SARS-CoV-2 variant Alpha has a spike-dependent replication advantage over the ancestral B.1 strain in human cells with low ACE2 expression
Fig 3
VOC Alpha and B.1 share efficient prevention of induction of innate immunity despite VOC Alpha-specific enhanced production of viral subgenomic RNA transcripts.
Calu-3 cells were infected with B.1 or VOC Alpha (MOI of 2) and cell lysates were generated either 24 or 48 hours postinfection, following total RNA extraction. Samples were generated in 4 individual experiments each conducted in quadruplicates. (A) Expression of cell-associated envelope was determined in Calu-3 at 24 and 48 hours postinfection by Q-RT-PCR. TBP was used for normalization. (B) Expression of cell-associated sgN in Calu-3 cells at 24 and 48 hours postinfection was determined by Q-RT-PCR. TBP was used for normalization. (C) RNA-seq analysis was conducted from total cell lysates that were obtained 24 hours postinfection to quantify sgRNA proportions in infected cells. Stacked bars depict the relative proportion of total sgRNA in Calu-3 cells mapping to each viral sgRNA from the RNA-seq dataset. (D) Canonical, as well as ORF9b and N* sgRNAs were quantified from the RNA-seq dataset. Data were normalized to the respective B.1- or VOC Alpha-specific genomic reads. (E) Expression of the indicated genes was determined by Q-RT-PCR. Shown is the mean fold change +/− SD. RVFV cl.13, which is devoid of its IFN antagonist NSs, was included as an induction control for the expression of IFNs and ISGs. IFN, interferon; MOI, multiplicity of infection; Q-RT-PCR, quantitative real-time PCR; RVFV cl.13, Rift Valley Fever Virus clone 13; sgN, subgenomic nucleocapsid; sgRNA, subgenomic RNA; TBP, TATA-binding protein; VOC, variant of concern. See S1 Data.