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Fig. 2 | Molecular Medicine

Fig. 2

From: Regulation of Transcription Functions of the p53 Tumor Suppressor by the mdm-2 Oncogene

Fig. 2

Expression and cellular localization of mdm-2 mutants

(A) Expression of mdm-2 mutants in SAOS-2 cells and in vivo interaction with p53. SAOS-2 cells were transfected with mdm-2 expression plasmids and selected with G418. Pooled drug resistant colonies were then transiently transfected with pC53-C1N3, which expresses wild-type human p53. The cells were labeled with 35S-methionine and analyzed by immunoprecipitation using the 4B2 or 4B11 monoclonal antibodies against mdm-2. The two doublet bands of varying intensity in each lane, at or below the 43 kD marker, are background bands nonspecifically binding to the antibodies, as shown by the vector transfected cells. Mdm-2 mutants able to bind p53 in vivo include Δ440–491, Δ340–491, Δ222–325, and Δ222–437. Mutant Δ90–150 showed weak interaction with p53. (B) Cellular localization of mdm-2 mutants. SAOS-2 cells were transfected with mdm-2 expression plasmids and selected with G418. Pooled G418-resistant cells were stained with anti-mdm-2 monoclonal antibody 2A10 by immunofluorescence. Shown here is a representative nuclear-localized mutant Δ440–491 (I) and a cytoplasmic mutant Δ150–230 (II).

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