Fig. 5. In vivo analysis of PEA-15 mutants. (A) Elk-1-dependent transcriptional activity was measured in serum-stimulated CHO-K1 cell lysates transfected with expression plasmids encoding either wild-type PEA-15, mutant PEA-15 or control vector. The data are from an experiment done in triplicate and expressed as percentage inhibition of transcription relative to a vector-only control. (B) NIH 3T3 cells expressing either wild-type or mutant GFP–PEA-15 were serum starved for 16–20 h and then stimulated with 20% serum for 3 h and stained for ERK. The proteins were imaged by immunofluorescence in confocal microscopy. PEA-15 mutants such as I121R that are unable to bind ERK2 in vitro do not prevent ERK translocation to the nucleus upon serum stimulation (arrows), contrary to wild-type GFP–PEA-15 (arrows). The scale bar represents 20 µm.