SARS-CoV-2 infects cells following viral entry via clathrin-mediated endocytosis

Abstract

With more than 13 million cases and 570,000 deaths, and with the resulting social upheaval, the COVID-19 pandemic presents one of the greatest challenges ever to the scientific community. It is thus vital to fully understand the biology of SARS-CoV-2, the causative agent of COVID-19. SARS-CoV-2 uses the spike glycoprotein to interact with the cell surface and to drive fusion of the viral membrane with cellular membranes, thus allowing transfer of viral RNA to the cytosol. Here we use purified spike glycoprotein protein and lentivirus pseudotyped with spike glycoprotein to determine that SARS-CoV-2 undergoes rapid endocytosis following binding to the plasma membrane. Using chemical inhibitors and loss of function approaches, we demonstrate that this cellular entry is through clathrin-mediated endocytosis. Thus, it appears that SARS-CoV-2 first engages the plasma membrane, then rapidly enters the lumen of the endosomal system, strongly suggesting that fusion of the viral membrane occurs with the lumenal membrane of endosomes. This discovery has important implications for the development of chemical probes to reduce or block infection.

A review on possible mechanistic insights of Nitazoxanide for repurposing in COVID-19

“…Receptor mediated entry follows the clathrin mediated endocytic pathway which typically results in early endosome formation and maturation to the late endosome before the contents are released in the cytosol. Such cytosolic release is a crucial step in furthering viral pathogenesis ( Bayati et al, 2020 ; Glebov, 2020 ). Nitazoxanide can delay cytosolic release through its effect on multiple targets ( Rossignol, 2014 ).…”

COVID‐19: Current knowledge in clinical features, immunological responses, and vaccine development

“…Importantly, it has been shown that after engaging the cell membrane, either viral RNA enters the cytosol or the ACE2/SARS-CoV-2 complex is endocytosed with the entire virus and the viral membrane fuses with the luminal side of the endosome allowing for viral RNA transfer to the cytosol. 102 Various groups have demonstrated that SARS-CoV-2 is able to infect cell lines expressing ACE2 more effectively than those lacking ACE2 expression. 103,104 Another protein that plays a role in SARS-CoV-2 entry is the cellular serine protease, TMPRSS2, which primes the S protein and is essential for viral spread and pathogenesis.…”

Coronavirus entry: how we arrived at SARS-CoV-2

“…SARS-CoV-2 entry also fits well with the “early” and “late” pathway model proposed by Gallagher. Entry specifically via CME has been proposed as a route of internalization of SARS-CoV-2 in 293 T/ACE2 cells [ 55 ]; however, as discussed by the authors there are conflicting reports regarding specific endocytosis pathways for SARS-CoV and for coronaviruses in general, and so data need to interpreted cautiously. Another key set of findings comes from a CRISPR screen where RAB7A and genes involved in cholesterol biogenesis, among others, were identified as critical components of the SARS-CoV-2 entry pathway [ 56 ], indicating a key role for modified late endosomes—in this case using a human lung A549 cell line expressing ACE2.…”