FIGURE 6.

Rcn2, Caf20, and Gga1 physically interact with Slt2. Western blot analysis of the in vitro co-purification assays. The strain YMF3 (SLT2::6MYC) was transformed with plasmids expressing Rcn2, Caf20, or Gga1 fused to GST under control of the GAL1 promoter. Transformants were grown for 2 h in the presence of galactose and then Congo red for a final 30 μg/ml was added (+) or not (−), and cultures were incubated for a four additional hours. Cell extracts (input) were incubated with glutathione-Sepharose beads to purify GST-complexes (pull-down), and immunodetection was performed with anti-Myc and anti-GST antibodies that recognize Slt2-Myc and GST-fused proteins as indicated. Similar results were obtained in three different experiments, and selected images correspond to representative blots. Numbers below the anti-Myc blot indicate the amount of Slt2-Myc pulled down by GST-Rcn2, GST-Caf20, and GST-Gga1 normalized with respect to the amount pulled down by GST, both in the absence (−) or presence (+) of Congo red.