Figure 9.
The lateral mobile fractions of NHE3 PBMIImut and NHE3 Δ4 were significantly increased compared with wild-type NHE3. A, OK cells depleted in NHE3 expression by the acid suicide technique were transiently transfected with HA-tagged NHE3 WT, NHE3 PBMIImut, and NHE3 Δ4 and labeled with monoclonal HA antibody as primary antibody and Alexa Fluor 488 anti-mouse as secondary antibody at 4 °C. FRAP was measured for NHE3 WT (■), NHE3 PBMIImut (●), and NHE3 Δ4 (▴) as described under “Experimental Procedures”. Data were collected as 30 images every 10 s. B, the mobile fractions (%) of NHE3 WT, NHE3 PBMIImut, and NHE3 Δ4 were determined at the 200-s time point. Each experiment analyzed FRAP results from more than five regions of interest from multiple cells. Experiments were repeated three times, and results are shown as mean ± S.D. Error bars represent S.D. p values were determined by unpaired t tests. NS, not significant; Mf, mobile fraction.