Project description:ARID1a (BAF250), a component of human SWI/SNF chromatin remodeling complexes, is frequently mutated across numerous cancers, and its loss of function has been putatively linked to glucocorticoid resistance. Here, we interrogate the impact of siRNA knockdown of ARID1a compared to a functional interference approach, both in the HeLa human cervical cancer cell line. We report that ARID1a knockdown resulted in a significant global decrease in chromatin accessibility in ATAC-seq analysis, as well as affecting a subset of genome-wide GR binding sites in GR ChIP-seq analysis. Interestingly, the specific effects on gene expression were limited to a relatively small subset of glucocorticoid-regulated genes, notably those involved in cell cycle regulation and DNA repair. The vast majority of glucocorticoid-regulated genes were largely unaffected by ARID1a knockdown or functional interference, consistent with a more specific role for ARID1a in glucocorticoid function than previously speculated. Using liquid chromatography mass spectrometry, we have identified a chromatin-associated protein complex comprising GR, ARID1a, the cell cycle regulating P53 binding protein P53BP1, DNA repair proteins including Poly(ADP-Ribose) Polymerase 1 (PARP1), DNA damage-binding protein 1 (DDB1), DNA mismatch repair protein MSH6 and splicing factor proline and glutamine rich protein (SFPQ), as well as the histone acetyl transferase KAT7, an epigenetic regulator of steroid-dependent transcription, DNA damage repair and cell cycle regulation. Not only was this protein complex ablated with both ARID1a knockdown and functional interference, but DNA damage repair was also significantly impaired. Recovery from dexamethasone-dependent cell cycle arrest was also significantly impaired. Taken together, our data demonstrates that although glucocorticoids can still promote cell cycle arrest in the absence of ARID1a, the purpose of this arrest to allow time for DNA damage repair is hindered.

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Effects of ARID1A knockdown on gene expression in human cholangiocyte cell line MMNK1

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