Entry of the Ã?±-coronavirus porcine epidemic diarrhea virus (PEDV) requires specific proteases to activate spike (S)\nprotein for the membrane fusion of the virion to the host cell following receptor binding. Herein, PEDV isolate 85-7\ncould proliferate and induce cellââ?¬â??cell fusion in a trypsin independent manner on Vero cells, and eight homologous\nmutation strains were screened by continuous proliferation in the absence of trypsin on Vero cells. According to the\nwhole genome sequence comparative analysis, we identified four major variations located in nonstructural protein 2,\nS, open reading frame 3, and envelope (E) genes, respectively. Comparative analyses of their genomic variations and\nproliferation characteristics identified a single mutation within the S2ââ?¬Â² cleavage site between C30 and C40 mutants:\nthe substitution of conserved arginine (R) by a glycine (G) (R895G). This change resulted in weaker cellââ?¬â??cell fusion,\nsmaller plaque morphology, higher virus titer and serious microfilament condensation. Further analysis confirmed\nthat this mutation was responsible for optimal cell-adaptation, but not the determinant for trypsin-dependent entry\nof PEDV. Otherwise, a novel variation (16ââ?¬â??20 aa deletion and an L25P mutation) in the transmembrane domain of the\nE protein affected multiple infection processes, including up-regulation of the production of the ER stress indicator\nGRP78, improving the expression of pro-inflammatory cytokines IL-6 and IL-8, and promoting apoptosis. The results of\nthis study provide a better understanding of the potential mechanisms of viral functional proteins in PEDV replication,\ninfection, and fitness.
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