It is believed that adult tissues in mammals lack the plasticity needed to assume new developmental fates because of the absence of efficient pathways of dedifferentiation. However, the well-documented ability of the transcription factor pancreatic and duodenal homeobox gene 1 (PDX-1) to activate pancreatic lineage development and insulin production following ectopic expression in liver suggests a surprising degree of residual plasticity in adult liver cells. This study seeks a mechanistic explanation for the capacity of PDX-1 to endow liver cells with pancreatic characteristics and function. We demonstrate that PDX-1, previously shown to play an essential role in normal pancreatic organogenesis and pancreatic β-cell function and to possess the potential to activate multiple pancreatic markers in liver, can also direct hepatic dedifferentiation. PDX-1 represses the adult hepatic repertoire of gene expression and activates the expression of the immature hepatic marker α-fetoprotein. We present evidence indicating that PDX-1 triggers hepatic dedifferentiation by repressing the key hepatic transcription factor CCAAT/enhancer-binding protein α. Hepatic dedifferentiation is necessary though not sufficient for the activation of the mature pancreatic repertoire in liver. Conclusion: Our study suggests a dual role for PDX-1 in liven inducing hepatic dedifferentiation and activating the pancreatic lineage. The identification of dedifferentiation signals may promote the capacity to endow mature tissues in mammals with the plasticity needed for acquiring novel developmental fates and functions to be implemented in the field of regenerative medicine.