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Changes in cell metabolism and plasma membrane potential have been linked to shifts between tissue growth and differentiation, and to developmental patterning. How such changes mediate these effects is poorly understood. Here, we use the developing wing of Drosophila to investigate the interplay between cell metabolism and a key developmental regulator-the Hedgehog (Hh) signalling pathway. We show that reducing glycolysis both lowers steady-state levels of ATP and stabilizes Smoothened (Smo), the 7-pass transmembrane protein that transduces the Hh signal. As a result, the transcription factor Cubitus interruptus accumulates in its full-length, transcription activating form. We show that glycolysis is required to maintain the plasma membrane potential and that plasma membrane depolarization blocks cellular uptake of N-acylethanolamides-lipoprotein-borne Hh pathway inhibitors required for Smo destabilization. Similarly, pharmacological inhibition of glycolysis in mammalian cells induces ciliary translocation of Smo-a key step in pathway activation-in the absence of Hh. Thus, changes in cell metabolism alter Hh signalling through their effects on plasma membrane potential., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

At a cellular level, nutrients are sensed by the mechanistic Target of Rapamycin (mTOR). The response of cells to hypoxia is regulated via action of the oxygen sensor Hypoxia-Inducible Factor 1 (HIF-1). During development, injury and disease, tissues might face conditions of both low nutrient supply and low oxygen, yet it is not clear how cells adapt to both nutrient restriction and hypoxia, or how mTOR and HIF-1 interact in such conditions. Here we explore this question in vivo with respect to cell proliferation using the ciliary marginal zone (CMZ) of Xenopus . We found that both nutrient-deprivation and hypoxia cause retinal progenitors to decrease their proliferation, yet when nutrient-deprived progenitors are exposed to hypoxia there is an unexpected rise in cell proliferation. This increase, mediated by HIF-1 signalling, is dependent on glutaminolysis and reactivation of the mTOR pathway. We discuss how these findings in non-transformed tissue may also shed light on the ability of cancer cells in poorly vascularised solid tumours to proliferate.

Hedgehog ligands control tissue development and homeostasis by alleviating repression of Smoothened, a seven-pass transmembrane protein. The Hedgehog receptor, Patched, is thought to regulate the availability of small lipophilic Smoothened repressors whose identity is unknown. Lipoproteins contain lipids required to repress Smoothened signaling in vivo. Here, using biochemical fractionation and lipid mass spectrometry, we identify these repressors as endocannabinoids. Endocannabinoids circulate in human and Drosophila lipoproteins and act directly on Smoothened at physiological concentrations to repress signaling in Drosophila and mammalian assays. Phytocannabinoids are also potent Smo inhibitors. These findings link organismal metabolism to local Hedgehog signaling and suggest previously unsuspected mechanisms for the physiological activities of cannabinoids.

Hedgehog (Hh) is a lipoprotein-borne ligand that regulates both patterning and proliferation in a wide variety of vertebrate and invertebrate tissues. When Hh is absent, its receptor Patched (Ptc) represses Smoothened (Smo) signaling by an unknown catalytic mechanism that correlates with reduced Smo levels on the basolateral membrane. Ptc contains a sterol-sensing domain and is similar to the Niemann-Pick type C-1 protein, suggesting that Ptc might regulate lipid trafficking to repress Smo. However, no endogenous lipid regulators of Smo have yet been identified, nor has it ever been shown that Ptc actually controls lipid trafficking. Here, we show that Drosophila Ptc recruits internalized lipoproteins to Ptc-positive endosomes and that its sterol-sensing domain regulates trafficking of both lipids and Smo from this compartment. Ptc utilizes lipids derived from lipoproteins to destabilize Smo on the basolateral membrane. We propose that Ptc normally regulates Smo degradation by changing the lipid composition of endosomes through which Smo passes, and that the presence of Hh on lipoproteins inhibits utilization of their lipids by Ptc.