The inhibition of cell migration by IRF5 is non-transcriptional and occurs in the cytoplasm. A) Graphical summary of data from the dual luciferase assay measuring ISRE transactivation in the presence of full-length IRF5 or the indicated deletion mutants. Data are from 3 independent experiments performed in duplicate. B) Western blot analysis of cytoplasmic (C) and nuclear (N) extracts from 231 cells transiently transfected with the indicated plasmids. α-tubulin was used as a control for extract purity. C) Schematic representation of full-length IRF5 with a mutated 5′ NLS (mNLS). D) Graphical summary of data from the migration assay comparing EV migratory function to full-length IRF5 and the mNLS mutant in 231 cells. Data are from at least 3 independent experiments performed in duplicate; ***p ≤ 0.0001.