{"doi":"10.1128/mbio.02287-23","title":"Calpain-2 mediates SARS-CoV-2 entry via regulating ACE2 levels","abstract":"ABSTRACT Since the beginning of the coronavirus disease 2019 (COVID-19) pandemic, much effort has been dedicated to identifying effective antivirals against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A number of calpain inhibitors show excellent antiviral activities against SARS-CoV-2 by targeting the viral main protease (M pro ), which plays an essential role in processing viral polyproteins. In this study, we found that calpain inhibitors potently inhibited the infection of a chimeric vesicular stomatitis virus (VSV) encoding the SARS-CoV-2 spike protein but not M pro . In contrast, calpain inhibitors did not exhibit antiviral activities toward the wild-type VSV with its native glycoprotein. Genetic knockout of calpain-2 by CRISPR/Cas9 conferred resistance of the host cells to the chimeric VSV-SARS-CoV-2 virus and a clinical isolate of wild-type SARS-CoV-2. Mechanistically, calpain-2 facilitates SARS-CoV-2 spike protein-mediated cell attachment by positively regulating the cell surface levels of ACE2. These results highlight an M pro -independent pathway targeted by calpain inhibitors for efficient viral inhibition. We also identify calpain-2 as a novel host factor and a potential therapeutic target responsible for SARS-CoV-2 infection at the entry step. IMPORTANCE Many efforts in small-molecule screens have been made to counter SARS-CoV-2 infection by targeting the viral main protease, the major element that processes viral proteins after translation. Here, we discovered that calpain inhibitors further block SARS-CoV-2 infection in a main protease-independent manner. We identified the host cysteine protease calpain-2 as an important positive regulator of the cell surface levels of SARS-CoV-2 cellular receptor ACE2 and, thus, a facilitator of viral infection. By either pharmacological inhibition or genetic knockout of calpain-2, the SARS-CoV-2 binding to host cells is blocked and viral infection is decreased. Our findings highlight a novel mechanism of ACE2 regulation, which presents a potential new therapeutic target. Since calpain inhibitors also potently interfere with the viral main protease, our data also provide a mechanistic understanding of the potential use of calpain inhibitors as dual inhibitors (entry and replication) in the clinical setting of COVID-19 diseases. Our findings bring mechanistic insights into the cellular process of SARS-CoV-2 entry and offer a novel explanation to the mechanism of activities of calpain inhibitors.","journal":"mBio","year":2024,"id":455331,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":6,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9602,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":622888,"name":"Avan Antia","orcid":"0000-0002-5092-6173","position":1,"is_corresponding":false},{"id":909020,"name":"Luis Alberto Casorla-Perez","orcid":"0000-0002-4643-2525","position":2,"is_corresponding":false},{"id":324219,"name":"Maritza Puray‐Chavez","orcid":"0000-0002-7983-0757","position":3,"is_corresponding":false},{"id":324222,"name":"Sebla B. Kutluay","orcid":"0000-0001-5549-7032","position":4,"is_corresponding":false},{"id":106969,"name":"Matthew A. Ciorba","orcid":"0000-0002-7656-0982","position":5,"is_corresponding":false},{"id":106970,"name":"Siyuan Ding","orcid":"0000-0002-5338-260X","position":6,"is_corresponding":false},{"id":227335,"name":"Qiru Zeng","orcid":"0000-0002-3731-9950","position":0,"is_corresponding":true}],"reference_count":37,"raw_metadata":null,"created_at":"2026-07-19T02:03:17.458329Z","pmid":"38349185","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}