The intracellularly acting protein toxin of Pasteurella multocida (PMT) causes numerous effects in cells, including activation of inositol 1,4,5-trisphosphate (IP₃) signaling, Ca²⁺ mobilization, protein phosphorylation, morphological changes, and DNA synthesis. The direct intracellular target of PMT responsible for activation of the IP₃ pathway is the G_q/11-protein, which stimulates phospholipase C (PLC) 1. The relationship between PMT-mediated activation of the G_q/11-PLC-IP₃ pathway and its ability to promote mitogenesis and cellular proliferation is not clear. PMT stimulation of p42/p44 mitogen-activated protein kinase occurs upstream via G_q/11-dependent transactivation of the epidermal growth factor receptor. We have further characterized the effects of PMT on the downstream mitogenic response and cell cycle progression in Swiss 3T3 and Vero cells. PMT treatment caused dramatic morphological changes in both cell lines. In Vero cells, limited multinucleation, nuclear fragmentation, and disruption of cytokinesis were also observed; however, a strong mitogenic response occurred only with Swiss 3T3 cells. Significantly, this mitogenic response was not sustained. Cell cycle analysis revealed that after the initial mitogenic response to PMT, both cell types subsequently arrested primarily in G₁ and became unresponsive to further PMT treatment. In Swiss 3T3 cells, PMT induced up-regulation of c-Myc; cyclins D1, D2, D3, and E; p21; PCNA; and the Rb proteins, p107 and p130. In Vero cells, PMT failed to up-regulate PCNA and cyclins D3 and E. We also found that the initial PMT-mediated up-regulation of several of these signaling proteins was not sustained, supporting the subsequent cell cycle arrest. The consequences of PMT entry thus depend on the differential regulation of signaling pathways within different cell types.