CGRP inhibits SARS-CoV-2 infection of bronchial epithelial cells, and its pulmonary levels correlate with viral clearance in critical COVID-19 patients.

Upon infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), patients with critical coronavirus disease 2019 (COVID-19) present with life-threatening respiratory distress, pulmonary damage, and cytokine storm. One unexplored component in COVID-19 is the neuropeptide calcitonin gene-related peptide (CGRP), which is highly abundant in the airways and could converge in multiple aspects of COVID-19-related pulmonary pathophysiology. Whether CGRP affects SARS-CoV-2 infection directly remains elusive. We show that in critical COVID-19 patients, CGRP is increased in both plasma and lungs. Importantly, CGRP pulmonary levels are elevated in early SARS-CoV-2-positive patients and restored to baseline upon subsequent viral clearance in SARS-CoV-2-negative patients. We further show that CGRP and its stable analog SAX directly inhibit infection of bronchial Calu-3 epithelial cells with SARS-CoV-2 Omicron and Alpha variants in a dose-dependent manner. Both pre- and post-infection treatments with CGRP and/or SAX are enough to block SARS-CoV-2 productive infection of Calu-3 cells. CGRP-mediated inhibition occurs via activation of the CGRP receptor and involves down-regulation of both SARS-CoV-2 entry receptors at the surface of Calu-3 cells. Together, we propose that increased pulmonary CGRP mediates beneficial viral clearance in critical COVID-19 patients by directly inhibiting SARS-CoV-2 propagation. Hence, CGRP-based interventions could be harnessed for management of COVID-19.IMPORTANCEThe neuropeptide CGRP is highly abundant in the airways. Due to its immunomodulatory, vasodilatory, and anti-viral functions, CGRP could affect multiple aspects of COVID-19-related pulmonary pathophysiology. Yet, the interplay between CGRP and SARS-CoV-2 during COVID-19 remains elusive. Herein, we show that pulmonary levels of CGRP are increased in critical COVID-19 patients, at an early stage of their disease when patients are SARS-CoV-2-positive. Upon subsequent viral clearance, CGRP levels are restored to baseline in SARS-CoV-2-negative patients. We further show that pre- and post-infection treatments with CGRP directly inhibit infection of Calu-3 bronchial epithelial cells with SARS -CoV-2, via activation of the CGRP receptor leading to decreased expression of both SARS-CoV-2 entry receptors. Together, we propose that increased pulmonary CGRP is beneficial in COVID-19, as CGRP-mediated inhibition of SARS-CoV-2 infection could contribute to viral clearance in critical COVID-19 patients. Accordingly, CGRP-based formulations could be useful for COVID-19 management.

Short-term exposure to cigarette smoke upregulates cathepsin S and alters expression of tight junction ZO-1.

A long-term exposure to cigarette smoke (CS) alters the integrity of airway epithelial barrier, contributes to lung dysfunction, and elicits the expression and activity of lung cathepsin S (CatS), a cysteine protease that participates in the remodeling of connective tissue and cell junctions. Here, we observed that a short-term (4 days) exposure of mice to CS increased the expression and activity of CatS, while the expression level of zonula occludens 1 (ZO-1), an epithelial tight junction protein that stabilizes barrier assembly, was reduced in lung tissue lysates. Present data support that proteolytically active CatS may contribute to the defect of ZO-1 in CS-exposed mice.

Proteomic analysis of SARS-CoV-2 particles unveils a key role of G3BP proteins in viral assembly.

Considerable progress has been made in understanding the molecular host-virus battlefield during SARS-CoV-2 infection. Nevertheless, the assembly and egress of newly formed virions are less understood. To identify host proteins involved in viral morphogenesis, we characterize the proteome of SARS-CoV-2 virions produced from A549-ACE2 and Calu-3 cells, isolated via ultracentrifugation on sucrose cushion or by ACE-2 affinity capture. Bioinformatic analysis unveils 92 SARS-CoV-2 virion-associated host factors, providing a valuable resource to better understand the molecular environment of virion production. We reveal that G3BP1 and G3BP2 (G3BP1/2), two major stress granule nucleators, are embedded within virions and unexpectedly favor virion production. Furthermore, we show that G3BP1/2 participate in the formation of cytoplasmic membrane vesicles, that are likely virion assembly sites, consistent with a proviral role of G3BP1/2 in SARS-CoV-2 dissemination. Altogether, these findings provide new insights into host factors required for SARS-CoV-2 assembly with potential implications for future therapeutic targeting.