Role of NPC1 in Regulating Store-Operated Calcium Entry: Lessons from Niemann Pick Type C Disease

Abstract

Niemann-Pick Type C is an autosomal recessive neurodegenerative disease characterized by mutations in the gene encoding NPC1, a protein critical for the transport of cholesterol to the endoplasmic reticulum (ER) and other cellular membranes. Mutations in NPC1 result in accumulation of cholesterol and other lipids in endo-lysosomes of cells. It has previously been established that non-physiological depletion of cholesterol can alter the activity of the molecular components comprising store-operated calcium entry (SOCE). This event is choreographed by interaction of ER membrane protein STIM1 and plasma membrane Orai channels to facilitate the movement of calcium ions across the plasma membrane in response to depletion of ER calcium. Despite this relationship, little is known regarding the mechanism by which mammalian cells can regulate cholesterol to control SOCE and its subsequent downstream signaling pathways. Thus, we sought to examine SOCE in the context of this neurodegenerative disorder whose principal characteristic is an alteration of cholesterol homeostasis. Using a combination of dynamic confocal and super resolution microscopies, and proteomics we have discovered that SOCE-mediated calcium flux is enhanced in Niemann Pick Type C disease. Moreover, there are larger conducting STIM1/Orai1 puncta at steady state and enhanced formation of these puncta following depletion of ER calcium. We also determine that a downstream consequence of these dynamics is enhanced translocation of the nuclear transcription factor, NFAT. Pharmacological inhibition of NPC1 recapitulates this SOCE phenotype in mouse hippocampal neurons. These neurons also exhibit increased basal calcium signaling and enhanced excitability, a consequence of NPC1 inhibition. Thus, a protein of the lysosome, NPC1, has the ability to tune SOCE through its ability to transfer cholesterol at ER-lysosome membrane contact sites, facilitating gene and protein expressional changes which may contribute to the disease phenotype.