Fig. 2. RPB3 interacts with HCR. (A) AH109 yeast cells were co-transformed with the indicated constructs and plated onto SD medium lacking leucine and tryptophan (-LW) to verify the expression of both bait (W⁺) and prey (L⁺) plasmids, or onto medium lacking leucine, tryptophan, histidine and adenine (-LWHA) to examine the interaction between bait and prey proteins. (B) Y187 yeast cells were co-transformed with the indicated constructs and assayed for ß-galactosidase activity. (C) In-vitro-translated and S³⁵-labelled HCR was subjected to GST pull-down analysis using GST or GST-RPB3 beads. (D) Whole-cell extracts of NIH 3T3 cells transfected with both Flag-HCR and Myc-RPB3, or with Flag-HCR and empty Myc-tag vectors (Myc-Vector) were immunoprecipitated with anti-Flag monoclonal antibody and analysed by western blot using an anti-Myc monoclonal antibody.