Investigating cellular transcriptomic alterations caused by VEEV replicase and SARS-CoV-2 mRNA co-transfection

The viral RNA-dependent RNA polymerase (replicase) from Venezuelan equine encephalitis virus constitutes a vital component of the bipartite trans-amplifying mRNA vaccine. In this vaccine strategy aimed at targeting SARS-CoV-2, the replicase mRNA is administered alongside the mRNA encoding the SARS-CoV-2 spike protein. Our investigation sought to determine whether the replicase induces amplification of cellular mRNAs. To this end, cells were transfected with mRNAs encoding the replicase and SARS-CoV-2 spike protein, while control groups received transfections of mRNAs encoding an unrelated protein along with the SARS-CoV-2 spike. We observed no significant upregulation of genes in the treatment group compared to the control group. This suggests that the replicase does not induce off-target amplification of cellular mRNAs. There were two conditions tested in this experiment: In Condition 1, A549 cells underwent treatment involving transfection with mRNAs encoding the VEEV replicase and SARS-CoV-2 spike proteins. In Condition 2, serving as the control, A549 cells were transfected with mRNAs encoding an unrelated mRNA, Xef, along with the SARS-CoV-2 spike protein. Subsequent analysis focused on comparing the cellular transcriptomes between these two conditions at 4 hours and 12hours post-transfection.