c-Myb and Pax-5 cooperatively activate the

RAG-2 promoter in 293T cells. (A)RAG-2 promoter activity in 293T cells with expression vector for Pax-5, c-Myb, or both. A quantity of 5 μg luciferase construct linked to the −86/+147 promoter region was transfected into 293T cells in the presence of 0.04 μg, 0.2 μg, 1 μg, or 5 μg of pEFBOS–Pax-5 alone or with 0.2 μg of pAct–c-Myb (indicated as +), or 0.008 μg, 0.04 μg, 0.2 μg, or 1 μg of pAct–c-Myb alone or together with 1 μg of pEFBOS–Pax-5 (indicated as +). pSRα-LacZ was included as an internal control. Total amount of DNA was adjusted with either pAct vector or pEF-BOS vector. Luciferase and β-galactosidase activities were assayed 48 hours later. The luciferase activity without Pax-5 and c-Myb was set to 1. Error bars indicate deviation of 3 experiments. (B) Promoter activity of wild-type versus c-Myb binding site–mutated RAG-2 promoter in the presence of Pax-5, c-Myb, or both. Luciferase construct linked to −86/+147 promoter region containing wild-type c-Myb binding site (WT) or its mutation (c-Myb1m or c-Myb2m) was transfected into 293T cells in the absence or presence of Pax-5, c-Myb, or both Pax-5 and c-Myb expression vectors, and the promoter activities were assessed as above. Error bars indicate deviation of 3 experiments.