Fig. 8

(A-B) GO analysis for biological pathways was performed using DAVID. Numbers in parentheses indicate number of genes enriched in each GO term. P-value cutoff = 0.001. In situ hybridization of (C-E) medium-wave sensitive (opn1mw1) and (F-H) short-wave sensitive opsin (opn1sw1). Transcripts of both genes are only detected in a few cells of the ventral retina in tet2^-/-;tet3^-/- embryos (arrows in E,H; n>8). natriuretic peptide a (nppa) is normally expressed in the heart (I), and is not detected in the retina of wild-type embryos (J; n>8). (K) In tet2^-/-;tet3^-/- embryos, ectopic nppa expressing cells were detected throughout the retina and brain (arrows) of all embryos examined (n = 16/16). (L) Bisulfite sequencing did not identify any changes in DNA methylation in any of the sixteen RNA-seq-identified target loci examined (S4 Table), including the nppa gene body. Bisulfite reads covering part of the first intron and second exon of nppa gene body are shown as black (methylated) or white (unmethylated). (M) Site-specific 5hmC quantification detected a significant, 20-fold reduction in 5hmC levels in the nppa gene body of 72hpf tet2^-/-;tet3^-/- embryonic eye tissue, when compared to phenotypically wild-type siblings (p = 0.0038; two-tailed, unpaired t-test).