The molecular basis of signal-dependent transcriptional activation has been
extensively studied in macrophage polarization, however our understanding remains limited regarding the molecular determinants of repression. Here we show that IL-4- activated STAT6 transcription factor is required for the direct transcriptional repression of a large number of genes during in vitro and in vivo alternative macrophage polarization. Repression results in decreased lineage-determining transcription factor, p300 and RNA polymerase II binding followed by reduced enhancer RNA expression, H3K27 acetylation and chromatin accessibility. The repressor function of STAT6 is HDAC3-dependent on a subset of IL-4-repressed genes. In addition, STAT6-repressed enhancers show extensive overlap with the NF-κB p65 cistrome and exhibit decreased esponsiveness to lipopolysaccharide after IL-4 stimulus on a subset of genes. As a consequence, macrophages exhibit diminished inflammasome activation, decreased IL-1β production and pyroptosis. Thus, IL-4-STAT6 signaling pathway establishes an alternative polarization-specific epigenenomic signature resulting in dampened macrophage responsiveness to inflammatory stimuli.