XAP2 is not required for assembly of the AhR with hsp90 in vitro. (A) Immunoblot analysis of XAP2 in RL. Lanes: 1, XAP2 transcribed and translated in vitro in RL; 2, RL alone. Each reaction was resolved by SDS-PAGE and used for immunoblot analysis with XAP2 polyclonal antibodies. (B) Immunoabsorption of AhR-hsp90-XAP2 complex. Lanes: 1, pcDNA3/βmAhR-FLAG cotranscribed and translated in vitro with XAP2 in RL; 2, same mixture as in lane 1 incubated with M2 affinity gel blocked with FLAG peptide; 3, pcDNA3/βmAhR-FLAG transcribed and translated in vitro in reticulocyte lysate; 4, same mixture as in lane 3 incubated with M2 affinity gel blocked with FLAG peptide. Each translation mixture was subjected to immunoabsorption with the M2 affinity gel, washed, resolved by SDS-PAGE, and used in an immunoblot analysis with the antibodies raised against the AhR (RPT1), hsp90 (hsp86/84), and XAP2.