Coordinated transcription factor and chromatin dynamics enable innate immune memory in single macrophages

Cells of the innate immune system need to interpret dynamic microenvironments and rapidly respond to sequential signals to control inflammation and infection. It is unclear how complex immune signals are encoded and decoded through a small number of transcription factors and ultimately coordinate cellular decision-making. Here, we use single live cell analysis and ATAC sequencing to show that sequential inflammatory signals via the NF?B network induce memory through reprogramming of signaling network state and chromatin accessibility in primary macrophages. We use transcriptomic profiling and deep learning to show that these regulatory modes combine to explain macrophage decision-making in response to new inflammatory signals. Macrophage response to prior stimulus predicts memory state and transcriptional response to subsequent stimulus. Surprisingly, we find that innate immune memory is mediated via intracellular individual-cell factors and not via paracrine signaling between macrophages. This work comprehensively profiles the dynamic regulation of signaling networks and chromatin accessibility and their role in tuning macrophage responses to evolving inflammatory signals. Overall design: Murine BMDMs were treated with media along (M), CpG (C), PolyI:C (P) for 4 hours at which point the media was replaced with fresh media containing media alone (M), LPS (L), PolyI:C (P), CpG (C), or TNF (T). Treated BMDM were lifted from plates and ATACseq was performed as described in the methods. High throughput sequencing was performed with 50bp bi-directional reads.