The trypsin-enhanced infection of porcine epidemic diarrhea virus is determined by the S2 subunit of the spike glycoprotein

Yubei Tan; Limeng Sun; Gang Wang; Yuejun Shi; Wanyu Dong; Yanan Fu; Zhen Fu; Huanchun Chen; Guiqing Peng

doi:10.1128/JVI.02453-20

The trypsin-enhanced infection of porcine epidemic diarrhea virus is determined by the S2 subunit of the spike glycoprotein

J Virol. 2021 May 10;95(11):e02453-20. doi: 10.1128/JVI.02453-20. Epub 2021 Mar 10.

Authors

Yubei Tan^{1

2}, Limeng Sun^{1

2}, Gang Wang^{1

2}, Yuejun Shi^{1

2}, Wanyu Dong³, Yanan Fu^{1

2}, Zhen Fu^{1

2}, Huanchun Chen^{1

2}, Guiqing Peng^{4

2}

Affiliations

¹ State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China.
² Key Laboratory of Preventive Veterinary Medicine in Hubei Province, The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, 430070, China.
³ Department of Veterinary Medicine, College of Animal Science and Technology, Zhejiang A&F University, Hangzhou, 311300, China.
⁴ State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China penggq@mail.hzau.edu.cn.

Abstract

Porcine epidemic diarrhea virus (PEDV) is an enteric pathogen in the swine industry, causing high mortality in neonatal piglets. Efficient PEDV infection usually relies on the presence of trypsin, yet the mechanism of trypsin dependency is ambiguous. Here, we identified two PEDV strains, trypsin-enhanced YN200 and trypsin-independent DR13, in which the spike (S) protein of YN200 exhibits a stronger ability to induce syncytium formation and cleaved by trypsin than that of DR13. Using a full-length infectious YN200 cDNA clone, we confirmed that the S protein is a trypsin dependency determinant by comparison of rYN200 and rYN200-S_DR13 To explore the trypsin-associated sites of the YN200 S protein, we then constructed a series of mutations adjacent to the fusion peptide. The results show that the putative S2' cleavage site (R892G) is not the determinant for virus trypsin dependency. Hence, we generated viruses carrying chimeric S proteins: the S1 subunit, S2 subunit, and S2_{720∼892 aa} domain (NS2') were individually replaced by the corresponding DR13 sequences. Intriguingly, only the S2 substitution, not the S1 or NS2' substitutions, provides trypsin-independent growth of YN200. Additionally, the NS2' recombinant virus significantly abrogated effective infection, indicating a vital role for NS2' in viral entry. These findings suggest that the trypsin dependency of PEDV is mainly controlled by mutations in the S2 subunit rather than directly trypsin cleavage site.ImportanceWith the emergence of new variants, PEDV remains a major problem in the global swine industry. Efficient PEDV infection usually requires trypsin, while the mechanism of trypsin dependency is complex. Here, we used two PEDV strains, trypsin-enhanced YN200 and trypsin-independent DR13, and results showed that the S protein determined PEDV trypsin dependency by using a reverse genetic system of YN200. The S2 subunit was verified as the main portion of PEDV trypsin dependency, though the putative S2' site mutation cannot render trypsin-independent growth of YN200. Finally, these results provide some different insight to the PEDV trypsin dependency and might inspire vaccine development.