The mechanisms underlying the initiation and proliferation of liver regeneration (LR) has been extensively studied using the partial hepatectomy (PHx) model, while little is known about the termination of LR. PP2Ac alpha (protein phosphatase 2 A catalytic subunit a isoform) is the catalytic subunit of protein phosphatase 2 A (PP2A), accounting for most of intracellular serine/threonine phosphatase activity. We have previously observed that termination of LR delayed in PP2Ac alpha liver-specific knockout (LKO) mice after PHx. In our study, we used phospho explorer antibody array analysis to screen the potential phosphorylation targets of PP2Ac alpha, and PP2Ac alpha had a great influence on the hepatic phosphoproteomic signaling in the termination of LR after PHx. We then tested the phosphorylation changes and metabolic function of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-2 (PFKFB2), an isoform of the key glycolytic enzyme PFKFB, which was significantly regulated by PP2Ac alpha knockout. PP2Ac alpha knockout enhanced glycolysis in vivo and in vitro, while adenoviral-mediated RNAi of PFKFB2 reversed the extension of postoperative liver regeneration in KO mice along with the downregulation of glycolysis. Therefore, we demonstrated that PP2Ac alpha liver-specific knockout regulated the hepatocytes glycolysis via activating PFKFB2, thus enhancing liver regeneration during the termination stage. (C) 2020 Elsevier Inc. All rights reserved.