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. Author manuscript; available in PMC: 2015 Dec 1.
Published in final edited form as: Arch Biochem Biophys. 2014 Aug 7;563:136–144. doi: 10.1016/j.abb.2014.07.026

Figure 6. MAGE-A3 Is Not Ubiquitinated During KRAB-KAP1 Interactions.

Figure 6

HEK293T cells were transiently transfected with Flag tagged MAGE-A3 or empty vectors and KRAB domain GAL4-DBD constructs, KAP1, and HA tagged ubiquitin. Proteasome activity was inhibited by incubation for 5 hours in the presence of 25 µM MG132. Top panel, whole lysate immunoblot (IB) for KAP1; middle panel, Flag-MAGE-A3 tagged ubiquitinated proteins were immunoprecipitated with anti-Flag and detected with anti-HA; Lower panel: immunoprecipitation and immunoblotting confirmed expression of Flag-MAGE-A3. No ubiquitinated MAGE-A3 species were seen in blots of cells transfected with both KZNFs and MAGE-A3, panels A to F (A=ZNF382, B=ZNF136, C =ZNF45, D=MZF1, E=ZNF287 and F=ZNF274).