(A) Linear and circRNA stimulate expression of specific genes. Left: scatterplot of RNA-seq result of HeLa cells transfected with linear or circRNA encoding GFP-IRES sequence. Genes induced by at least 2-fold compared to mock control are plotted, and the color scale represents the enrichment of circRNA compared to linear RNA. Genes shown in bold were validated by qRT-PCR below. Right: Gene Ontology enrichment of genes induced by circular or linear RNA. Heatmap GO enrichment p value is shown.
(B) CircRNA stimulates expression of innate immune genes. HeLa cells were mock transfected or transfected with 500 ng of linear or circRNA encoding GFP-IRES. Relative expression of the indicated mRNA and transfected RNA are measured by qRT-PCR. Means ± SEM are shown (n = 3), *p < 0.05, Student’s t test, throughout.
(C) CircRNA transfection protects against viral infection. FACS analysis plots of HeLa cells transfected with 500 ng of Cy3-labeled linear or circRNA and exposed to VEEV-GFP virus.
(D) Percent of all HeLa cells infected with VEEV-GFP following transfection with linear or circRNA.
(E) Percent of HeLa cells that did not contain Cy3-labeled linear or circRNA infected with VEEV-GFP following transfection with linear or circRNA.