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. 2005 Jun 20;169(6):885–896. doi: 10.1083/jcb.200409150

Figure 5.

Figure 5.

Forced cytoplasmic localization of Gβ2 attenuates the suppressive effect of the wild-type Gβ on GR-induced transactivation, whereas forced nuclear localization enhances it. (A and B) EGFP-fused NES-Gβ2 and NLS-Gβ2 are exclusively localized in the cytoplasm and the nucleus, respectively, in HCT116 cells. HCT116 cells were transfected with pEGFP-C1-NES-Gβ2- or pEGFP-C1-NLS-Gβ2-expressing plasmid. The cells were fixed and their confocal images were obtained. Representative images are shown in A, whereas mean ± SEM values of signal intensities in the nucleus (black bars) and the cytoplasm (white bars) obtained from over 20 cells are shown in B. (C) NES-Gβ2 loses the suppressive effect on GR transactivation, whereas NLS-Gβ2 has a stronger inhibitory effect than the wild-type Gβ2 on GR transactivation in HCT116 cells. HCT116 cells were transfected with pCDNA4His/MaxB-NES-Gβ2- or -NLS-Gβ2 together with pRShGRα, pMMTV-Luc, and pSV40-β-Gal. Bars represent mean ± SEM values of the luciferase activity normalized for β-galactosidase activity in the absence or presence of 10−6 M of dexamethasone. (*), P < 0.01; n.s., not significant, compared with the baseline. (D) Gβ2 wild-type and its fusions with NES or NLS are similarly expressed in HCT116 cells. HCT116 cells were transfected with pCDNA4His/MaxB-Gβ2, -NES-Gβ2-, or -NLS-Gβ2. The cells were lysed and the expression of wild-type, NES-, and NLS-fused Gβ2 was examined in a Western blots using anti-His antibody.