Fig. 6

Aldh2 meets the demand of McSCs for purines. (A) Purine nucleotides rescue Aldh2-deficient melanocyte regeneration. Melanocyte regeneration assay in mitfa^vc7 embryos with or without CVT-10216 plus purine nucleotide cocktail. Melanocyte counts normalized to respective untreated controls. Each dot represents a single embryo, three experimental replicates. Data are mean±s.e.m. **P<0.0021, ***P<0.0002, ****P<0.0001; ns, not significant (one-way ANOVA with Tukey's multiple comparisons). (B) Purine, but not pyrimidine nucleosides, rescues Aldh2-deficient melanocyte regeneration. Melanocyte regeneration assay on mitfa^vc7 embryos with or without CVT-10216 and supplemented with deoxyadenosine (dA), deoxguanosine (dG) or deoxyuridine (dU) or thymidine (T) nucleosides (200 µM). Each datapoint represents a single embryo, three experimental replicates. ****P<0.0001; ns, not significant (one-way ANOVA with Tukey's multiple comparisons). Data are mean±s.e.m. (C) Purine nucleotides rescue McSC differentiation in ALDH2i-treated embryos. Representative confocal z-stacks of Tg(mitfa:GFP;crestin:mCherry) embryos treated with MoTP with or without CVT-12016, as well as 400 µM AMP/GMP purine nucleotides, or 400 µM UMP/thymidine pyrimidine nucleotides. Two experimental replicates, more than five embryos per condition. (D) Quantification of crestin:mCherry and mitfa:GFP niche areas from C. Each dot represents the sum of the GFP or mCherry niche area/ number of somites in view in one embryo. ****P<0.0001; ns, not significant (one-way ANOVA with Tukey's multiple comparisons). Data are mean±s.e.m. (E) Proposed model for Aldh2-mediated control of the McSC lineage. Regenerating McSCs start expressing crestin and low levels of mitfa. Next, McSCs increase their metabolic demands for purine nucleotides to express high levels of mitfa and generate progeny. This metabolic demand is met by Aldh2 metabolizing endogenous formaldehyde into formate, which is then used in the 1C cycle to fuel production of purine nucleotides. McSCs undergo cell division to generate progeny, which migrate away from the niche to the epidermis. ALDH2i (CVT-10216) delays the progression of the activated McSCs to generate progeny in regeneration.