Abstract
NODAL/Activin signaling orchestrates key processes during embryonic development via SMAD2. How SMAD2 activates programs of gene expression that are modulated over time however, is not known. Here we delineate the sequence of events that occur from SMAD2 binding to transcriptional activation, and the mechanisms underlying them. NODAL/Activin signaling induces dramatic chromatin landscape changes, and a dynamic transcriptional network regulated by SMAD2, acting via multiple mechanisms. Crucially we have discovered two modes of SMAD2 binding. SMAD2 can bind pre-acetylated nucleosome-depleted sites. However, it also binds to unacetylated, closed chromatin, independently of pioneer factors, where it induces nucleosome displacement and histone acetylation. For a subset of genes, this requires SMARCA4. We find that long term modulation of the transcriptional responses requires continued NODAL/Activin signaling. Thus SMAD2 binding does not linearly equate with transcriptional kinetics, and our data suggest that SMAD2 recruits multiple co-factors during sustained signaling to shape the downstream transcriptional program.