FIG 9.

HMGN5 promotes neurite outgrowth and controls chromatin structure in hippocampal neurons. (A) Representative confocal micrographs of βIII-tubulin-stained hippocampal neurons transfected with GFP, HMGN5-GFP, or HMGN5-GFP-3′ UTR. βIII-tubulin staining is shown in IBW contrast, while GFP signal is shown in green. Scale bar, 20 μm. (B) Neurite length measurement of DIV3 hippocampal neurons transfected with GFP, HMGN5-GFP, or HMGN5-GFP-3′ UTR (n = 220 to 260 neurites from 85 to 90 cells from three independent experiments, mean ± SD). Statistical significance was evaluated by a Kolmogorov-Smirnov test (***, P < 0.001; ns, not significant). (C) Representative confocal micrographs of DAPI- and H3K9me3-stained hippocampal neurons transfected with GFP, HMGN5-GFP, or HMGN5-GFP-3′ UTR. GFP signal is shown in green, DAPI staining is shown in IBW contrast, and H3K9me3 staining is shown in red. The confocal plane was chosen as the best focus on nuclei. Images were collected on the same day with identical exposure and scaling settings. Scale bar, 5 μm. (D) Measurement of the number of heterochromatic foci in the DAPI staining of hippocampal neurons transfected with GFP, HMGN5-GFP, or HMGN5-GFP-3′ UTR (n = 65 cells over three independent experiments, mean ± SEM). Statistical significance was evaluated by a two-tailed unpaired t test (*, P < 0.05; ns, not significant). (E) Quantification of mean fluorescence intensity of H3K9me3 foci in hippocampal neurons transfected with GFP, HMGN5-GFP, or HMGN5-GFP-3′ UTR (n = 20 to 40 foci, mean ± SEM). Statistical significance was evaluated by a two-tailed unpaired t test (*, P < 0.05; ns, not significant).