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. 2021 Jun 17;17(6):e1009644. doi: 10.1371/journal.ppat.1009644

Fig 4. Mechanistic analysis of UPR activation by SARS-CoV-2 proteins.

Fig 4

HEK-293T cells were transfected with plasmids encoding SARS-CoV-2 S (S-HA) or ORF8 (ORF8-FLAG), mock-transfected, or treated with tunicamycin (Tn). At 8 h p.t., cells were treated with 60 μM IREi and 100 μM AEBSF and then harvested at 24 and 36 h p.t. (A) Western blot analysis of ORF8-FLAG, S-HA, HERP, BiP, PERK, ATF4, p-eIF2α and ATF6 proteins. The specific p-eIF2α and ATF6-Nt bands are indicated with a red asterisk. Protein band quantifications for HERP, BiP, p-eIF2α and ATF6-Nt, normalised by eIF2α as a loading control and given relative to the mock, are provided below the respective immunoblots. (B) RT-PCR analysis of XBP1-u and XBP1-s mRNAs, performed as described in Fig 2D. Immunoblots and agarose gels are representative of three biological replicates. “h p.t.” = hours post-transfection.