Distinct hypoxic molecular mechanisms differentially regulate human vs.mouse Interleukin 1β gene expression in monocytes/macrophages

Abstract

Abstract Interleukin 1β (IL-1β) is an important cytokine, whose regulation is distinct from that of Tumor Necrosis Factor (TNF). We previously reported that LPS/TLR4 initiates hypoxia in mice, with HIF-1α binding the murine Il1b gene, suggesting that TLR4 directly activates Trained Innate Immune Memory (TIIM). To address whether human monocyte IL1B gene regulation parallels that of mouse, mRNA and chromatin immunoprecipitation (ChIP) kinetics, with small molecule inhibitors under either normoxia or hypoxia, were evaluated. IL1B transcription kinetics in Lipopolysaccharide (LPS)-treated human THP-1 monocytes revealed an Immediate-Early (IE) phase of IL1B gene induction, requiring Spi1/PU.1, NF-κB and C/EBPβ transcription factors (TF), followed by a late-phase of lower-level continuous non-tolerized TIIM expression that depends upon hypoxia-inducible TFs. We observed that Spi1/PU.1 is required for IL1B gene transcription, performing a critical role in inducing the promoter for IE expression via a TAF1-independent mechanism. Late HIF-2α-dependent IL1B expression is due to LPS pseudo-hypoxic stabilization of the labile HIF factor. In contrast to IE activation, late-phase transcription revealed minimal dependence on NF-κB due to human-specific competition for an overlapping NF-κB/HIF enhancer element. Our studies reveal that IL1B in naïve THP-1 human monocytes binds HIF-2α, not HIF-1α, while PMA treatment morphologically polarizes THP-1 macrophages to bind both HIF isoforms at IL1B. Additionally, bioinformatic screening revealed other immunologically important genes regulated by similar mechanisms, providing insight for novel therapeutic targeting of transcription factors other than NF-κB. Interleukin Foundation, Denver Colorado