Markus Meyerhöfer, Yawen Zhou, Aaron Gallego-Crespo, Viral Shah, Malte Andreas Behrendt, Maria Saura-Pañella, Björn Häupl, Oleksandr Todoriuk, Monika Hartmann, Matthias Klein, Catherine Wölfel, Patricia S. Hähnel, Christian Michel, Sabine Muth, Thomas Kindler, Tobias Bopp, Hansjörg Schild, Sarah J. Horton, Markus P. Radsak, Matthias Theobald, George S. Vassiliou, Brian J.P. Huntly, Michael W.M. Kühn, Falk Butter, Thomas Oellerich, Daniel Sasca
ABSTRACT: The lysine acetyltransferase (KAT) activity of EP300 lysine acetyltransferase (p300)/CREB-binding protein (CREBBP) has traditionally been linked to transcriptional activation. This has been attributed largely to acetylation of histone residues such as histone H3 lysine 27 acetylation (H3K27ac), a defining hallmark of active regulatory elements. Here we show that, in acute myeloid leukemia (AML), inhibition of p300/CREBBP catalysis can paradoxically increase transcription. We combined time-resolved dynamics of nascent and total transcription with chromatin binding dynamics of p300/CREBBP and their associated transcription factors (TFs)/coregulators (inferred from chromatin pulldown proteomics, acetyl proteomics, and motif enrichment) to uncover mechanisms of transcriptional rewiring after p300/CREBBP catalytic inhibition. In parallel, we dissected the functional contribution of individual p300/CREBBP acetyl-interactome members to KAT inhibition using genome-wide CRISPR-Cas9 dropout and focused Perturb-seq screens. Together, these approaches revealed that KAT inhibition paradoxically retains p300/CREBBP, and promotes cooperative TF assembly and increased H3K27 acetylation at a subset of regulatory elements. The effect was most pronounced at interferon regulatory factor (IRF) motif-enriched loci, including interferon-stimulated genes (ISGs), where KAT inhibition triggered p300/CREBBP accumulation and enhanced combinatorial TF binding, enabling recruitment of the ISG activator STAT1. Consequently, ISG loci were converted into transcriptionally active states that induced cell-cycle arrest, differentiation, and apoptosis. Therapeutically, combining KAT inhibition with interferon-alpha augmented ISG expression, synergistically drove AML cell death in vitro, and significantly extended survival in both AML xenografts and murine models. These findings refine our understanding of p300/CREBBP KAT activity, demonstrating that cooperative TF assembly can reconfigure p300/CREBBP-containing complexes under catalytic inhibition to induce transcription, with translational implications for reprogramming interferon-driven programs through catalytic inhibition in AML and beyond.