Extended Data Fig. 11: Volume EM of notochord in notoroids.

a, Low resolution SBF SEM image of a notoroid oriented to show transverse sections of the notochord. Neuroepithelium (n) and mesenchymal cells (m) are shown. Orange (b) and green (c) coloured boxes show regions of interest with putative notochord cells. b-c) High resolution SBF SEM images of notochord. Regions of interest shown in panels d-i are denoted with colour boxes. d-i) Cropped and enlarged SBF SEM images showing representative features of notochord in notoroids (d-f), mesenchymal cells (g) and neuroepithelial cells near the notochord (i). Notochord cells have abundant cytoplasmic glycogen (magenta stars), lipid droplets (yellow stars) and vacuoles (cyan arrows), are covered by a basal lamina (yellow arrows) and surrounded by a sheath of ECM fibres (f). The cells are closely packed, connected by desmosomes (purple arrow in h) and often ciliated (pink arrow in e). Dilated ER (green arrows) and endolysosomal structures (blue arrows) are frequent. In some cells, intermediate filaments (black arrow in d) and rough ER (dark green arrow in h) were observed. Mesenchymal cells are star-shaped with long protrusions in contact with neighbouring cells. Neuroepithelial cells in proximity to the notochord have some vacuoles and show an accumulation of mitochondria (orange arrow) towards the basal lamina. Scale bars: 50 µm (a), 10 µm (b-c), 3 µm (d-i).