JCS01395 Supplemental Material

Files in this Data Supplement:

• Supplemental Figure 1 (Adobe PDF) -

  Fig. S1. Mutation of NES2 reduces the export rate of Cdc25B3 in G2. Fluorescence loss in photobleaching (FLIP) of YFP-Cdc25B3 and YFP-Cdc25B3(L62A) in G2 HeLa cells. Cells were synchronised with thymidine, released and microinjected with expression vectors five hours after release. Three hours later, the FLIP assay was performed as described for G1 cells. The nuclear fluorescence of the bleached cells was quantified. After background subtraction, the value of nuclear fluorescence at the start of the experiment was normalised to 1.

• Supplemental Figure 2 (Adobe PDF) -

  Fig. S2. Endogenous Cdc25B partly translocates to the cytoplasm after treatment with cycloheximide. When treating cells with cycloheximide for 1 hour, both the nuclear and cytoplasmic staining was significantly reduced indicating a short half-life of endogenous Cdc25B. Therefore, to be able to study the localisation pattern changes we added the proteasome inhibitor MG132 to the cells. HeLa cells were treated for two hours with or without MG132 (MG) and SB202190 (SB), and for 1 hour with or without cycloheximide (CHX) as indicated. The nuclear and cytoplasmic B1:5 (Cdc25B) fluorescence of Cyclin B1 positive cells from deconvolution images was quantified, the background was subtracted and the  cytoplasmic signal was divided with the nuclear signal.