Inactivated Wild-Type Human Coronavirus (229E)

To gain a deeper understanding of the signaling and molecular events that orchestrate the overall host response within the nucleus of coronavirus-infected cells, researchers investigated transcriptome dynamics in A549 and HuH7 cells infected with human coronavirus 229E (HCoV-229E), respectively, revealing a core signature of upregulated genes in these cells. Compared with treatment with the canonical inflammatory cytokine interleukin (IL)-1, HCoV-229E replication was found to attenuate the induction of transcription factor (TF) NF-κB and limit the nuclear concentration of NF-κB subunits through (i) an unusual mechanism involving partial degradation of IKKβ, NEMO, and IκBα, and (ii) upregulation of TNFAIP3 (A20), although constitutive IKK activity and basal TNFAIP3 expression levels have been shown to be required for efficient viral replication. The data showed that a large number of genes were activated in cells infected with HCoV (but not treated with IL-1), while the inducible p65 NF-κB was not recruited. Both HCoV-229E replication and IL-1 were shown to upregulate a small set of genes encoding immune regulators that bind p65 at their promoters and require IKKβ activity and p65 for expression. In addition, HCoV-229E and IL-1 activated a common set of 440 p65-binding enhancers that differed from an additional 992 HCoV-229E-specific enhancer regions by their combination of TF binding motifs. Taken together, this study demonstrates that cytoplasmic RNA viruses fine-tune NF-κB signaling at multiple levels and deeply reprogram host cell chromatin architecture to orchestrate the timely and coordinated expression of genes involved in multiple signaling, immunoregulatory and metabolic processes.

To systematically analyze the host cell gene response induced by HCoV-229E, the researchers first performed a series of whole transcriptome studies using the A549 lung epithelial cancer cell model. To detect transcriptional changes caused by viral replication, cells were infected with low multiplicity (replication-competent) HCoV-229E (MOI 0.001) or inoculated with replication-incompetent (heat- or UV-inactivated) HCoV-229E. At 16 and 48 hours post-infection (p.i.), increasing amounts of HCoV-229E RNA were detected in HCoV-229E-infected cells, but not in control cells inoculated with heat- or UV-inactivated HCoV-229E. Transcriptome analysis revealed that HCoV-229E infection resulted in differential expression of 108 genes at 16 hours post-infection and 239 genes at 48 hours post-infection (Figure 1A, top). An almost identical set of differentially expressed genes was also found in control cells inoculated with heat-inactivated virus (i.e., virus did not replicate) (Figure 1A, lower left and middle panels), suggesting that the observed differential gene expression was largely due to soluble factors present in the virus stock or interactions between cells and non-infectious virus particles. However, after 2 days of infection, a specific set of 26 up-regulated genes and 11 down-regulated genes were also identified that were differentially regulated only in cells infected with replication-competent HCoV-229E but not in cells inoculated with inactivated virus (Figure 1A, lower right panel).

Figure 1. A549 lung epithelial cells were infected with HCoV-229E and collected for further analyses at 16, 24 and 48 h p.i.. As controls, cells were mock-infected or incubated with heat-inactivated HCoV-229E and collected at the time points indicated above. (Poppe M, et al., 2017)