Expression level of EGFP-tagged AID mutant proteins correlates with the specific cell compartment in which they reside. (A) Schematic representation of mutations affecting either the NLS (mutNLS, V18S, R19V, according to Shinkura et al. [13], with numbers indicating amino acid positions within the AID protein) or the NES (mutNES, L189A, F193A, and L196A), with both domains being represented as black boxes. (B) FACS analysis of the tetracycline-inducible clones, with values of the MFI of the EGFP-positive population. By comparison with endogenous AID expression levels in BL2 (corresponding to a MFI value of 15 in the KI clone [Fig. 1 C]), AID overexpression in EGFP-positive cells can be estimated at ∼4 for mutNES-AID, 24 for WT-AID, and 90 for mutNLS-AID constructs. (C) Confocal images of EGFP signals from inducible clones, with or without 2-h LMB treatment, are shown together with nuclear staining with propidium iodide (PI) of the same field. Clones with a lower percentage of inducible expression are shown, as indicated by the presence of EGFP-negative cells in the field, a shut-off of expression regularly observed in clones kept in culture in induced conditions. Bars, 20 μm.