Fig. 3: P. canaliculata regenerating eyes express genes driving vertebrate eye development.

A Schematic representation of the main morphologically defined time points during eye regeneration. Blastema/cell proliferation (yellow), lens (cyan), retina (red). B MDS plot of RNA-Seq samples collected during eye regeneration time course. Dimension 1 explains 41.8% of variance and Dimension 2 13.72%. n = 4 samples/time point (5 animals/sample) (see Supplementary Fig. 3A). C Number of differentially expressed (DE) genes (up-regulated in red and down-regulated in blue) compared to Intact samples [Log Fold Change (LogFC ≥ 0 and FDR ≤ 1e−5]. D GO enrichment analysis using significantly up-regulated genes (LogFC ≥ 0 and FDR ≤ 1e−5) in 1 dpa vs Intact and in 3, 6, 9, 12, 15, 21 and 28  dpa vs 1 dpa comparisons to highlight the features recovered throughout regeneration. GO terms used in this representation were manually selected from GO enrichment analysis run on the up-regulated genes obtained from comparing each time point with the previous one (p value ≤ 0.01, q value ≤ 0.05) (see Supplementary Fig. 4 and Supplementary Data 3B, C). E Heatmap of gene expression changes during eye regeneration time course. These genes represent the main stages of eye regeneration, and many of them are known, from previous studies, to be involved in vertebrate eye development (z scores calculated from TPMs). Gene categories on the right of the heatmap are based on GO annotations. F TEM images of the photoreceptor cytoplasm in the retina of Intact, 1, 2 and 3 months post amputation (mpa) eyes. Nuclei (yellow), photic vesicles (cyan), ribosomes and rough endoplasmic reticulum (empty arrowhead) and mitochondria (full arrowhead). The images are representative of data collected through three independent experiments. G SEM images of the retina, photoreceptor microvilli and apical cilia observed in Intact and 3 mpa eyes. At both time points, the retina is constituted by long photoreceptors (empty arrowhead), neuropile (on the bottom, full arrowhead) and microvilli (on the top). The photoreceptor microvilli at 3 mpa show an apical flat and wide expansion of the membrane like the Intact eyes. Cilia (full arrow) have been observed intercalated in the photoreceptor microvilli both in Intact and 3 mpa retina. The images are representative of data collected through three independent experiments.