Following partial hepatectomy (PH), there is a rapid and highly orchestrated series of biochemical events which occur prior to cellular proliferation. Some of these events are presumably intimately linked with the eventual regeneration of the liver, whereas others are likely to be stress related or required for the continued differentiated function of the liver while regeneration is occurring. The regulation of the AP-1 transcription factor c-Jun during hepatic regeneration has been studied here. There is a progressive increase in c-Jun-mRNA levels after sham operation, one-third PH, and two-thirds PH. A concomitant increase in activating protein 1 (AP-1) binding activity is also observed. The c-Jun protein is a major constituent of the AP-1 complex in quiescent and early regenerating liver. The activity of c-Jun amino-terminal kinase (JNK), which phosphorylates the activation domain of the c-Jun protein, is markedly stimulated after one-third and two-thirds PH. c-Jun amino-terminal kinase-1 is a constituent of this stimulated JNK activity after PH. When primary cultures of adult rat hepatocytes are incubated with epidermal growth factor or transforming growth factor-alpha, AP-1 transcriptional activity is increased and the activation domain of the c-Jun protein is further potentiated. Phosphopeptide mapping of the endogenous c-Jun protein in proliferating cultured hepatocytes demonstrates phosphorylation of the c-Jun activation domain. Pretreatment of animals prior to PH with a neutralizing antibody to tumour necrosis factor-alpha (TNFalpha), inhibits hepatocyte DNA synthesis and JNK activation. It is concluded that the stimulation of one-third or two-thirds PH activates JNK through a mechanism that requires TNFalpha, which phosphorylates the c-Jun activation domain in hepatocytes, resulting in enhanced transcription of AP-1-dependent genes. Although nuclear factor-kappa B (NFkappaB) binding activity is induced during liver regeneration following PH, the physiological consequence of this induction is unknown. The role of NFkappaB during liver regeneration has been assessed by delivering to the liver a super-repressor of NFkappaB activity using an adenoviral vector expressing a mutated form of IkappaB. This adenovirus (Ad5IkappaB) was almost exclusively expressed in the liver and inhibited NFkappaB DNA binding activity and transcriptional activity in cultured cells as well as in the liver in vivo. Following PH, Ad5IkappaB, but not a control adenovirus (Ad5betagal), resulted in the induction of apoptosis as demonstrated by histological staining and TUNEL analysis. In addition, infection with Ad5IkappaB but not Ad5betagal decreased the mitotic index following PH. These two phenomena, increased apoptosis and cell cycle arrest, were associated with liver failure in animals infected with the ad5IkappaB but not Ad5betagal as demonstrated by elevated serum bilirubin and ammonia levels. Thus, the induction of NFkappaB during liver regeneration following PH appears to be a required event to prevent apoptosis and to allow for normal cell cycle progression.