Hepatocytes are functionally heterogeneous and are divided into two distinct populations based on their metabolic zonation: the periportal and pericentral hepatocytes. During liver injury and regeneration, the cellular dynamics of these two distinct populations remain largely elusive. Here we show that major facilitator super family domain containing 2a (Mfsd2a), previously known to maintain blood–brain barrier function, is a periportal zonation marker. By genetic lineage tracing of Mfsd2a⁺ periportal hepatocytes, we show that Mfsd2a⁺ population decreases during liver homeostasis. Nevertheless, liver regeneration induced by partial hepatectomy significantly stimulates expansion of the Mfsd2a⁺ periportal hepatocytes. Similarly, during chronic liver injury, the Mfsd2a⁺ hepatocyte population expands and completely replaces the pericentral hepatocyte population throughout the whole liver. After injury recovery, the adult liver re-establishes the metabolic zonation by reprogramming the Mfsd2a⁺-derived hepatocytes into pericentral hepatocytes. The evidence of entire zonation replacement during injury increases our understanding of liver biology and disease.