GM-CSF and SLF combined treatment stimulates MAP kinase activity in factor-starved MO7e cells. Factor-starved MO7e cells were treated with GM-CSF (100 U/mL), SLF (50 ng/mL), or the combination of GM-CSF plus SLF for 10 minutes or were treated with MIP-1α (50 ng/mL), IP-10 (50 ng/mL), or PD098059 (50 μmol/L) for 1 hour before treatment with GM-CSF plus SLF for 10 minutes. Cell lysates containing eqivalent amounts of total protein were combined with MBP in the presence of [³²P]ATP and MAP kinase activity assessed by SDS-PAGE and autoradiography, as described in the Materials and Methods. Combined treatment of MO7e cells with GM-CSF and SLF stimulated an increase in MAP kinase activity (upper panel, lane 7) to a greater extent than that evoked by treatment with either cytokine alone (upper panel, lanes 3 and 4). Pretreatment with MIP-1α, IP-10, or PD098059 suppressed MAP kinase activity stimulated by GM-CSF in combination with SLF (upper panel, lanes 8, 9, and 10). Two predominant isoforms of MAP kinase, noted to the right of the lower panel as ERK1 and ERK2, were detected in MO7e lysates. ERK1 and ERK2 protein content were not altered by any of the treatments indicated (lower panel). Similar results were obtained in two additional experiments.