Fig. 6. Pygo protein forms a complex with Arm in vivo and contains a transactivation domain(s). (A) Co-immunoprecipitation of Pygo and Arm. 293T cells were transfected with plasmids expressing either Myc-tagged Pygo (amino acid 105 to 815) or HA-tagged Arm (amino acid 128 to 844) alone and together as indicated. Whole-cell lysates were prepared 36 hours after transfection. Lysates were immunoprecipitated with mouse monoclonal anti-Myc antibody. Immunoprecipitated materials and a fraction of each lysate were resolved by SDS-PAGE and analyzed by western blotting with antibodies as shown. IP, immunoprecipitation; IB, immunoblot. (B) Analysis of the transcription activation domain(s) in Pygo. Cells were transfected with the pG5E1b-luciferase reporter construct (Hsu et al., 1994) and with vectors expressing GAL4 DNA-binding domain alone (pM1) (Sadowski et al., 1992) or with GAL4-Pygo fusion protein. A GAL4-Jun AC-containing Jun activation domain (amino acids 5 to 89) fused with GAL4 was used as a positive control. Luciferase acitivities are expressed as relative activities compared with cells transfected with the plasmid containing the GAL4 DNA-binding domain alone.

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