Figure 1.

Spatial distribution of PKA activity in polarized cells visualized by pmAKAR3. CHO-α4 cell expressing (A) pmAKAR3 or (B) pmAKAR3(TA) were plated at low densities on the α4β1-specific ligand CS1 and allowed to adhere for 30 min before image acquisition. Pseudocolored FRET images are shown (top), with the corresponding YFP/CFP emission ratio value color-coded scale. The corresponding YFP emission images are shown (bottom) for reference. Yellow arrowheads indicate regions of protrusive behavior visible by time-lapse imaging. Cells were stimulated with 25 μM forskolin at T = 0. (C) FRET ratio values of whole cell area (as represented in A and B) were computed and plotted against the indicated time points. Depicted is the mean and SD of data from three replicate measurements (n = 3 cells). (D) CHO-α4 cell expressing pmAKAR3 were plated on CS1 at a low density for 15 min before imaging. The representative cell as shown was selected based on a lack of a polarized morphology.