Fig. 4.

Simultaneous disruption of NLS1 and NLS2 abolishes the nuclear localization of LRH-1. a Basic amino acids within either or both NLSs were mutated to alanines in the full-length LRH-1 tagged with Myc. These constructs were transfected into COS-7 cells. After 24 h, cells were fixed and immunostained with anti-Myc antibody. The nuclei were counterstained with DAPI. Fusion proteins were visualized by confocal microscopy. Wild-type (a) and single cluster mutants in NLS1 (b, c) or NLS2 (d, e) are exclusively localized to the nucleus. The subcellular localization of double cluster mutants in representative cells is shown in f–i. Scale bar 25 μm. b Statistics of subcellular distribution of wild-type (a in a) and double cluster mutants (f–i in a) are summarized. A minimum of 100 cells were counted for each experiment and the means ± SEM three independent experiments are presented