Fig. 1

ADD1 was detected in the nucleus and cytoplasm, whereas ADD2 and ADD3 were detected only in the cytoplasm. a Schematic representation of the NLS motif in ADD1, ADD2, and ADD3. b HA-ADD1, HA-ADD2, and HA-ADD3 were transiently expressed in NIH3T3 cells. The cells were lysed, and the interaction of HA-ADD proteins with importin α was analyzed by co-immunoprecipitation. The HA-ADD proteins were immunoprecipitated (IP) with anti-HA antibody or pre-immune serum (IgG) as the control. The immunocomplexes were washed and analyzed by immunoblotting (IB) with anti-importin α or anti-HA. An equal amount of whole cell lysates (WCL) was analyzed by immunoblotting to verify the levels of importin α and HA-ADD proteins. c HA-ADD1, HA-ADD2, and HA-ADD3 and their respective mutants with the deletion of residues adjacent to the NLS were transiently expressed in NIH3T3 cells. The cells were fixed and stained for the nuclei and HA-ADD proteins. The representative images are shown. Scale bar, 10 μm. The distribution of the HA-ADD proteins in the nucleus and cytoplasm was measured (n > 800). The values (mean ± SD) are from three experiments. N > C, distributed mainly in the nucleus; N < C, distributed mainly in the cytoplasm; N = C, distributed equally in the nucleus and cytoplasm