FIGURE 7.

The p47^phox-p22^phox interaction in the membrane of CHO 91/47/67 cells is dependent on the phosphorylation status of Thr-147. CHO 91/47/67 cells expressing no p22^phox (mock), wild type (WT), T147A, T147D, or P156Q p22^phox were stimulated with dimethyl sulfoxide (−) or 100 nm PMA (+) for 10 min, disrupted by sonication, fractionated, and the membrane fraction was solubilized as described under “Experimental Procedures.” p47^phox was immunoprecipitated (IP) from solubilized membrane; the immunoprecipitated proteins and those in a whole cell lysate were subjected to SDS-PAGE and Western blot analysis for p47^phox and p22^phox content. A, blots of p22^phox (top panel) and p47^phox (middle panel) in immunoprecipitates and p22^phox in whole cell lysates (bottom panel) from a representative experiment are shown. B, p47^phox-associated p22^phox in immunoprecipitates was normalized to p47^phox content and expressed as percent of that in PMA-stimulated wild type p22^phox-expressing cells. The open bar represents the mean of unstimulated conditions for all p22^phox constructs. The filled bars are the PMA-stimulated conditions for the indicated p22^phox construct. The data are the mean ± S.E. (error bars; n = 3). The p47^phox-associated p22^phox is significantly different from the wild type condition where indicated (*, p < 0.05; NS, not significant).