Normal and SV40 virus-transformed WI-38 human lung fibroblasts were serum starved and refed, or synchronized by double thymidine block and released from the block. At different time points in the cell cycle, steady state levels of P120 mRNA and P120 protein content of the cells were determined by densitometric scans of Northern and Western blots. At the same time points, [3H]thymidine uptake was measured and flow cytometric analysis performed for DNA content and P120 antigen staining. Levels of P120 protein and P120 mRNA were approximately 4 times greater in non-synchronous, exponentially growing transformed cells than in similarly growing normal cells. Early G1-phase cells, synchronized either with serum deprivation or with metabolic block, contained only a trace amount of P120 protein and mRNA. The P120 gene was transcribed early in G1 and P120 protein synthesis initiated in middle G1. A dramatic increase of P120 protein level occurred in S-phase with a corresponding mRNA peak preceding the P120 protein peak. These results indicate that P120 is overexpressed in transformed WI-38 cells and that P120 is temporally regulated during the cell cycle of both transformed and normal fibroblasts. The dramatic increase in P120 protein expression at the G1 to S boundary suggests that P120 may play a role in the regulation of cell cycle and increased nucleolar activity that is associated with cell proliferation.