FcγR1 activates Rac. (A) The activation of total Rac and Rac2 was measured following FcγR1 stimulation in U937 cells (See Methods) using GST-PAK1_CRIB domain pull down assay. U937 cells were treated with PMA or PKC inhibitor (GF109203X) to determine the effect of PKC on Fcγ receptor desensitization. The protein bound to GST-PAK1-CRIB is resolved on SDS-PAGE and immunoblotted with antibody for total Rac. Same experiment was repeated and immunoblotted with antibody specific for Rac2. Lane 1, no stimulation; lane 2 & 3, preincubated with PMA (200 ng/ml) for 5 min. at 37°C followed by FcγR stimulation for 1 and 5 min. respectively, lane 4 & 5, cells were preincubated with GF 109203X (2.5 μM) for 15 min. on ice and then treated with PMA (200 ng/ml) for 1 & 5 min. followed by FcγR1 stimulations, lane 6 and 7, FcγR1 stimulation (alone) on Rac-GTP levels. The experiment was repeated three times. (B) Graphic representation of the signaling mechanism reported in the present study.
Joshi et al. BMC Immunology 2014 15:18 doi:10.1186/1471-2172-15-18