BN Conference 2025

Loss-of-function mutations of Niemann Pick C1 intracellular cholesterol transporter (NPC1) cause a rare genetic condition (NPC1 disease, NPCD) in which cholesterol (Chol) and sphingolipids are entrapped in late endosomes/lysosomes and lipid raft integrity is disrupted. Sonic hedgehog (Shh) signaling, which is essential for the development of neurons and glia as well as the formation of synapses, is one of the signaling pathways impacted by this Chol dyshomeostasis. In fact, cholesteroylation and its interaction with the Patched receptor to control the Chol pool required for the activation of Smoothened and downstream effectors at the primary cilium are part of Shh activation. Interestingly, lysosomal Chol accumulation also causes mTORC1 to become hyperactive. This has been demonstrated to affect Shh signaling in cells lacking Tsc2 by mislocalizing Smoothened and lowering the concentration of Gli2 in primary cilia. Methods: Immunofluorescence was employed for imaging and protein quantification was performed using Western blots analysis. Results: Npc1 knockout (KO) cell models display hallmarks of impaired autophagy, evidenced by elevated levels of p62 and LC3II. These cells also exhibit significantly shorter primary cilia. Remarkably, treatment with myriocin has been shown to counteract these cellular defects, restoring both normal autophagy markers and primary cilia length. Additionally, myriocin successfully normalizes the decreased Gli1 levels observed in Npc1 KO cells. Conclusions: Given the established role of p62 in modulating mTORC1, our findings point to a complicated interaction between myriocin, cholesterol homeostasis, autophagic flux, and mTORC1 signaling in the context of NPCD. They also suggest that myriocin therapy may be used to restore Shh signaling and correct ciliary defects.