C-Jun is a component of the transcription factor AP-1 that binds and activates transcription at TRE/AP-1 elements. The transcriptional activity of c-Jun is regulated by phosphorylation at Ser63 and Ser73 (1,2). Extracellular signals including growth factors, transforming oncoproteins and UV irradiation stimulate phosphorylation of c-Jun at Ser63/73 and activate c-Jun dependent transcription. Mutation of Ser63/73 renders c-Jun nonresponsive to mitogenic and stress induced signaling pathways. The MAP kinase homologue, SAPK/JNK, binds to the N-terminal region of c-Jun and phosphorylates c-Jun at Ser63/73. In addition, the activity of SAPK/JNK is stimulated by the same signals that activate c-Jun (3,4). Nonphosphorylated c-Jun (Ser63) II Cell Extracts: Total cell extracts from NIH/3T3 cells, prepared without treatment, serve as a negative control.