Fig 5.

SubAB suppresses LPS-induced NF-κB p65/p50 heterodimer binding to the iNOS promoter. RAW 264.7 cells (2 × 10⁶ cells/dish) were treated with or without LPS (10 μg/ml) in the presence or absence of SubAB (WT; 0.5 μg/ml) or mSubAB (MT; 0.5 μg/ml) for 4 h. Cells were fixed with formaldehyde, lysed in SDS buffer, and then sonicated to fragment the chromatin. Immunoprecipitation was performed using anti-p65 (p65 IP) or anti-p50 (p50 IP) antibodies or normal rabbit IgG. Normal IgG IP refers to immunoprecipitation of chromatin with a nonimmune IgG. After purification of immunoprecipitated DNA, targeted promoter regions of iNOS were amplified by qPCR. Results are expressed as the percent input for each ChIP fraction, and fold changes were calculated to normal IgG IP. Data are representative of three independent experiments.