Fig. 2

Cav3.1 and CaM associate at a level that supports FRET. Fluorescence spectral confocal images of tsA-201 cells expressing constructs of GFP-Cav3.1 (donor molecule) or mKate-CaM (acceptor molecule) excited at either 457 nm or 561 nm. Plots show the excitation (vertical line) and the associated emission spectrum for each condition. a, b Applying excitation at 457 nm to cells expressing either GFP-Cav3.1 alone (a) or in conjunction with the mKate acceptor molecule (b) results in a typical GFP emission spectrum. c, d Applying excitation at 561 nm to cells expressing either mKate-CaM alone (c) or in conjunction with GFP-Cav3.1 (d) results in a pure mKate emission spectrum. e Coexpressing GFP-Cav3.1 and mKate-CaM results in a double emission profile when excitation at 457 nm promotes FRET in a subset of cells (arrows). A control image of the same cells excited by a 561 nm laser line at right reveals that a GFP-Cav3.1 expressing cell (arrowhead) that does not exhibit FRET failed to express the mKate-CaM construct. All results were derived from at least 3 separate experiments. Scale bars 10 μm