Fig. 1

Identification of NEK7 as regulator of dendrite length and branching. a (Left) Cultured neurons at the depicted stages were subjected to microarray analysis (black arrowheads indicate sampling time points). Of the 1456 microtubule cytoskeleton-associated (MCA) genes, 377 were differentially expressed. (Right) Cluster analysis of differentially expressed genes. Expression profiles of non-MCA and MCA genes are depicted in gray and blue, respectively. Mean expression profiles are shown in red. Red boxes highlight overall upregulated clusters. b Hippocampal culture lysates from 1 to 21 DIV were immunoblotted against NEK7, and α-tubulin as loading control. c Quantification of NEK7 levels in western blots as in (b), normalized to α-tubulin; n = 3 independent hippocampal cultures. *P < 0.05 by two-tailed t-test. d Mouse cortex homogenates obtained at the indicated stages were immunoblotted against NEK7 and α-tubulin as loading control. e Quantification of NEK7 levels in western blots as in (d), normalized to α-tubulin; n = 3 independent homogenates. *P < 0.05 by two-tailed t-test. f Lysates of somato-dendritic and axonal compartments of hippocampal neurons at 9 DIV were immunoblotted against NEK7, Tau, and pan-histone, using α-tubulin as loading control. g One DIV neurons infected with control and Nek7 shRNA virus were lysed at 5 DIV and immunoblotted against NEK7. α-Tubulin served as loading control. h GFP-expressing control and NEK7-depleted neurons, transfected at 7 DIV were fixed at 14 DIV and stained with anti-GFP antibody. Scale bar, 50 μm. i Quantification of total dendrite length in control and NEK7-depleted neurons transfected at 1 DIV and analyzed at 5 DIV; n = 3 independent experiments. Total number of neurons analyzed: 90 (Control), 83 (shNek7#1), and 63 (shNek7#2); n.s. non-significant by two-tailed t-test. j Quantification of total dendrite length in 14 DIV neurons as in (h); n = 4 independent experiments. Total number of neurons: 35 (Control), 25 (shNek7 #1), and 28 (shNek7#2), *P < 0.05, **P < 0.01, ***P < 0.001 by two-tailed t-test. k Sholl analysis of 14 DIV neurons as in (h), schematic representation on the right. Mean number of intersections are plotted. Statistics as in (j). Error bars: s.e.m. Columns in all graphs show means and dot overlays individual data points