Endotoxin Tolerance Disrupts Chromatin Remodeling and NF-κB Transactivation at the IL-1β Promoter

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Abstract
The NF-κB family plays a crucial role in the pathogenesis of highly lethal septicemia by modulating transcription of many innate and adaptive immunity genes. Two phases of NF-κB activation occur: cytosolic activation and nuclear transactivation. Septicemia with multiorgan failure is associated with chronic activation of cytosolic NF-κB with translocation and accumulation of increased levels of nuclear p65 in blood leukocytes. Paradoxically, NF-κB-dependent transcription of many proinflammatory genes responding to bacterial LPS endotoxin (LPS) is persistently repressed during septicemia; this phenomenon of LPS tolerance is associated with immunosuppression and poor prognosis. This report suggests an explanation for this paradox. Using an in vitro human leukocyte model and chromatin immunoprecipitation assays, we find that both the cytosolic activation and nuclear transactivation phases of NF-κB occur in LPS responsive THP-1 promonocytes with recruitment and binding of NF-κB p65 at the IL-1β promoter. However, transcriptionally repressed LPS-tolerant THP-1 cells do not bind NF-κB p65 at the IL-1β promoter, despite cytosolic activation and accumulation of p65 in the nucleus. In contrast, NF-κB p50, which also accumulates in the nucleus, constitutively binds to the IL-1β promoter NF-κB site in both LPS-responsive and LPS-tolerant cells. The level of p65 binding correlates with a binary shift in nucleosome remodeling between histone H3 phosphorylation at serine 10 and methylation of histone H3 at lysine 9. We conclude that LPS tolerance disrupts the transactivating stage of NF-κB p65 and altered nucleosome remodeling at the IL-1β promoter in human leukocytes.