FIG. 4.

NF-κB activity in Rac2-deficient mast cells. Wild-type (WT) and Rac2^−/− C57BL/6 BMMCs were stimulated with 10 ng of SCF per ml for 1 h and then harvested for nuclear extracts and cytosol lysates. (A) Western blot analysis with an anti-NF-κB antibody (Ab). Levels of total p38 MAPK proteins were used as a loading control. Wild-type BMMCs were transduced with retroviral vectors expressing wild-type Rac2 or wild-type Rac1 [designated WT (Rac2) and WT (Rac1)]. (B) Electrophoretic mobility shift assay. We used 5 μg of nuclear extracts for binding to the NF-κB consensus sequence labeled with [³²P]dCTP, and DNA-protein complexes were examined on a 7% polyacrylamide gel. A 50-fold excess of unlabeled oligonucleotide (NF-κB comp) was added to the competition assay with wild-type (WT) nuclear extracts. No nuclear extract protein was used in the negative control lane. The results shown are representative of three experiments.