TMPRSS2 activation of Omicron lineage Spike glycoprotein is regulated by Collectrin-like domain of ACE2

Abstract

Continued high levels spread of SARS-CoV-2 globally enabled accumulation of changes within the Spike glycoprotein, leading to resistance to neutralising antibodies and concomitant changes to entry requirements that increased viral transmission fitness. Herein, we demonstrate a significant change in ACE2 and TMPRSS2 use by primary SARS-CoV-2 isolates that occurred upon arrival of Omicron lineages. Mechanistically we show this shift to be a function of two distinct ACE2 pools based on cleavage or non-cleavage of ACE2 by TMPRSS2 activity. In engineered cells overexpressing ACE2 and TMPRSS2, ACE2 was cleaved by TMPRSS2 and this led to either augmentation or progressive attenuation of pre-Omicron and Omicron lineages, respectfully. In contrast, TMPRSS2 resistant ACE2 restored infectivity across all Omicron lineages through enabling ACE2 binding that facilitated TMPRSS2 spike activation. Therefore, our data support the tropism shift of Omicron lineages to be a function of evolution towards the use of uncleaved pools of ACE2 with the latter consistent with its role as a chaperone for many tissue specific amino acid transport proteins.

Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86/JN.1 lineages combining increased fitness and antibody evasion

“…This protease cleaves and primes the spike for fusion, but also cleaves ACE2 (Heurich et al ., 2014). Initial SARS-CoV-2 variants used both cleaved and uncleaved ACE2 as a receptor, whereas a shift towards preferential use of uncleaved ACE2 for Omicron variants has been proposed (Aggarwal et al, 2023). Our results strongly suggest that a delicate balance between ACE2 and TMPRSS2 levels is necessary for optimal replication of recent variants in Vero E6 cells.…”

Neutralisation of SARS-CoV-2 Omicron subvariants BA.2.86 and EG.5.1 by antibodies induced by earlier infection or vaccination

“…SARS-CoV-2 variants exhibit variable affinities for the angiotensin-converting enzyme 2 (ACE2) host receptor and dominance in using either the fusion or the endocytic entry pathway in immortalised cell lines (6,7). To account for this, we attempted viral isolation on different cell lines permissive to SARS-CoV-2 infection with respiratory sample material obtained from individuals with an RT-PCR- and sequencing-confirmed BA.2.86 infection.…”

Rapid spread of the SARS-CoV-2 JN.1 lineage is associated with increased neutralization evasion

“…Vero (African green monkey kidney, female, kidney; CRL-1586, ATCC; RRID: CVCL 0574, kindly provided by Andrea Maisner), 293T (human, female, kidney; ACC-635, DSMZ; RRID: CVCL 0063), Vero E6 cells stably expressing TMPRSS2 (Vero-TMPRSS2; JCRB1819, CellBank Australia; RRID: CVCL_YQ49), and Vero E6 cells stably expressing ACE2 and TMPRSS2 (Vero-ACE2+TMPRSS2) 15 were cultivated using Dulbecco’s modified Eagle medium (DMEM, PAN-Biotech), supplemented with 10% fetal bovine serum (Biochrom), 100 U/ml of penicillin, and 0.1 mg/mL of streptomycin (PAN-Biotech). Calu-3 cells (human, male, lung; HTB-55, ATCC; RRID: CVCL_0609, kindly provided by Stephan Ludwig) were cultivated using DMEM/F-12 medium (GIBCO), supplemented with 10% fetal bovine serum (Biochrom), 100 U/ml penicillin and 0.1 mg/mL streptomycin (PAN-Biotech), 1x non-essential amino acid solution (from 100x stock, PAN-Biotech) and 1 mM sodium pyruvate (PAN-Biotech).…”