Figure 3
Role of Gα 16 mutants in STAT3, ERK1/2, NF-κB and c-Jun phosphorylation. HEK293 cells were transfected with pcDNA3, wild type or QL mutants of Gα16, 2A, 3A or 5A. The transfectants were deprived of serum overnight, and cell lysates were prepared for SDS-PAGE separation. Phosphorylated form or native ERK1/2 (A), STAT3 (B), and NF-κB (C) were detected by Western blotting as indicated. (D) For measuring Gα16-triggered JNK activity, COS-7 cells were transiently transfected with each of the constructs mentioned together with a plasmid encoding HA-tagged JNK. Serum starvation was performed as mentioned and JNK assay was performed as described in Methods. Expression of tagged JNK was determined by anti-HA antibody. The activation of JNK was monitored by detecting the phosphorylation of GST-fused-cJun. The fold induction of the phosphorylation of various effectors were quantified and plotted on the right hand side for comparisons. * QL counterparts of the mutants stimulate phosphorylation of the detected proteins significantly over cells expressing wild type complements (Tukey-Kramer's test, p < 0.05). ‡ QL counterparts of 2A-, 3A- and 5A-stimulated phosphorylation were significantly lower than that of Gα16QL (Tukey-Kramer's test, p < 0.05).