Fig. 4

NEK7 promotes Eg5 accumulation in distal dendrites through S1033 phosphorylation. a Neurons transfected with the indicated plasmids at 7 DIV were fixed and stained with FLAG and Eg5 antibodies. Magnifications show Eg5 staining in proximal (P) and distal (D) dendrite regions. Scale bar, 50 μm. b Quantification of the Eg5 intensity in ~50 µm segments of proximal and distal dendrites of neurons as in (a); n = 3 independent experiments (total number of neurons: 27 (Control), 24 (Nek7 depleted), and 23 (rescue with WT Nek7), 2 to 6 dendrites analyzed per neuron); n.s. non-significant, *P < 0.05, **P < 0.01, by two-tailed t-test. c Eg5 intensity in somas of neurons as in (a). Statistics as in (b). d FLAG staining in the same neurons as in (a) was quantified as control. Intensities were plotted relative to the intensities in the somas of the same cells. Statistics as in (b). e Seven DIV neurons were infected with control or Nek7 shRNA virus, and co-transfected with FLAG-Eg5 (wild-type or S1033 mutants) and GFP plasmids, and fixed at 14 DIV. Images show dendrites after staining with GFP and FLAG antibodies. Scale bar, 25 µm. f Quantification of FLAG-Eg5 intensity in ~50 μm segments of proximal and distal regions of dendrites in 14 DIV neurons as in (e), normalized to GFP intensity and to FLAG-Eg5 intensities in the somas of the same cells; n = 3 independent experiments (total number of neurons: 20 to 22 per condition, 2 to 5 dendrites measured per neuron); n.s. non-significant, *P < 0.05, **P < 0.01 by two-tailed t-test. Error bars: s.e.m. Columns in all graphs show means and dot overlays individual data points