CBP (CREB-binding protein) and p300 are versatile transcriptional coactivators that link transcriptional activators to the basal transcriptional apparatus. This study identifies CBP and p300 as coactivators of the nuclear factor-κB (NF-κB) component p65 (RelA). Both CBP and p300 potentiate p65-activated transcription of E-selectin and VCAM-1–CAT reporter constructs. The N- and C-terminal domains of CBP/p300 functionally interact with a region of p65 containing the transcriptional activation domain, as demonstrated by mammalian two-hybrid assays and glutathione S-transferase fusion protein binding. Coimmunoprecipitation/Western blot studies also show direct physical interactions between CBP/p300 and p65. The adenovirus E1A 12S protein, which complexes with CBP and p300, inhibits p65-dependent gene expression, which can be rescued by overexpression of CBP or p300. These findings suggest that CBP and p300 act as coactivators of p65-driven gene activation and may play a crucial role in the cytokine-induced expression of various immune and inflammatory genes.CBP (CREB-binding protein) and p300 are versatile transcriptional coactivators that link transcriptional activators to the basal transcriptional apparatus. This study identifies CBP and p300 as coactivators of the nuclear factor-κB (NF-κB) component p65 (RelA). Both CBP and p300 potentiate p65-activated transcription of E-selectin and VCAM-1–CAT reporter constructs. The N- and C-terminal domains of CBP/p300 functionally interact with a region of p65 containing the transcriptional activation domain, as demonstrated by mammalian two-hybrid assays and glutathione S-transferase fusion protein binding. Coimmunoprecipitation/Western blot studies also show direct physical interactions between CBP/p300 and p65. The adenovirus E1A 12S protein, which complexes with CBP and p300, inhibits p65-dependent gene expression, which can be rescued by overexpression of CBP or p300. These findings suggest that CBP and p300 act as coactivators of p65-driven gene activation and may play a crucial role in the cytokine-induced expression of various immune and inflammatory genes.