SARS-CoV-2 non-structural protein 6 triggers NLRP3-dependent pyroptosis by targeting ATP6AP1
Authors: Judeng Zeng
Tony Gin
Lina Yi
Gavin Matthew Joynt
Maggie Haitian Wang
Xiaodong Liu
Wei Hu
William Ka Kei Wu
Yuelong Shu
Huanle Luo
Ziheng Huang
Xuan Zou
Zhe Liu
Shisong Fang
Jun Yu
Jing Lu
Hung Chan
Huarong Chen
Matthew Tak Vai Chan
Yingzhi Liu
David Shu Cheong Hui
Francis Ka Leung Chan
Lin Zhang
Sunny Hei Wong
Siew Chien Ng
Christopher Hon Ki Cheng
Xiao Sun
Wenye Xu
Journal: Cell Death and Differentiation (2022)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: NSP6; SARS-CoV-2; COVID-19; inflammasome; pyroptosis
Affiliations: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China., Department of Anaesthesia and Intensive Care and Peter Hung Pain Research Institute, The Chinese University of Hong Kong, Hong Kong, China., Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong, China., Department of Gastroenterology, Shenzhen Hospital, Southern Medical University, Shenzhen, Guangdong, China., Guangdong Provincial Institution of Public Health and Guangdong Provincial Center for Disease Control and Prevention, Guangdong, China., CUHK Shenzhen Research Institute, Shenzhen, China., Shenzhen Center for Disease Control and Prevention, Shenzhen, Guangdong, China., School of Public Health (Shenzhen), Sun Yat-sen University, Shenzhen, Guangdong, China., Shenzhen Center for Disease Control and Prevention, Shenzhen, Guangdong, China. ., School of Public Health (Shenzhen), Sun Yat-sen University, Shenzhen, Guangdong, China. ., Guangdong Provincial Institution of Public Health and Guangdong Provincial Center for Disease Control and Prevention, Guangdong, China. ., Department of Anaesthesia and Intensive Care and Peter Hung Pain Research Institute, The Chinese University of Hong Kong, Hong Kong, China. .

Abstract:  A recent mutation analysis suggested that Non-Structural Protein 6 (NSP6) of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a key determinant of the viral pathogenicity. Here, by transcriptome analysis, we demonstrated that the inflammasome-related NOD-like receptor signaling was activated in SARS-CoV-2-infected lung epithelial cells and Coronavirus Disease 2019 (COVID-19) patients' lung tissues. The induction of inflammasomes/pyroptosis in patients with severe COVID-19 was confirmed by serological markers. Overexpression of NSP6 triggered NLRP3/ASC-dependent caspase-1 activation, interleukin-1Œ≤/18 maturation, and pyroptosis of lung epithelial cells. Upstream, NSP6 impaired lysosome acidification to inhibit autophagic flux, whose restoration by 1Œ±,25-dihydroxyvitamin D3, metformin or polydatin abrogated NSP6-induced pyroptosis. NSP6 directly interacted with ATP6AP1, a vacuolar ATPase proton pump component, and inhibited its cleavage-mediated activation. L37F NSP6 variant, which was associated with asymptomatic COVID-19, exhibited reduced binding to ATP6AP1 and weakened ability to impair lysosome acidification to induce pyroptosis. Consistently, infection of cultured lung epithelial cells with live SARS-CoV-2 resulted in autophagic flux stagnation, inflammasome activation, and pyroptosis. Overall, this work supports that NSP6 of SARS-CoV-2 could induce inflammatory cell death in lung epithelial cells, through which pharmacological rectification of autophagic flux might be therapeutically exploited. 

Retracted: No

PubPeer comments:
 Could the authors check figure 3E? Is it expected that these two images should appear so similar? ***  , 
 Similar question within Figure 5D. Should these images show the same cells? It seems they have purportedly been treated differently, if I understand correctly. Region of possible overlap outlined with magenta boxes, after rotation. ***  , 
 Another question for the authors. It seems that the authors are affiliated with multiple institutions (see below), yet the ethics statement in the paper reads very generically:  The study was approved by the institutional Ethics Committee. Could the authors provide greater detail?  , 
 I, as the corresponding author, will investigate these allegations with the first author and other co-authors. Thank you for bringing these up. , 
 #3 This work was done to support an ongoing public health response and data collection is part of the continuing public health investigation of an emerging outbreak and therefore the individual informed consent was waived (Lancet Infect Dis (2020) 20:911u20139. doi: 10.1016 s1473-3099(20)30287-5). The study was approved by the ethics committee of the Shenzhen Center for Disease Control and Prevention (CDC). , 
 #5 Thank you for addressing this question. , 
 #1 #2 Another senior member and I went through the raw data. We believe they are unintentional mistakes that were made during figure preparation. We will upload all the raw data to a pubic repository for readers to download (please give us some time; there are tens of folders, each of which contains up to hundreds of image files. We need time to annotate them properly). My apology. ,