or Epilepsies Numerous Mechanisms for the Epilepsies: Genetic Acquired Gene-environment interaction
6. Temporal Lobe Epilepsy (TLE) Involves the hippocampus Most common epilepsy in adults Can be severe and intractable to medicines Not genetic
8. Insult Acute damage Latent period Progressive damage Hyperexcitability Seizures Epilepsy after severe Head Trauma Epilepsy after severe Head Trauma Circuit reorganization But most epilepsy does not follow trauma….
10. How does the brain become epileptic after insults that have little cell death? - No clear structural changes - Change is in the function of the brain cells - Genes that neurons express govern function Which gene or genes are changed to make neurons epileptic? Toth et al., J Neurosci, 1998; Bender et al., 2003; Dube et al, TiNS 2007
11. 0.2 Ci/gm * 0.1 m * 0.0 Control HT-activity KA-activity HCN1 expression is repressed after epilepsy-provoking insults Chen, 2001; Brewster 2002,2005; Santoro & Baram 2003; Dyhrfjeld-Johnsen, 2008
12. Dysregulation of HCN1 is common in many animal models of epilepsy Chen, Baram, Soltesz, Nature Med, 2001 Brewster…Baram J Neurosci. 2002 Ludwig …Biel, EMBO J. 2003 Budde….Pape. J Neurosci. 2005 Kamal… de Graan, Ramakers, Eur J Neurosci, 2006 Kuisle…Baram, Luthi, J Physiol. 2006 Schridde, Strauss… van Luijtelaar 2006 Shin & Chetkovich, JBC, 2007 Jung…Poolos, J Neurosci. 2007, 2010 Zhang…Sanchez, Epilepsia, 2007 Powell… O’Brien, Epilepsia. 2008 Dugladze...Gloveli, PNAS 2008 Van Gaase...deGraan, 2008 Marcelin...Bernard, Neurobiol Dis. 2008 Lewis...Baram, Chetkovich , J Neurosci, 2009 Huang…Shah, J Neurosci, 2009 Santoro…Blumenfeld, Epilepsia 2010. Hablitz & Yang, Epilepsia 2010
13. How is HCN1 expression repressed during epileptogenesis? - The mechanisms might be informative about epileptogenesis - They might provide a ‘HOOK’ into large-scale gene-network changes that constitute the epileptogenic process Chen, 2001; Santoro & Baram 2003, Bender & Baram 2008; McClelland, Ann Neurol 2011
15. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical binding to the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changes of the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing ~30 genes, (in addition to hcn1) whose repression contribute to epilepsy.
16. NRSF Neuron-Restrictive Silencer Factor The original dogma NRSF silences gene expression - Group of genes important to neurons So NRSF is found in all types of cells except for neurons NRSF is found in neurons! Regulates, instead of silences, neuronal genes Basal levels reported as low in the brain, and to increase with seizure activity
25. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical binding to the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changes of the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing ~30 genes, (in addition to hcn1) whose repression contribute to epilepsy.
27. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical binding to the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changes of the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing ~30 genes, (in addition to hcn1) whose repression contribute to epilepsy.
28. An epigenetic mark of repression is selectively and persistently augmented after KA-SE at the hcn1 gene Active State Repressed State
29. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical binding to the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changes of the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing of ~30 genes, in addition to hcn1, that together contribute to epilepsy.
33. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical binding to the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changesof the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing ~30 genes, (in addition to hcn1) whose repression contribute to epilepsy.
34. Which genes did we rescue that modified the disease process? MICROARRAY Check the mRNA levels of 22,000 genes 0 day +1 -7 days +2 Surgery-Cannulae Induce Status Epilepticus Kainic Acid injections Periodically Infuse NRSE-ODNs
35. Which genes did we rescue that modified the disease process? 22,000 genes Genes repressed after insult leading to epilepsy. 470 genes 30 genes Genes rescued Using decoy ODNs
36. Decoy NRSE-ODNs alter the outcome of KA-induced status epilepticus. ALL of these genes have putative NRSEs
37. NRSF mediated epigenetic processes in neuroplasticity NRSF regulates hcn1 after an insult that leads to epilepsy through its physical bindingto the gene. Enduring repression of hcn1 expression by NRSF involves epigenetic changesof the chromatin of the gene. Blocking NRSF function after an insult that provokes epilepsy markedly attenuates the development of epilepsy. The prevention of epilepsy is a result of rescuing ~30 genes, (in addition to hcn1) whose repression contribute to epilepsy.
38. Temporal Lobe Epilepsy (TLE) Involves the hippocampus Most common epilepsy in adults Can be severe and intractable to medicines Not genetic
39. Understanding the mechanisms involved in this type of aberrant neuroplasticity may help in other neurological disorders. Neuroplasticity makes us who we are!
45. NRSF – Histone Modification NRSF is found in neurons! Direct repression, Histone modification, DNA methylation,… Activity influences NRSF levels and NRSF represses a set of genes.
46. ALLgenes validated by GSEA based on the Johnson paper list of genes containing an NRSE. +indicates genes validated by GSEA based on MSigDB:V$NRSF_01 list of genes %indicates genes (14) found with ANOVA, Benjamini-Hoch FDR < 0.25. $indicates genes (6) found with one-way ANOVA, unadjusted p value < 0.05 Johnson DS, Mortazavi A, Myers RM, Wold B (2007) Genome-wide mapping of in vivo protein-DNA interactions. Science 316: 1497-1502.
47. Heatmap of ALL NRSE-containing genes down regulated after KA-SE The majority of genes containing NRSE sites are recovered by the administration of NRSE-ODNs
48. Candidate Gene Analysis Genes repressed in KA+SCRLD compared to Sham+SCRLD Rescued Genes: increased in KA-NRSE compared to KA-SCRLD 4 GROUPS Sham + SCRLD Sham + NRSE KA + SCRLD KA + NRSE Genes repressed in KA+NRSE compared to Sham+SCRLD Also performed ANOVA on the groups.
