In the clinic, glucocorticoids are widely used due to their anti-inflammatory properties. Despite their huge benefits, prolonged use is often associated with severe side effects as well as the development of glucocorticoid resistance in some cases. Mutations in an important SWI/SNF subunit, ARID1a, have previously been linked to glucocorticoid resistance and are associated with a vast number of human cancers. Determining the functional role of ARID1a in glucocorticoid receptor (GR) signalling is therefore of great importance. Chromatin-remodelling by the SWI/SNF complex is crucial for genomic GR signalling, with chromatin accessibility being dynamically altered at GR binding sites in target genes to regulate transcription. We therefore initially hypothesized ARID1a to be a key regulator of GR mediated transcriptional regulation. This hypothesis was tested using RNA sequencing (RNA-SEQ) to determine the genome-wide effects of loss of ARID1a on glucocorticoid target gene expression in HeLa cells. We found that ARID1a siRNA knockdown had no impact upon RNA expression of the majority of robustly regulated glucocorticoid responsive genes. Instead, the data demonstrated the importance of glucocorticoid regulation on cell-cycle progression through ARID1a, revealing disruption of P53 pathways and a potential role in DNA repair mechanisms. As both GR and P53 directly interact with ARID1a, we tested the impact of loss of ARID1a upon protein interactions. Liquid chromatography mass spectrometry was used to identify GR interacting proteins. These proteomics studies have enabled us to assess how ARID1a knockdown alters components of the protein complex bound to GR at the DNA template. The proteomics data corroborates our RNA-SEQ data; again revealing a loss of GR association with p53 related and DNA repair proteins, thus supporting a novel role of GR in control of cell-cycle through interactions with ARID1a.
06 Nov 2017 - 08 Nov 2017