Fig. 1
The effects of Rp58 disruption on p27kip1 within the embryonic cortex. a Immunostaining of Rp58 and p27kip1 to reveal their co-presence in cells of the VZ/SVZ, IZ and CP. Arrows point to double-positive cells, while arrowheads point to Rp58+ cells. b–d Knockdown or overexpression of Rp58 did not significantly disrupt p27kip1 co-immunostaining of GFP+ cells in the IZ (F2,6 = 3.1 p = 0.11, One-way ANOVA, >300 cells counted from 3 independent brains per condition) (c), while there was a significant effect in the VZ/SVZ (F2,6 = 45, p < 0.0005, One-way ANOVA, >400 cells counted from 3 independent brains per condition). Arrows point to double-positive cells, while arrow heads point to GFP+ cells. e, f Knockdown of Rp58 leads to a decrease in the proportion of GFP electroporated cells which co-label with βIII-tubulin, indicative of impaired neurogenesis (F2,6 = 18 p = 0.003, One-way ANOVA, >600 cells counted from 3 independent brains per condition). Arrows point to GFP+ cells which do not co-label with βIII-tubulin. g GFP labelled cells were dissociated from successfully electroporated brain tissue and plated for 2 h before fixation, immunostaining and data collection. Knockdown of Rp58 leads to a reduction in neurogenesis, while forced expression does not significantly disrupt p27kip1 expression (F2,6 = 9 p = 0.011, One-way ANOVA, >450 cells counted from 3 independent experiments per condition). Arrow points to double-positive cells, while arrow head points to GFP+ cells.. (H) Forced expression of Neurog2 but not Rp58 in P19 embyrocarcinoma cells induces neurogenesis, as evaluated by immunostaining for βIII-tubulin. *p < 0.05, **p < 0.01, ***p < 0.001; “ns” denotes not significant. Scale bar represents 50 μm