Fig. 1.

DNMT3a mediates developmental plasticity. We used CRISPR/Cas9 to produce single (DNMT3aa^−/−) and double (DNMT3aa^−/−ab^−/−) knock-out lines plus a no-template control (control) (a). In each genotype, we tested the effect of developmental and acclimation temperatures (23 °C blue, 28 °C red) on fecundity, survival, and deformities in F₁ and F₂ fish. We used adult F₂ fish to determine locomotor and metabolic thermal performance curves. The points in the experiment where data in the different figures were collected are marked on top of the panel. At 23 °C developmental temperature (b), survival to the 26-somite stage (early survival) was lowest in the DNMT3aa^−/−ab^x/x (ab mixed genotype) fish (purple circles; p < 0.0001 compared to control), and survival was somewhat reduced in the DNMT3aa^−/− line compared to control at 28 °C (p < 0.04). Deformities (c) increased from control (grey circles) to DNMT3aa^−/− (green circles; p = 0.004) and DNMT3aa^−/−ab^x/x fish (p < 0.0001) at 23 °C developmental temperature and were elevated in DNMT3aa^−/−ab^x/x fish at 28 °C compared to control (p = 0.002). Survival to swim bladder inflation (late survival) (d) at 23 °C was lowest in DNMT3aa^−/−ab^x/x fish (p < 0.0001 compared to control) and intermediate in DNMT3aa^−/− fish (p = 0.02 compared to control). Late survival was reduced in the two knock-out lines at 28 °C (both p < 0.01 compared to control). Means ± s.e. are shown in B-D (black horizontal and vertical lines, respectively), and each symbol represents data from one clutch (n = 19–28 clutches per treatment). Letters indicate significant differences between groups. There were no homozygous knock-out survivors (e) at the ab locus in the F₁ DNMT3aa^−/−ab^x/x line at 23 °C developmental temperature, but 35% of survivors were DNMT3aa^−/−ab^−/− homozygous fish at 28 °C