Final Report Summary - EPITARGENE (SRF target genes in epilepsy.) Executive Summary:Epilepsy is a chronic neurological disorder, affecting 1-3% of human population, with temporal lobe epilepsy (TLE) being the most common type in adults. Unfortunately, current anti-epileptic drugs are ineffective in more than 30% of TLE patients. Therefore, it is very important to understand the molecular mechanism underlying this pathology. Multiple pieces of evidence imply that aberrant synaptic plasticity may underlie epilepsy. Inactivation of SRF (Serum Response Factor), one of the major transcription regulators, was shown to cause deficiency in hippocampal synaptic plasticity and learning. In addition, it’s involvement in impaired axonal outgrowth, guidance and synaptic targeting has been reported.Project EpiTarGene was based on the hypothesis that regulation of gene transcription by SRF can explain mechanisms of aberrant plasticity observed in epilepsy. The aim was to identify genes regulated by SRF in epilepsy. In the course of the experiments we obtained an inducible, forebrain specific SRF knockout line of mice (SRF KOs) that lack SRF protein in the brain during adulthood. Next, we employed microarrays to monitor global gene expression in the kainic acid (KA) model of aberrant plasticity.Statistical analysis of microarray results revealed that in basic conditions (saline treated animals) SRF KOs do not display any significantly downregulated transcripts, except the SRF itself (Tukey p < 0.05 fold change < 0.66). As an outcome of our experiments, 431 genes altered after seizures and significantly changed in SRF KOs were identified (ANOVA interaction genotype: treatment p < 0.0005 correction FDR <1%, fold induction after KA > 1.5 or < 0.66). More than 260 genes showed increased expression after seizures in WT animals and were significantly down-regulated in SRF KOs. Among those genes we discovered and experimentally verified potential plasticity related genes, including: lipocalin 2 (Lcn2, NGAL), that is a small, secreted protein originally identified as a protein associated with plasticity related protease MMP-9.In our experiments we aimed at the evaluation of the role of Lcn2 in structural synaptic plasticity and showed that an increased level of Lcn2 may exert rapid effects on the dendritic spines morphology. Lcn2 caused elongation, thinning and a decrease in the proportion of mature spines, and could possibly lead to the lowering of the hyperexcitability of the network.We believe that our data will have an impact on current understanding of the pathogenesis of epilepsy and influence the development of new therapeutic opportunities through selective targeting of aberrant plasticity-related SRF effectors.
