JCS01108 Supplemental Figures

Files in this Data Supplement:

• Figure S1 -

  Fig. S1. Aurora-A phosphorylates only CDC25B. Recombinant MBP-CDC25A, B and C were incubated in vitro in a kinase assay buffer in the presence of Aurora-A kinase. Reaction was analysed by western blotting using SE96 antibody (top panel) or MBP antibody (middle panel) as a loading control. Lower panel: Quantification of ³²P incorporation from the assay performed in the presence of [g-³²P]ATP.

• Figure S2 -

  Fig. S2. Competition experiments for the SE96 labelling. Upper panel: Staining with SE96 antibodies in the presence or in the absence of phosphorylated peptide (QNKRRRS(p)VTPPEEQ; P-Peptide), unphosphorylated peptide (QNKRRRSVTPPEEQ; Peptide) or an irrelevant phosphorylated peptide (MEVEELS(p)PLALGR; Irrelevant P-Peptide); SE96 (red) and DAPI (blue). Lower panel: staining with SE96 in the presence of MBP-CDC25B phosphorylated or not by recombinant Aurora-A (as described in the legend to Fig. 1).

• Figure S3 -

  Fig. S3. Double labelling S353-phosphorylated CDC25B and a-tubulin. HeLa cells were fixed and double immunofluorescence staining was performed with SE96 and an a-tubulin monoclonal antibody (Sigma); SE96 (red), a-tubulin (green) and DAPI (blue).