Figure 4
From: miR-200 family controls late steps of postnatal forebrain neurogenesis via Zeb2 inhibition
miR-200 induces calretinin expression through Zeb2 inhibition.
(a) Zeb2 mRNA (red) is widely expressed in the forebrain with particularly prominent presence in the SVZ and RMS. (b) Images showing GFP cells in the RMS stained with Zeb2 antibody 4 days after in vivo electroporation in control or miR-200-gof conditions. (c) Quantification of mean Zeb2 staining intensity 4 days after in vivo electroporation cells at 4 dpe in control or miR-200-gof conditions. This showed a significant reduction in Zeb2 protein expression in neuronal precursors, regardless of their calretinin expression status. Differences between groups of cells were analyzed pairwise with a t-test (control vs miR-200 calretinin positive P < 2.2e-16; control vs miR-200 calretinin negative P < 2.2e-16); n = number of cells used for analysis; an = number of animals from which analyzed cells were issued. (d) Zeb2 expression normalizes the miR-200-gof mediated induction in calretinin expression. Differences between groups were analyzed pairwise with the Man and Whitney test (control (n = 5 animals) vs miR-200 (n = 5 animals) P = 0.008816, miR-200 (n = 5 animals) vs miR-200 + Zeb2 (n = 7 animals) P = 0.04236). (e) Calretinin immunostaining of coronal forebrain section through the SVZ of Gsh2-Cre; Zeb2+/+ (wt) or Gsh2-Cre; ZebFl/Fl knockout (Zeb2 −/−) animals at P5 at the level indicated in the schema. (f) The number of calretinin immunoreactive cells in the aSVZ. is much higher in knockout (Zeb2 −/−) than in control (wt) animals. (g) qRT-PCR analysis in FACS sorted SVZ cells from P2 animals reveals a massive increase in calretinin mRNA expression in knockout (Zeb2 −/−) compared to control (wt) animals. In (f,g) difference between groups was analyzed using t- test. Scale bars: 1 mm in a, 20 μm in (b) 200 μm in (e).
