Fig. 6 . The interaction between Pex10p and Pex4p depends on the presence of Pex22p. (A) PEX5/PEX22 cells, (B) pex5/PEX22 cells and (C) PEX5/pex22 cells containing Pex10-C_ub-RUra3p and expressing the N_ub constructs of Pex4p, Pex10p, Pex13p, Pex15p, Pex19p, Tom22p, Guk1p and the empty plasmid were diluted to an OD₆₀₀ of 1, and 4 µl of this and of tenfold serial dilutions were spotted onto media containing 100 µM CuSO₄ and 5-FOA. The media lacked tryptophan and histidine to select for the plasmids. Cells were incubated at 30°C for 48 hours. (D) as (C) but cells contained Pex4-C_ub-RUra3p and expressed N_ub-Pex4p or the empty vector in wild-type or pex22 cells. (E) The Pex4p-Pex4p interaction is specific. Cells co-expressing Pex4-C_ub-RUra3p and the N_ub-constructs of Pex4p, Ubc6p, Guk1p or the empty vector were analyzed as in (A-C). (F) Yeast cells co-expressing Pex4-6ha together with either Pex4-9myc or Pex11-9myc were subjected to anti-ha precipitation after DSP crosslinking. The co-precipitated proteins were detected with anti-myc antibody after reverting the cross-link, 12.5% SDS-PAGE, and transfer onto nitrocellulose. The blot was stripped and re-probed with anti-ha-antibody to ensure that similar quantities of Pex4-6ha had been loaded (boxed).