Klf4 protein expression pattern and colocalization of Klf4/p63 and Klf4/dlc during late gastrulation and early somite stages.
(A) Klf4 is expressed in the epiblast of the deep cell layer (DEL), yolk syncytial layer (YSL), and enveloping layer cells (EVL) of 60%, 70%, 80% and 90% epiboly embryos. Insets show respective XZ or YZ projections of confocal images from an 80% epiboly embryo along the axes shown in the main panel (d). Klf4 is expressed in the ectoderm and EVL of bud (tb), 5s and 10s embryos. (B) Images of p63 expression during different embryonic developmental stages are shown (a-d). Colocalization of Klf4 and p63 was detected in ventral ectoderm of embryos at various stages, including 90% epiboly, bud, 5s and 10s (e-h). Enlargements of Klf4 and p63 merged images (e-h) are shown from the areas indicated in lower magnification images (a-d). Percentage of colocalization of Klf4 and p63 in respective stage is indicated within each panel. Some non-colocalized cells showing only expression of Klf4 (arrowhead) or p63 (arrow) were observed in the ventral ectoderm of embryos at 5s and 10s. Insets show YZ projections of confocal images. (C) Images of wild-type embryos stained with anti-Klf4 antibody and dlc antisense RNA at 90% epiboly, bud and 5s stages are shown. Enlargements of merged images (d, h, l) show the areas indicated in corresponding lower magnification pictures (c, g, k). Insets show YZ projections from confocal images. Arrowheads indicate examples of ionocyte progenitors with colocalized Klf4 and dlc in the epidermal ionocyte domain. Arrows indicate ionocyte progenitors expressing only dlc. dlc+ ionocyte progenitor number at indicated embryonic stages is shown (m). Scale bar, 50 μm. Error bars indicate standard error.