Fig. 2
Knockdown of FgfA1 induces loss of the apical tuft. A In situ hybridization showing the expression of FgfA1 mRNA in the aboral ectoderm of wild type embryos (arrows) at/after 24hpf. B Live images of 48 hpf larvae injected with scrambled control shRNA (ctrl shRNA) or FgfA1 shRNA. C Images of fixed, 48 hpf larvae treated with 20 μm SU5402 or vehicle control (DMSO) and stained with DAPI (nuclei), phalloidin (F-actin), and anti-acetylated tubulin antibody (cilia). Arrows in B, C point to apical tuft cilia and dotted circles indicate loss of apical tuft. The oral pole is to the left in A–C; scale bars = 20 µm. D Quantitative analysis of apical tuft cilia length in the shRNA experiment (grey boxes) and pharmacological experiment (cyan boxes). Box plots are presented as: median—middle line, 25th and 75th percentiles—box, 5th and 95th percentiles—whiskers. Sample sizes for each treatment: wild type N = 10, ctrl shRNA N = 8, FgfA1 shRNA N = 10, DMSO N = 10, SU5402 N = 12. P-values from ANOVA with Tukey HSD post hoc: wild type vs. ctrl shRNA: p = 0.5839021, wild type vs. FgfA1 shRNA: p = 0.0000064, ctrl shRNA vs FgfA1 shRNA: p = 0.0000001, wild type vs DMSO: p = 0.0154365, DMSO vs SU5402: p = 0.0004114, FgfA1 shRNA vs. SU5402: p = 0.3147784. Letters indicate groups that are significantly different. E Cladogram of hard corals and sea anemones plotting the distribution of taxa with a larval apical tuft (cartoons, right). The apical tuft was likely lost in the ancestor of Scleractinia (black circle) and regained in the ancestor of the clade containing Astrangia and Oculina and at least one species of Caryophyllia (magenta circles). An Fgf signaling pathway controls apical tuft development in Astrangia poculata (this study) and Nematostella vectensis [48]. The cladogram was inferred from two studies of overlapping taxa [27, 38]. References indicating presence/absence of apical tuft by taxon: Pocillopora [60], Stylophora [1], Caryophyllia [61], Lophelia [30], Astrangia [58], Oculina [5], Acropora [22], Galaxea [1], Porites [51], Nematostella [21], other sea anemones: Anthopleura [7], Exaiptasia [6], Gonactinia [8]