The biological significance of phosphatidylcholine-specific phospholipase C (PC-PLC) in hepatocarcinogenesis and the proliferation and differentiation of rat liver cancer cells was investigated. The Ca 2+ -dependent activities of PC-PLC gradually increased during N-nitrosodiethylamine (DEN)-induced hepatocarcinogenesis and peaked at weeks 18-20 when the tumour formed. There was a close relationship between Ca 2+ -dependent PC-PLC activities and cellular DNA content, membranous -glutamyltranspeptidase ( -GT), and tyrosine protein kinase. In contrast, Ca 2+ -independent PC-PLC activities decreased during hepatocarcinogenesis. Similarly, when CBRH-7919 rat liver cancer cells were treated with phorbol 12-myristate 13-acetate, a proliferation stimulator of the cells, -GT and Ca 2+ -dependent activities of PC-PLC and the expression of -fetoprotein increased significantly. However, when these cells were induced by retinoic acid to differentiate, Ca 2+ -dependent PC-PLC and -GT activities decreased significantly, together with -fetoprotein expression. There was a close relationship between Ca 2+ -dependent PC-PLC and -GT activities during differentiation as there was during proliferation. We suppose that Ca 2+ -dependent PC-PLC is involved in rat hepatocarcinogenesis induced by DEN and that it plays an important role in the phorbol ester-induced proliferation or retinoic acid-induced differentiation of liver cancer cells.