Synaptic Activity Causes Minute-scale Changes in BAF Complex Composition and Function [ATAC-Seq]

Genes encoding subunits of the BAF (mammalian SWI/SNF) ATP-dependent chromatin remodeling complex are among the those most enriched for deleterious de novo mutations in intellectual disabilities and autism spectrum disorder, but the causative molecular pathways are not understood. Synaptic activity in neurons is critical for learning and memory and proper neural development. While BAF is required for activity-dependent developmental processes, such as dendritic outgrowth, the immediate molecular consequences of neuronal activity on BAF complexes are unknown. Here we report that neuronal activity induces dramatic remodeling of the subunit composition of BAF complexes within 15 minutes, concurrent with both phosphorylation and dephosphorylation of its subunits. These biochemical effects are a convergent phenomenon downstream of multiple calcium-activated signaling pathways in neurons and fibroblasts and correspond to changes in BAF-dependent chromatin accessibility. Our studies imply that BAF decodes signals at the membrane by altering the combinatorial composition of its subunits. Overall design: ATAC-seq: to investigate the targets of the BAF complex in neurons, primary cortical neurons from E16.5 embryonic mice were cultured for 7 days in vitro and depolarized with 50 mM KCl for 10 minutes or 30 minutes to mimic neuronal activity, and/or treated with 0.1% DMSO or 100 nM FHT-1015 (an inhibitor of BAF ATPase activity). To investigate the effect of the PBAF complex, neurons were infected with Arid2KO (KO) or non-targeting (NT) CRISPR gRNAs, cultured for 7 days in vitro, and depolarized (30 mins) or left unstimulated. Also, to investigate the effects of Smarcc2 serine phosphorylations, gene knockout, vector (Vec), Smarcc2 wildtype (WT), Smarcc2 S586E, and Smarcc2 S586A were introduced into cortical neurons with non-targeting (NT) or Smarcc2KO (KO) CRISPR gRNAs, cultured for 18 days, and depolarized (30 mins) or left unstimulated. Unstimulated conditions in all cases included 1 uM TTX/100 uM D-AP5 overnight treatment to quiet spurious background activity.