Edited AZI has a high binding affinity with DDX1 in the nucleus. (A) Immunofluorescence of FLAG-tagged AZI-TC or AZI-G proteins in 32D cells. Nonedited AZI protein (AZI-TC) was accumulated in the cytoplasm, whereas edited AZI protein (AZI-G) was predominantly localized in the nucleus. (B) Schematic of experimental procedure. Briefly, 32D cells were transduced with nonedited (AZI-TC) or edited (AZI-G) overexpression lentivirus vectors. The transduced cells were then sorted for IP and mass spectrometry (MS). (C) IP of AZI from 32D cells transduced with nonedited (AZI-TC) or edited (AZI-G) lentivirus vectors. (D) The differential AZI interacting proteins identified by mass spectrometry in 32D cells transduced with the nonedited (AZI-TC) or edited (AZI-G) constructs. (E) Western blot analysis of AZI-FLAG co-immunoprecipitated (Co-IP). (F) The relative enrichment of DDX1 (referring to Figurepanel E) normalized to IP efficiency. (G) Proximity of AZI and DDX1, determined by immunofluorescence. (H) Proximity of AZI and DDX1, determined by proximity ligation assay. (I) The ratio of cytoplasmic and nuclear subcellular localization of AZI (edited and nonedited) with DDX1. (J) The binding affinities between AZI (edited and nonedited) and DDX1 in the nucleus and cytoplasm, as determined by western blot analysis after AZI-FLAG IP. ***P < .001; ****P < .0001; Error bars represent SD. DAPI, 4′,6-diamidino-2-phenylindole; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NS, not significant.