Treatment of 3T3-L1 preadipocytes with differentiation inducers triggers a cascade in which C/EBPbeta is rapidly expressed, followed by C/EBPalpha and PPARgamma. C/EBPalpha and PPARgamma then activate the expression of adipocyte genes that produce the differentiated phenotype. Circumstantial evidence indicates that C/EBPbeta activates transcription of the C/EBPalpha and PPARgamma genes, both of which possess C/EBP regulatory elements in their proximal promoters. Although C/EBPbeta is expressed immediately upon induction of differentiation, acquisition of DNA binding activity is delayed for approximately 14h. Chromatin immunoprecipitation (ChIP) analysis conducted 24h after induction revealed that C/EBPbeta binds to C/EBP regulatory elements in the proximal promoters of the C/EBPalpha and PPARgamma genes. After an additional delay ChIP analysis showed that C/EBPalpha binds to its own promoter and to the promoters of the PPARgamma and 422/aP2 genes. These findings support the view that once expressed, C/EBPalpha is responsible for maintaining the expression of PPARgamma and C/EBPalpha, as well as adipocyte proteins (e.g., 422/aP2) in the terminally differentiated state. Together these findings provide compelling evidence that C/EBPbeta, C/EBPalpha, and PPARgamma participate in a cascade during adipogenesis.