Fig. 8

Wnt induces Notch signaling in the primordium via Fgf.

(A and A') The expression of the Fgf target pea3 is strongly increased in apc^mcr primordia at 36.5 hpf. (B and B') In apc^mcr primordia (B') atoh1a expression is increased compared to a sibling (B), but it still is restricted to individual cells. (C and C') her4 expression is slightly expanded in the primordium after Wnt upregulation by apc^mcr (C'). (D–F') The Fgf inhibitor PD173074 inhibits the expression of pea3 (D') and prevents the upregulation of atoh1a (E'). The expression of atoh1a is lost in the first proneuromast, as atoh1a is Fgf dependent. The more trailing central cells express atoh1a because it becomes self-regulatory. The Fgf inhibitor downregulates her4 in sibling (F) and apc^mcr mutant primordia (F'). (G–H') Loss of Wnt signaling in the primordium by heat-shock induction of dkk1b does not disrupt proneuromast formation in NICD (H') compared to heat-shocked sibling primordia (G') suggesting that Notch acts downstream of Wnt and Fgf in proneuromast formation. (I–L'') Notch inhibits Wnt signaling in the primordium. The expression of Wnt targets lef1 (I'), sef (J'), wnt10a (K') and dkk1b (L') expression is downregulated in NICD primordia. Conversely, in mib1^ta52b primordia lef1 (I''), sef (J'') and wnt10a (K'') are upregulated. (L'') dkk1b is downregulated in mib1^ta52b primordia, because Fgf signaling is secondarily lost (see text). (M–R') wnt10a expression expands only in the absence of Notch signaling in the primordium. (M–N') In apc^mcr primordia wnt10a expands towards the trailing region but is restricted to more central cells (arrows) (N,N'). (O,O') Downregulation of Notch with the γ-secretase inhibitor DAPT, causes a much more complete expansion of wnt10a in the trailing region of apc^mcr primordia, demonstrating that Notch signaling inhibits wnt10a in the primordium. (Q–R') wnt10a expression is expanded in the absence of Notch signaling when primordia are treated with Fgf inhibitor (Q,Q') but is once again restricted to the leading region of the primordium if NICD is activated in the absence of Fgf (R,R'). (S) Treatment of sibling embryos with the GSK3β inhibitor and Wnt activator BIO causes the upregulation of pea3. (S') BIO treatment of NICD embryos rescues the loss of pea3 in NICD primordia demonstrating that Wnt activates Fgf signaling upstream of Notch and that the loss of Fgf signaling in NICD is secondary to the loss of Wnt signaling. Wnt signaling is lost because Notch negatively feeds back to Wnt downstream of Fgf. All scale bars are 25 μm.