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Acta Veterinaria et Zootechnica Sinica ; 53(5):1587-1597, 2022.
Article Dans Chinois | CAB Abstracts | ID: covidwho-1994513


HEK293 cells were used as the cell model to investigate the role of human aminopeptidase N (hAPN) in the invasion of porcine deltacoronavirus (PDCoV) into human cells. The proliferation of PDCoV on HEK293 cells was firstly identified by RT-qPCR/RT-PCR. And then, hAPN knockout cell line was constructed by CRISPR/Cas9 technology and cell viability of HEK293 hAPN knockout and wild-type cells was verified by CCK-8 assay. Effect of hAPN knockout and overexpression on PDCoV replication was detected by RT-qPCR and Western blot. Meanwhile, interaction of PDCoV S protein and hAPN protein was analyzed by homology modeling and molecular docking. Results showed that PDCoV virus copies rapidly increased at 12-36 h and reached peak level at 36 h, it could propagate at least for passage 2 on HEK293 cells. There was no significant difference in cell viability between hAPN knockout cells and wild-type cells. Knockout of hAPN inhibit PDCoV replication and overexpression of hAPN enhance PDCoV replication. Homology modeling and molecular docking analysis showed S1 protein could bind hAPN domain II. Residues TYR92, THR51, THR48, PHE16 and MET14of S1 protein receptor binding motif 1 (RBM1) can form hydrogen bonds with residues PHE490, GLN531, ARG528 and SER529 of hAPN. This study indicates that hAPN plays a critical role in HEK293 cells during PDCoV infection, which provides new theoretical evidence for further studies on the mechanism of PDCoV entry into host cells and cross-species transmission.

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