FIG. 4.
E2A and Id2 proteins modulate TrkB and p21Cip1 promoter activity in RA-treated cells. (A) TrkB (□) and p21Cip1 (░⃞) promoter constructs used in Fig. 3 were cotransfected with E12, E47, and Id2 expression vectors or the empty vector. Normalized expression levels produced by the respective promoter constructs after 7 days of RA treatment were made relative to those obtained in cycling cells. Average values and standard deviations obtained from three independent transfection experiments are shown. (B) BrdU incorporation as shown by immunofluorescence after 5 days of treatment with RA. Cells were cotransfected with a transfection reporter GFP plasmid and the Id2 expression vector (top panel) or the empty vector (bottom panel). Although some BrdU incorporation was observed in GFP-negative cells in both panels (see arrow in lower panel), cells positive for both BrdU incorporation and GFP expression were much more frequent if the Id2 expression plasmid was added (arrows in upper panel). (C) Quantification of the BrdU incorporation shown in panel B. As indicated in Materials and Methods, cells were counterstained with DAPI, and a minimum of 1,000 cells were evaluated in each transfection. The graph shows the percentages of BrdU-positive cells relative to GFP-positive cells.