(A) Cartoon of the structure of SF-1 showing the position of the nonsense mutations reported here (p.E11X, p.Q107X) together with that of the previously reported p.Y138X change (18). (B) The p.E11X and p.Q107X mutants show impaired transcriptional activity in an assay using the minimal promoter of Cyp11a in tsa201 cells. The activity of the previously described p.G35E mutant is shown for comparison (19). Data represent the mean±s.e.m. of three independent experiments, each performed in triplicate. (−), empty vector; WT, wild-type; RLU, relative lights units. (C) Expression and cellular localization of WT and mutant GFP-SF-1 fusion proteins. WT GFP-SF-1 localizes in the nucleus with relative nucleolar exclusion. A truncated protein corresponding to the p.E11X nonsense mutant reported here is located throughout the cell, whilst low levels of cytoplasmic fusion protein are seen with p.Q107X. In contrast, the previously reported p.Y138X change is strongly localized to the nucleus with signal in the nucleolus too (18). These studies largely support reports that the nuclear localization signal of SF-1 requires codons 89–101 with bipartite basic motifs at either end of this region being particularly important (33).