Figure 1.

PLCγ1 and DAG Accumulate across the Synapse as It Forms

(A–C) CTLs expressing PLCγ1-EGFP and Lifeact-mApple upon first contact with EL4-blue target (t = 0), initial clustering and depletion of probes with time (min:s); 83% conjugates, n = 24; 14 independent experiments.

(A) Single confocal slices from Video S1A shown as merged and separate channels.

(B) En-face views across 3 μm slice of the synapse as single and merged channels with intensity plot for PLCγ1-EGFP.

(C) Pixel intensity (y axis) plots against distance across the synapse (x axis, μm) for boxed region shown in (A).

(D–F) CTLs expressing EGFP-PKCγ-C1 to detect DAG and Lifeact-mApple upon first contact between CTL and EL4-blue target (t = 0), initial accumulation and depletion of probes with time (min:s); 100% conjugates, n = 20; 10 independent experiments.

(D) Single confocal slices from Video S1B shown as merged and separate channels.

(E) En-face views across 3 μm slice of the synapse as single and merged channels with intensity plot for DAG (EGFP-PKCγ-C1).

(F) Pixel intensity (y axis) plots against distance across the synapse (x axis, μm) for boxed region shown in (D).

Scale bars = 3 μm. See also Figures S1 and S2.