CpG content determines tissue-resident macrophage enhancer selection

Tissue-resident macrophages (TRMs) acquire specialized identities through transcription factor (TF) networks operating in distinct chromatin environments. Early growth response 2 (EGR2) contributes both to alveolar macrophage (AM) identity and to interleukin (IL)-4-driven polarization of bone marrow-derived macrophages (BMDMs), yet the regulatory principles enabling its function across these contexts remain unclear. By integrating RNA-seq and ATAC-seq data from wild-type and Egr2-deficient macrophages, we show that macrophage identity is constrained by DNA methylation-dependent cis-regulatory landscapes. AM-specific enhancers are predominantly CpG-rich and hypomethylated, a feature shared with additional TRM populations, whereas BMDM-accessible regulatory regions are largely CpG-poor. Although IL-4 induces EGR2 together with KLF4 and DEC1 in BMDMs, this TF module fails to engage CpG-rich enhancers, indicating that the BMDM regulatory network lacks additional AM-specific components required for their activation. These findings identify CpG content as a determinant of enhancer competence and provide a mechanistic explanation for the context-dependent activity of EGR2 in macrophages, placing macrophage plasticity under the control of sequence-encoded epigenetic constraints.