Glycolytic flux controls retinal progenitor cell differentiation via regulating Wnt signaling

Metabolic pathways are remodeled in response to energy and other homeostatic demands and are dynamically regulated during embryonic development, suggestive of a role in guiding cellular differentiation. Here, we show that glycolytic flux is required and sufficient to bias multipotent retinal progenitor cells (RPCs) to acquire a rod photoreceptor fate in the murine retina. In an RPC-specific conditional knock-out ofPhosphatase and tensin homolog(Pten-cKO) and in an RPC-specific conditional gain-of-function of dominant activePFKB3(cytoPFKB3), glycolytic gene expression and activity are elevated, correlating with precocious rod photoreceptor differentiation and outer segment maturation. Conversely, glycolytic inhibition in retinal explants, achieved either with 2-deoxy-d-glucose, a competitive inhibitor of glucose metabolism, by lowering media pH, which disables PKM2, a rate-limiting enzyme, or by inhibiting lactate/H⁺symporters, which lowers intracellular pH, suppresses RPC proliferation and photoreceptor differentiation. Mechanistically, we show that Wnt signaling, the top-upregulated pathway inPten-cKO retinas, is a glycolysis-dependent pathway. Pharmacological and genetic perturbation of Wnt signaling using aCtnnb1-cKO phenocopies glycolytic inhibition, suppressing RPC proliferation, photoreceptor differentiation and outer segment maturation. Thus, developmental rewiring of glycolytic flux modulates Wnt signaling to drive rod photoreceptor differentiation and maturation, an instructive role that may be exploited therapeutically for cell replacement strategies.