Sox2 and Pax6 Play Counteracting Roles in Regulating Neurogenesis within the Murine Olfactory Epithelium
Fig 7
Sox2 transduction enhances Ascl1 expression, whereas Pax6 transduction leads to a reduction in Ascl1 expression and a reciprocal increase in the prevalence of downstream neural precursor markers.
(A) A proneural bHLH cascade operates during olfactory neurogenesis and defines a hierarchy of GBCs as they progress from transit amplifying (TA) progenitor cells into differentiated neurons, by way of two immediate neural precursor (INP) stages. (B-M) At 8-days post-infection (9-days post-lesion), tissue was immunolabeled for Ascl1, Neurog1, and NeuroD1 along with GFP in clones derived from EV (B, F, J), Sox2 (C, G, K), Pax6 (D, H, L) and SEP (E, I, M) RVV-transduction. Hollow arrows mark GFP (+) cells within a given clone that co-express Ascl1, Neurog1, or NeuroD1. In contrast, solid arrows mark non-transduced, i.e., GFP-lacking, cells that express these factors. Dashed lines mark the basal lamina and scale bar in K corresponds to 20 μm in panels B-M. (N-P) Quantitative analysis of the effect of the various transduction conditions on the percentage of cells in a clone that express Ascl1 (N), Neurog1 (O) and NeuroD1 (P). For the graph, the percentage of cells marked by joint expression of the individual transcription factor and GFP within each clone are graphed in a box-and-whiskers scatter plot; the median number of each distribution is marked by the horizontal black line within the box. For Ascl1+ GBCs, the median percentages for EV vs. Sox2 vs. Pax6 vs. SEP are 0, 36, 0, 20, respectively. For Neurog1+ GBCs, the median percentages were 0, 37, 17, 25, respectively. Finally, for NeuroD1+ GBCs, the median values were 0, 50, 33, 19, respectively. Multiple pair-wise comparisons (horizontal line) were found to be statistically significant using Dunn’s method (* p < 0.05) following the demonstration of overall significance using Kruskal-Wallis One-Way ANOVA tests. (Q) Percentage of clones that contain cells positive for labeling with the bHLH factors. To compare percentages statistically, data were first arcsin transformed prior to normality testing, standard parametric ANOVA, and post-hoc Holm-Sidak pair-wise testing (**p < 0.01). Asterisks signify comparisons to EV, except for the comparison in number of Ascl1-conatining clones between Sox2 and Pax6 vectors, which is designated by the orthogonal lines.