Cdkn1c expression depends on Hh in tailbud and on MRFs anteriorly MO-injected wild type (B–D) or myf5hu2022 incross embryos (A,B) co-injected with shha mRNA (D,E) analysed for cdkn1c mRNA in 12–15 ss dorsal (A,B,D) or 24 hpf lateral flatmount, dorsal to right (C,E). Anterior is to top. Bars: flatmounts 100 μm, sections 50 μm. (A–C). Adaxial cdkn1c mRNA accumulation initiates independent of MRF activity (arrowheads), but fails to be maintained (bracket). Myod MO reduces cdkn1c expression in older somites (brackets) but tailbud is unaffected (arrowheads). Myf5 and myod knockdown ablates somite cdkn1c mRNA except in small groups of cells lateral to the un-migrated adaxial cells (arrows). Neural, cloacal and transient notochordal expression is unaffected. (D). Over-expression of Shha promotes cdkn1c up-regulation in anterior PSM and somite in contorls and after Myod knockdown, but not in myf5 + myod double morphants (upper brackets). Note that Shha up-regulates cdkn1c in the lateral tailbud of myf5 + myod double morphants (lower brackets), but this is not maintained in anterior PSM. (E). Over-expression of Shha promotes ectopic smyhc1 expression at 24 hpf in control or myod morphants, but not in myf5 + myod double morphant embryos.