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Severe acute respiratory syndrome‐related coronavirus 2 (SARS‐CoV‐2) primarily targets the respiratory system. Physiologically relevant human lung models are indispensable to investigate virus‐induced host response and disease pathogenesis. In this study, we generated human induced pluripotent stem cell (iPSC)‐derived alveolar organoids (AOs) using an established protocol that recapitulates the sequential steps of in vivo lung development. AOs express alveolar epithelial type II cell protein markers including pro‐surfactant protein C and ATP binding cassette subfamily A member 3. Compared to primary human alveolar type II cells, AOs expressed higher mRNA levels of SARS‐CoV‐2 entry factors, angiotensin‐converting enzyme 2 (ACE2), asialoglycoprotein receptor 1 (ASGR1) and basigin (CD147). Considering the localization of ACE2 on the apical side in AOs, we used three AO models, apical‐in, sheared and apical‐out for SARS‐CoV‐2 infection. All three models of AOs were robustly infected with the SARS‐CoV‐2 irrespective of ACE2 accessibility. Antibody blocking experiment revealed that ASGR1 was the main receptor for SARS‐CoV2 entry from the basolateral in apical‐in AOs. AOs supported the replication of SARS‐CoV‐2 variants WA1, Alpha, Beta, Delta, and Zeta and Omicron to a variable degree with WA1 being the highest and Omicron being the least. Transcriptomic profiling of infected AOs revealed the induction of inflammatory and interferon‐related pathways with NF‐κB signaling being the predominant host response. In summary, iPSC‐derived AOs can serve as excellent human lung models to investigate infection of SARS‐CoV‐2 variants and host responses from both apical and basolateral sides. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) regulate gene expression posttranscriptionally and are implicated in viral replication and host tropism. miRNAs can impact the viruses either by directly interacting with the viral genome or modulating host factors. Although many miRNAs have predicted binding sites in the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) viral RNA genome, little experimental validation has been done. We first identified 492 miRNAs that have binding site(s) on the spike (S) viral RNA by a bioinformatics prediction. We then validated the selected 39 miRNAs by examining S-protein levels after coexpressing the S-protein and a miRNA into the cells. Seven miRNAs were found to reduce the S-protein levels by more than 50%. Among them, miR-15a, miR-153, miR-298, miR-508, miR-1909, and miR-3130 also significantly reduced SARS-CoV-2 viral replication. SARS-CoV-2 infection decreased the expression levels of miR-298, miR-497, miR-508, miR-1909, and miR-3130, but had no significant effects on miR-15a and miR-153 levels. Intriguingly, the targeting sequences of these miRNAs on the S viral RNA showed sequence conservation among the variants of concern. Our results suggest that these miRNAs elicit effective antiviral defense against SARS-CoV-2 by modulating S-protein expression and are likely targeting all the variants. Thus, the data signify the therapeutic potential of miRNA-based therapy for SARS-CoV-2 infections. NEW & NOTEWORTHY MicroRNAs can impact viruses either by directly interacting with the virus genome or by modulating host factors. We identified that cellular miRNAs regulate effective antiviral defense against SARS-CoV-2 via modulating spike protein expression, which may offer a potential candidate for antiviral therapy.