Distinct chromatin landscapes dictate resident cardiac macrophage lifecycles and functions

AIMS: Three resident macrophage subsets co-exist in the homeostatic heart, exhibiting distinct origins transcriptional identities and functions. However, the underlying epigenetic architecture regulating resident cardiac macrophage subset heterogeneity is not known, which limits mechanistic understanding and therapeutic targeting. METHODS AND RESULTS: We performed bulk and single-cell ATAC-seq to map the chromatin landscape of murine resident macrophage subsets. Integration of ATAC-seq with RNA-seq of each cardiac macrophage subset revealed two primary chromatin accessibility-based cell states that correlated with tissue residency - a CCR2- macrophage chromatin-based cell state that corresponded with resident TLF+ (TIMD4 + LYVE1 + FOLR2+) and CCR2-MHC-IIhi subsets that do not require monocytes for maintenance, and a CCR2+ chromatin-based cell state that corresponded to CCR2 + MHC-IIhi macrophages, which rely on monocyte input. Using footprint analysis, we identified subset-specific differences in transcription factor binding that explained transcriptional differences, including enriched AP-1 binding in CCR2+ macrophages and enriched MAF-family factor binding in TLF+ macrophages. Importantly, we demonstrate myeloid-specific loss of Mafb or Maf skewed macrophage subset composition toward CCR2+ macrophages and increased MHC-II expression in TLF+ macrophages. Furthermore, we identify a link between IRF8 and MHC-II, in which myeloid-specific loss of Irf8 abolished MHC-II expression exclusively in fate-mapped resident CCR2- subsets but not in CCR2+ macrophages, demonstrating a subset-specific dependency of IRF8 in modulating antigen-presentation. CONCLUSION: Together, these data suggest tissue residency dictates chromatin accessibility and transcription factor binding in cardiac macrophages, linking epigenetic structure to monocyte-dependency, distinct gene expression patterns and critical functions such as antigen presentation.