The histone demethylase KDM5C gene is a direct target of the ARX homeobox transcription factor.

Intellectual Disability (ID) and Epilepsy often occur together with a dramatic impact on the development and quality of life of the affected children. PolyAlanine expansion mutations of Aristaless-related homeobox gene (ARX) cause a spectrum of X-chromosome phenotypes with ID (XLID) and various forms of malignant paediatric epilepsy, including infantile spasms. We show that lysine (K)-specific demethylase 5C (KDM5C/JARID1C/SMCX), a known XLID gene involved in chromatin remodeling, is regulated by ARX. Combining luciferase reporter and DNA binding assays, we demonstrate that ARX directly regulates and binds a Conserved Noncoding Element in the 5' region of the KDM5C gene. We studied ARX mutant proteins carrying various PolyAlanine expansion mutations, identified in XLID/Epilepsy patients. We prove that the changes in the repeat units in PolyAlanine tracts cause hypomorphic ARX mutations, which exhibit a decreased trans-activating activity and a reduced, but not abolished, binding to the KDM5C regulatory region. Furthermore, the altered functioning of the five ARX PolyAlanine mutants tested is likely to correlate with the degree of XLID/Epilepsy severity. By means of quantitative RT-PCR, we show a dramatic downregulation of the Kdm5C mRNA levels in murine Arx-KO ES cells and neurospheres. Such diminution in Kdm5C mRNA level leads to a decrease of the Kdm5C protein content during in vitro neuronal differentiation. Furthermore, we observed that Arx KO produced an increase of the Kdm5C-mediated levels of H3K4me3, potentially due to a compromised activity of Kdm5C. Since H3K4me3 is the hallmark of open chromatin, Arx-dependent Kdm5C defects could compromise cyclical rounds of methylation-demethylation and consequently the chromatin remodelling. In summary, we discover a new ARX-dependent disease path and suggest that the molecular pathogenesis of ARX PolyAlanine mutations may be in part caused by aberrant histone demethylation as a result of KDM5C defect. As chromatin modifications are reversible, it is possible that epigenetic drugs could compensate KDM5C-H3K4me3 deregulation. Even though many other ARX targets could have important roles in the expansion phenotype, our findings could potentially open up further studies aimed to cure or ameliorate ARX PolyAlanine-related phenotypes, and other cognition and epileptic disorders.