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. 2007 Feb 21;81(9):4722–4731. doi: 10.1128/JVI.02548-06

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Copyright © 2007, American Society for Microbiology

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FIG. 5. — GSK-3 phosphorylation of C/EBPα and C/EBPβ is inhibited by LANA. (A) HeLa cells transfected with Flag-C/EBPα, HA-GSK-3β, and GFP-LANA, as indicated, were metabolically labeled for 4 h with [³²P]orthophosphate 2 days posttransfection. Flag-C/EBPα was immunoprecipitated with anti-Flag M2 affinity beads and subjected to Western blotting to detect Flag-C/EBPα (lower) and to autoradiography to detect ³²P-labeled C/EBPα (upper). Detection of phosphorylated Flag-C/EBPα required cotransfection with HA-GSK-3β (lane 3), and this phosphorylation was blocked by cotransfected LANA (lane 5). (B) HeLa cells transfected with Flag-C/EBPβ, HA-GSK-3β, and GFP-LANA, as indicated, were metabolically labeled with [³²P]orthophosphate as described above. Flag-C/EBPβ was immunoprecipitated with anti-Flag M2 affinity beads and subjected to Western blotting to detect Flag-C/EBPβ (lower) and to autoradiography to detect ³²P-labeled C/EBPβ (upper). ³²P labeling of Flag-C/EBPβ increased in the presence of HA-GSK-3β (lane 3), and this increase was abolished by cotransfection of LANA (lane 7). LiCl is a GSK-3 inhibitor. Circled p, phospho.