Figure 2.

Co-immunoprecipitation of DDIT3 and CDK2. (a) HT1080 cells were co-transfected with the indicated GFP constructs and CDK2-DsRed1. Proteins in cell lysates were further immunoprecipitated with anti GFP serum and recombinant CDK2-DsRed1 protein was detected by western blot using CDK2 antibodies. Input shows proteins before immunoprecipitation of cell lysates while IP shows proteins present in immunoprecipitates. (b) HT1080 cells were transiently transfected with the indicated GFP constructs and cellular proteins were immunoprecipitated with CDK2 antibodies. Recombinant proteins in samples were detected with anti GFP serum. The most intense bands in the input show the correct size of the GFP-tagged protein products while additional unspecific bands were also reacting with the GFP antisera in these samples. (c) ClustalW alignment of DDIT3 and C/EBPα amino acid sequences. A region shown to bind CDK2 in C/EBPα is boxed. Letters indicate standard amino acid abbreviations. Amino acids with related physicochemical characteristics are displayed below the sequences as: '*' (identical residue), ':' (highly similar residue), '.' (weakly similar residue). The leucine zipper region is seen towards the C-terminal in both proteins.