PP2A binding to CKA is required for DCV motility. (A) Representative kymographs of DCV motility for each of the genotypes measured. Bars, 5 µm. (B) Scatter plot of DCV flux measurements. Each point is the analysis from one kymograph. Bars indicate the mean and SEM for each genotype. BFP-CKA expression after CKA depletion significantly rescued the retrograde flux of DCVs and increased the anterograde flux of DCVs. In contrast, expression of a BFP-CKA fusion protein that cannot bind PP2A (BFP-CKA^ΔPP2A) after CKA depletion significantly reduced the flux of DCVs in both directions compared with CKA depletion alone. Unpaired, two-tailed t test result: *, P < 0.05; **, P < 0.005; ***, P ≤ 0.0001; ns, not significant. n = 12 axons. (C) Quantification of autophagic vesicle (AV) accumulation in terminal boutons of NMJ4 when BFP-Cka, BFP-Cka^ΔPP2A, mts^DN, or Pp2A-29B RNAi were expressed. (D) Quantification of autophagic vesicle accumulation in terminal boutons of NMJ4 when PP2A activity is disrupted using the elav-GeneSwitch driver. Berger’s unconditional exact test results: *, P < 0.05; **, P < 0.005; ns, not significant. (E) A phospho shift of endogenous CKA and GFP-tagged CKA isoforms was seen when cells were treated with okadaic acid, a PP2A inhibitor. Representative blots are shown. IB, immunoblot. n = 2 repeats.