Extended Data Fig. 3: Differential cadherin code and cortical tension regulate self-organization in ETX embryos.
From: Stem cell-derived synthetic embryos self-assemble by exploiting cadherin codes and cortical tension
(a) Time course of formation of correctly-sorted ETX embryos following seeding. 0.5-h: 0/515 structures; 12-h: 79/1292 structures; 24-h: 160/1074 structures; 48-h: 134/888; 72-h: 93/702 structures. N = 3 for each condition. Data are presented as Mean ± SD. Statistics calculated by unpaired two-tailed Student’s t test. (b) Live cell imaging and tracking. H2B-RFP-XEN (green), H2B-CFP-ES (red) and EGFP-TS (blue) were overlaid with Imaris cell-tracking spheres. (c) Quantification of mobility for different types of cells during self-organization. Data are presented as Mean ± SD at different time points. (d) The bar graph shows the average mobility for different cell types during self-organization at different time ranges after cell seeding. Data are presented as Mean ± SEM. 12 structures from 3 independent experiments were imaged for quantification. Statistics calculated by unpaired two-tailed Student’s t test. (e) Examples of structures made from low (control) and high number of XEN cells, stained at day-1 and day-3. Experiments were repeated 3 times. Scale bar, 10 μm. (f) Schematic of morphological transitions when using low and high number of XEN cells. (g) Cortical stiffness measurements for indicated cell types before and after treatment with Blebbistatin (Bleb). Total measured cell numbers for each condition: ES: n = 58; ES + Bleb: n = 34; TS: n = 68; TS + Bleb: n = 31; XEN: n = 68; XEN + Bleb: n = 35. Data are presented as Mean ± SD. Statistics calculated by ANOVA with a multiple comparison test. (h) Day 3 well-sorted ETX embryos were cultured with either blebbistatin, cytochalasin D or DMSO (control) during consolidation stage for 24 hrs and immuno-stained to reveal the indicated markers. Quantification shows the percentage of disorganized ETX structures. n = 84 (Bleb treated), n = 83 (Cyto D treated) and n = 75 (control), N = 3 for each condition. Data are presented as Mean ± SD. Statistics calculated by unpaired two-tailed Student’s t test. Scale bar, 100 μm. (i) CPM modelling shows the effect of XEN cell stiffness (λP) on externalization efficiency (N = 474). The sorting efficiency calculated for each time-point is plotted as a heat-map, overlayed with contours (white dotted lines). Source numerical data are available in source data.
