Novel Mechanism of Attenuation of LPS-Induced NF-kB Activation by the Heat Shock Protein 90 Inhibitor, 17-N-allylamino-17-demethoxygeldanamycin, in Human Lung Microvascular Endothelial Cells

Heat shock protein (hsp) 90 inhibition attenuates NF-... activation and blocks inflammation. However, the precise mechanism of NF-... regulation by hsp90 in the endothelium is not clear. The authors investigated the mechanisms of hsp90 inhibition by 17-N-allylamino-17- demethoxygeldanamycin (17-AAG) on NF-... activation by LPS in primary human lung microvascular endothelial cells. Transcriptional activation of NF-... was measured by luciferase reporter assay, gene expression by real-time RT-PCR, DNA binding of transcription factors by chromatin immunoprecipitation assay, protein-protein interaction by coimmunoprecipitation/immunoblotting, histone deacetylase (HDAC)/histone acetyltransferase enzyme activity by fluorometry, and nucleosome eviction by partial microccocal DNase digestion. The effect of LPS on IKBα mRNA expression was associated with rapid deacetylation of histone-H3(Lys9) and a dramatic down-regulation of core histone H3binding. Even though treatment with an HDAC inhibitor produced the same effect as hsp90 inhibition, the effect of17-AAG was independent of HDAC. The authors conclude that hsp90 inhibition attenuates NF-...B transcriptional activation by preventing coactivator recruitment and nucleosome eviction from the target promoter in human lung endothelial cells.