Fig 6

(A) Four putative KLF-binding motifs were identified in the upstream region of the dlc promoter. The core binding motif (CACCC) is indicated by red text (a). Chromatin immunoprecipitation was performed using anti-Klf4 or anti-Myc antibodies, and qPCR of the isolated chromatin revealed significant enrichment of the KLF-binding motif located between -756 to -747 of the dlc promoter (b). (B) A diagram depicting the organization of dlc11k-mCherry, dlc3k-mCherry, and dlc3kM-mCherry constructs. KLF binding motif is shown in red text, and mutated bases are indicated by lowercase letters (a). Images of 5s embryos labeled with dlc antisense RNA probe (b), F1 transgenic embryos from Tg(dlc11k:mCherry) (c-e), Tg(dlc3k:mCherry) (f, g) and Tg(dlc3kM:mCherry) (h, i) are shown. Five somites are indicated (e). (C). Uninjected (un-inj) Tg(dlc3k:mCherry) (a) or Tg(dlc3kM:mCherry) (e), LacZ-injected Tg(dlc3k:mCherry) (b) or Tg(dlc3kM:mCherry) (f) and klf4-injected Tg(dlc3k:mCherry)(c) or Tg(dlc3kM:mCherry) (g) embryos were stained with anti-mCherry antibody at bud stage. Quantification of results from (a-c) is shown (d). Statistical significance was determined by Student’s t-test. ***p < 0.001. Error bars indicate standard error. CG, cranial ganglia; EI, epidermal ionocytes; S, somites; PSM, presomitic mesoderm.