Fig. 4. Mutation of leucine 62 to alanine impairs nuclear export of Cdc25B. (A) The amino acids 54 to 67 of Cdc25B, which constitutes NES2. The three leucines are indicated in bold and the residue subjected to site-directed mutagenesis is indicated above. (B) Localisation of YFP-Cdc25B3 or YFP-Cdc25B3(L62A) 30 hours after transfection in G0 hTERT immortalised human fibroblasts. (C) Localisation patterns of YFP-Cdc25B3, YFP-Cdc25B3 L62A, YFP-Cdc25B3 S323G and YFP-Cdc25B3 NLS- in G2 HeLa cells. Cells were synchronised by a double thymidine-block, transfected immediately after release and fixed after 8 hours. Cells with interphase nuclei were scored for nuclear, both nuclear and cytoplasmic, or cytoplasmic YFP fluorescence.