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. 2009 Dec 4;285(7):4909–4919. doi: 10.1074/jbc.M109.042341

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FIGURE 7. — ATM is required for the escape from the degradation of DYRK2. A, 293T cells ectopically expressed with FLAG-DYRK2 were transfected with scramble siRNA or ATM siRNA and then left untreated or treated with ADR for 24 h in the presence of MG-132. Lysates were immunoprecipitated with anti-FLAG-agarose. Immunoprecipitates (IP) were subjected to immunoblot analysis (IB) with anti-MDM2 (top panel) or anti-FLAG (second panel). Lysates were also subjected to immunoblot analysis with anti-ATM (third panel) or anti-tubulin (bottom panel). A ratio of MDM2 bound to DYRK2 was determined by the densitometric analysis using ImageJ program. The expression in unstressed cells transfected with scramble siRNA was defined as 1.00. B, 293T cells were transfected with FLAG-DYRK2 or the AQ mutants and then treated with ADR for 24 h in the presence of MG-132. Nuclear lysates were immunoprecipitated with anti-FLAG-agarose. Immunoprecipitates were subjected to immunoblot analysis with anti-ubiquitin (top panel), anti-MDM2 (second panel), or anti-FLAG (third panel). Lysates were also analyzed by immunoblotting with anti-MDM2 (fourth panel) or anti-lamin B1 (bottom panel).