Figure 6.

Hedgehog and Wnt Signaling Display Different Temporal Dynamics in PSC Lines with Inherent Tendencies to More Cortical or Ventral Differentiation Outcomes

(A) Early-stage differentiations classified as highly or partially ventralized express higher levels of SHH compared to dorsalized and caudalized differentiations, as well as levels of Hedgehog signaling readouts PTCH1, GLI1, and GAS1 consistent with increased pathway activity. Dorsalized differentiations express higher levels of Wnt signaling readouts AXIN2 and TNFRSF19 compared to ventralized differentiations but lower AXIN2 compared to partially caudalized differentiations (pairwise Welch’s t test, false discovery rate [FDR]-corrected p values: ^∗p < 0.05; ^∗∗p < 0.01; ^∗∗∗p < 0.001; ns, not significant). Only comparisons to the dorsalized class are shown. Differentiations per group: dorsalized, 86; highly ventralized, 23; partially ventralized, 23; and partially caudalized, 14.

(B) Early-stage differentiations from separate PSC lines vary in average expression of Hedgehog and Wnt signaling pathway activation, consistent with different tendencies in regional patterning (one-way ANOVA: ^∗p < 0.05; ^∗∗p < 0.01). Error bars represent standard error; n = 2–6 differentiations per line.

(C) Gene expression time course during differentiation for selected genes associated with forebrain regions and Hedgehog or Wnt signaling. Differentiations from ventral-prone lines (partially ventral iPSC22.1 and highly ventral GMESC01.1, iPSC01.1, iPSC06.1, and iPSC14.1, n = 8–11) were compared to differentiations of a dorsal-prone line (iPSC21.1, n = 1–2). Profiles represent average gene expression, and error bars represent standard deviation. Significance shown for dorsal versus ventral comparison at 17 and ∼35 dpi (Welch’s t test, FDR-corrected p values: ^∗p < 0.05; ^∗∗p < 0.01; ^∗∗∗p < 0.001).