Analysis of transcription factor expression during hindbrain neurogenesis. (A) Monocle3 pseudo-temporal ordering of 16 hpf, 24 hpf and 44 hpf hindbrain cells superimposed onto the aggregate UMAP. Cells are coloured based on their progression along pseudotemporal space (from pseudotime 0 in violet to the end of differentiation in yellow). (B) Individual pseudotemporal plots representing cell distribution at each developmental stage. (C) Heatmap showing selected TFs clustered by pseudotemporal expression pattern (q values<0.01). Pseudotime ordering is from left (progenitor state) to right (differentiated neurons). Selected transcription factors are shown for each group (G1-G7). The full gene list is in Fig. S13. (D-F) Expression of scrt1a, scrt1b and scrt2 during pseudotime. Whole-mount in situ hybridisation at 44 hpf for Scratch genes is shown in dorsal view (D′-F′), side view (D″-F″) and hindbrain sections (D‴-F‴). Scale bars: 50 µm. VN, ventral neurogenesis; DN, dorsal neurogenesis. (G) Using GENIE3, a directed network of interactions was predicted among the genes in the 44 hpf scRNA-seq dataset. The Scratch genes network was viewed and extracted in Cytoscape; boxes highlight TFs present in the above heatmap and colours match the group of origin in C.
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