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. 2016 Apr 12;12(4):e1005984. doi: 10.1371/journal.pgen.1005984

Fig 1. Notch pathway controls Type I>0 daughter proliferation switch in NB3-3A independently of programmed cell death.

Fig 1

(A) Embryonic Drosophila CNS at St11; NBs outlined. (B) NBs can divide by Type I or Type 0 mode, and many lineages display a Type I>0 switch [23]. (C-E) kuze29-4 mutants have normal NB numbers (n = 40 T2-A2 segments; Student’s two-tailed T-test; +/-SD). (F-I) The Type 0 cells in the NB3-3A lineage can be visualized by Eve expression. (F) In control at St16, an average of 9.5 Eve cells is observed. (H) ED225 programmed cell death (PCD) mutants show similar numbers (E). (I) kuze29-4;ED225 double mutants show additional Eve cells, when compared to both kuze29-4 and ED225 single mutants. (J) Quantification of Eve cell number (* p ≤0.05, ** p≤0.01, *** p≤0.001; n≥60 clusters; Kruskal-Wallis, Dunn’s posthoc; +/-SD). (K) Canonical Notch pathway. (L-M) In line with the extra Eve cells found in the kuze29-4;ED225 double mutants, staining for apoptotic cells, using anti-cleaved Caspase3 as read-out, revealed increased PCD in kuze29-4. There was however no sign of premature cell death of the NB itself. (N-O) NB and daughter proliferation analysis in NB3-3A. (N) As previously described [23], in control after St11 we exclusively detected cell division of the NB itself (Dpn+) (0% daughter divisions; n≥88 lineages). (O) In contrast, in kuze29-4, we readily detected dividing daughter cells (28% daughter divisions; n≥65 lineages; dorsal and intermed refers to dorsal and intermediate confocal layers, respectively). (Q) In wild type, the NB3-3A undergoes one Type I and 11 Type 0 divisions. The NB exits cell cycle at St15, and undergoes PCD at St17 [23]. There is limited PCD, in the early parts of the lineage. Additional Eve cells in kuz result from a failure in the Type I>0 switch. In kuz mutants, some of the aberrantly generated cells are removed by PCD. In ED225 mutants, the NB does not undergo PCD, but does not progress further [23]. In kuz,ED225 mutants, aberrantly generated cells survive, hence increasing the Eve cell numbers beyond that observed in kuz alone.