Cell cycle progression of mouse macrophage cells was impaired by interferon-gamma (IFN-gamma). The blockage of G1/S transition was associated with diminution of cyclin-dependent kinase-2 (CDK2)-associated kinase activities. The expression of p21Cip1 was not upregulated by IFN-gamma. Instead, the physiologic downregulation of p27Kip1 necessary for normal cell cycle progression did not take place sufficiently in the presence of IFN-gamma. During normal cell cycle progression, the levels of p27Kip1 were maximal at early G1 and then decreased gradually. In the presence of IFN-gamma, however, the levels of p27Kip1 discontinued to decrease at a late mid G1 point and were consistently as high as, or higher than, levels observed there. The steady, relatively high-level attachment of p27Kip1 to CDK2 contributed to the insufficient formation of active cyclin/CDK2, possibly deferring cells from entering S phase.