Regulation of nuc-2 remodeling in DCs. (A) DCs were either incubated with medium alone or stimulated with LPS (1 μg/mL) or IFN-γ (100 U/mL) or both for 3 hours. Nuclei were then isolated and digested with BstXI. After purification of genomic DNA and in vitro digestion with EcoRI, samples were analyzed by Southern blotting and indirect end-labeling (left panel). IL-12(p35) mRNA levels were determined by real-time RT-PCR (right panel). For this purpose, DCs were incubated with medium alone or stimulated with LPS (1 μg/mL) or IFN-γ (100 U/mL) or both for 6 hours. IL-12(p35) mRNA levels were normalized using β-actin mRNA levels and were compared with unstimulated conditions. (B) DCs were either incubated with medium alone or stimulated with LPS (1 μg/mL) and IFN-γ (100 U/mL) for the indicated periods of time. DNA accessibility in chromatin within the p35 locus (left panel) and IL-12(p35) mRNA levels (right panel) were assessed as described in panel A. The results are representative of at least 3 independent experiments performed with different donors.