Fig. 2

LMTK2 is complexed with both KLC1 and kinesin-1 and mutation of LMTK2(WD1388/1389) to AA disrupts this binding. a HEK293 cells were transfected with either control empty vector (Ctrl), EGFP-LMTK2 + FLAG-KLC1 + myc-kinesin-1A or EGFP-LMTK2(WD1388/1389AA) + FLAG-KLC1 + myc-kinesin-1A. LMTK2 was immunoprecipitated via the EGFP-tag and bound KLC1 and kinesin-1 detected by immunoblotting. Both inputs and immunoprecipitations (IP) are shown. b and c Endogenous LMTK2 and KLC1 are complexed in HEK293 cells (b) and rat cortical neurons (c). LMTK2-KLC1 PLAs were performed using rabbit anti-LMTK2 and goat anti-KLC1 antibodies. Control experiments involving omission of both or each primary antibody are also shown. Representative phase contrast images with PLA signals of cells are displayed; arrows indicate PLA signals and in (c) show signals in axons. To facilitate viewing of signals in these phase contrast images, the PLAs colours were electronically changed from red to pale blue. Scale bars = 20 μm. Graphs show PLA signals per cell in the different experiments. Data were analysed by Welch’s ANOVA and Games-Howell post hoc test. b N = 40–46 cells, c N = 43–54 cells; error bars are s.e.m., ***p < 0.001