MicroRNA-Mediated regulation of the immune response in Calu-3 Cells Infected with a SARS-CoV-2 E Gene Variant

Chenfan Sun¹Fang Xu¹Zhongji Pu²Han-Ping Zhu¹Yudong Li³Hangjing Lu¹Beibei Wu¹Yisheng Sun^1*Pingping Yao^1*Jianmin Jiang^1*

• ¹Zhejiang Center for Disease Control and Prevention (Zhejiang CDC), Hangzhou, China
• ²Xianghu Laboratory, Hangzhou, China
• ³Zhejiang Gongshang University, Hangzhou, China

SARS-CoV-2, the pathogen of COVID-19, disrupts the alveolar epithelial barrier and triggers exacerbation of airway inflammation. The envelope (E) protein plays a key role in promoting epithelial damage and sustaining inflammation. We previously identified a SARS-CoV-2 variant (F8) containing a 12-bp deletion in the E gene. Compared to the 8X strain, which possesses the wild-type E gene, F8 could induce a higher expression of inflammatory factors in Calu-3 cells. In this study, we analyzed miRNAs expression profiles in F8- and 8X-infected Calu-3 cells. Through RT-qPCR and functional verification, we discovered that F8 infection significantly upregulated miR-361-3p and downregulated let-7b-5p. Furthermore, miR-361-3p regulated the PI3K-Akt pathway by directly targeting and inhibiting TSPAN1, ultimately promoting PTEN expression. The downregulation of let-7b-5p might release the suppression on inflammatory cytokines (IL-6, IL-8, PTX3), and partially disorder ZO-1 expression in maintaining the barrier function. This study is the first to demonstrate that the SARS-CoV-2 E protein mutant F8 remodeled the host miRNAs network to coordinate the immune responses and barrier function. All findings provided valuable insights into the pathogenesis of SARS-CoV-2 variants.