Figure 4. The effects of Arf6 and Rab22 on M₂ and M₄ internalization in JEG-3 cells.

A. JEG-3 cells were transfected with Flag-M₂ alone (white bars) or with either Arf6WT (black bars), Arf6T27N (dominant-negative; dark grey bars) or Arf6Q67L (constitutively active; light grey bars). B. JEG-3 cells were transfected with Flag-M₂ alone (white bars) or with either Rab22WT (black bars), Rab22S19N (dominant-negative; dark grey bars), or Rab22Q69L (constitutively active; dark grey bars). C. JEG-3 cells were transfected with Flag-M₄ alone (white bars) or with either Arf6WT (black bars), Arf6T27N (dominant-negative; dark grey bars), or Arf6Q67L (constitutively active; light grey bars). D. JEG-3 cells were transfected with Flag-M₄ alone (white bars) or with either Rab22WT (black bars), Rab22S19N (dominant-negative; dark grey bars), or Rab22Q69L (constitutively active; light grey bars). Following transfection, cells were stimulated for 0, 15, or 30 minutes with 1mM carbachol. Internalization of M₂ and M₄ receptors was measured using the binding of [³H]NMS to intact cells as described in ‘Experimental Procedures.’ Data represent the means ± S.E. of three to eight independent experiments, each performed in triplicate.