Physical interaction of GABP and p300 in myeloid cells and effect of p300 on transcriptional coactivation of the CD18 promoter.
Transcriptional coactivator p300 physically contacts GABP and increases responsiveness of the CD18 promoter to RA. (A) A 0.75-mg extract from U937 cells treated with 10−5 M RA for 48 hours was incubated with antibodies to p300 (lane 2), GABPα (lane 3), or no antibody (lane 4), followed by precipitation with protein-G sepharose. Immunoprecipitated products and 50 μg whole-cell extract (lane 1) were immunoblotted with antibodies to p300 and GABPα, as indicated. Open arrow indicates p300, and filled arrow indicates GABPα. Lane 5 represents a brief exposure of lane 2. WCE indicates whole cell extract; IP, immunoprecipitation; WB, Western blot. (B) U937 cells that were stably transfected with CD18(−918)/luc were transiently transfected with the indicated quantities of the transcriptional coactivator p300. Transfected cells were divided into 2 equal aliquots, which were treated with 10−5 M RA or DMSO (0.1%), and RA activation was measured 14 hours later. All data represent the mean and standard error of at least 3 independent transient transfections.
