Extended Data Fig. 9: Perturbing Nodal/BMP signalling during gastruloid self-organization.
From: Recording morphogen signals reveals mechanisms underlying gastruloid symmetry breaking
(A) Flow cytometry measurements of single-cell Wnt activity levels with and without Activin A pretreatment, measured at t = 72 h. (B) Flow cytometry measurements of single-cell Wnt activity levels with and without BMP4 pretreatment, measured at t = 72 h (left) and t = 96 h (right). (C) Dynamics of Nodal activity during gastruloid development, measured by flow cytometry of pooled and dissociated gastruloids. Substantial Nodal activity is detectable from 36-60, preceding CHIR stimulation at 48 and subsequently decaying to negligible levels by 72 h. (D) Nodal signalling levels in adherent cells treated for 48 h with different concentrations of recombinant Activin A (in N2B27). Dotted line is the mean gastruloid Nodal activity level at 48 haa under standard conditions (that is, initial aggregate of n = 200 cells) plotted for comparison. (E) Effect of a transient ActA treatment (100 ng ml−1, applied from 45-48 haa) on Wnt activity levels at 96. Transient ActA treatment continues to cause an enrichment in Wnt-off cells at 96 haa, even when duration of treatment is reduced. (F) Co-treatment of adherent Wnt reporter cells with BMP4 affects responses to Wnt3a treatment. Whereas moderate BMP4 (25 ng ml−1) treatment facilitated the Wnt response (middle), higher BMP4 (125 ng ml−1) treatment suppressed the Wnt response.
