Inactivation of the Niemann Pick C1 cholesterol transporter 1 (NPC1) restricts SARS-CoV-2 infection

The Niemann Pick C1 (NPC1) protein is an intracellular cholesterol transporter located in the late endosome/lysosome (LE/Ly) and is involved in cholesterol mobilization. Loss-of-function mutations of the NPC1 gene lead to accumulation of cholesterol and sphingolipids in LE/Ly, resulting in severe fatal NPC1 disease. Cellular alterations associated with NPC1 inactivation affect both the integrity of lipid rafts and the endocytic pathway. Because the angiotensin-converting enzyme 2 (ACE2) and type 2 serine transmembrane protease (TMPRSS2) of the SARS-CoV-2 Spike (S) protein also localize to lipid rafts, we sought to investigate the hypothesis that NPC1 inactivation would generate an intrinsically unfavorable barrier to SARS-CoV-2 entry. In this study, we demonstrate that NPC1 pharmacological inactivation or CRISP/R-Cas mediated ablation of NPC1 dramatically reduced SARS-CoV-2 infectivity. More specifically, our findings demonstrate that pharmacological inactivation of NPC1 results in massive accumulation of ACE2 in the autophagosomal/lysosomal compartment. A >40-fold decrease in virus titer indicates that this effectively prevents VSV-Spike-GFP infection by impeding virus binding and entry. A similarly marked decrease in viral infectivity is observed in cells that had NPC1 expression genetically abrogated. These observations were further confirmed in a de novo SARS-CoV-2 infection paradigm, where cells were infected with the naturally pathogenic SARS-CoV-2. Overall, this work offers strong evidence that NPC1 function is essential for successful SARS-CoV-2 infection, thus implicating NPC1 as a potential therapeutic target in COVID-19 management. Significance A significant convergence exists between the cellular alterations associated with NPC1 inactivation and the mechanistic processes of SARS-CoV-2 infectivity. These alterations affect the integrity of lipid-enriched plasma membrane microdomains and the endocytic pathway. Furthermore, the cholesterol-regulated ACE2 receptor protein that facilitates SARS-Cov-2 viral binding and entry is targeted to the autophagolysosomal compartment upon NPC1 inactivation, thus hindering virus-host cell interaction. To our knowledge, this study provides the first evidence that NPC1 function represents a crucial factor for SARS-CoV-2 infection and suggests therapeutic opportunities.