Interactions between RNAP subunits, mRNA capping enzyme, and putative helicase of the yeast VLEs.

(A) Western blot of immunoprecipitated (α-GFP IP) and mock immunoprecipitated (mock IP) proteins from IFO1267_pRKL2-5 (yEGFP3-K2ORF6p, K2ORF7p-FLAG) or control IFO1267_pRKL2-15 cells (K2ORF7p-FLAG), respectively. The strains used are indicated above the lanes. The antibodies used for Western blot are indicated on the left hand side of the strips. The proteins detected are indicated on the right hand side of the strips. Positions of the identified proteins corresponded with theoretical molecular weight of the full length recombinant proteins, as determined by positions of the protein mass markers. Input represented approximately 1/100 of the sample and IP represented approximately 1/2 of the sample in this and the other immunoprecipitation experiments. Mock immunoprecipitations in all experiments were done using empty agarose beads. The same experimental scheme is used throughout this figure. (B) Western blot analysis of immunoprecipitations from lysates from IFO1267_pRKL2-7 (yEGFP3-K2ORF6p, HA-K2ORF4p) and control IFO1267_pRKL2-13 (HA-K2ORF4p) cells (indicated above the lanes). The (α-GFP) and anti-HA (α-HA) antibodies used are indicated on the left hand side; the detected proteins on the right hand side. (C) Western blot analysis of immunoprecipitations from lysates from strains IFO1267_pRKL2-6 (yEGFP3-K2ORF6p, K2ORF3p-HA), IFO1267_pRKL2-10 (yEGFP3-K2ORF4p, K2ORF3p-HA), and IFO1267_pRKL2-14 (control). (D) Western blot analysis of immunoprecipitations from lysates from IFO1267_pRKL2-9 cells (yEGFP3-K2ORF4p, HA-K2ORF6p). (E) Western blot analysis of immunoprecipitations from lysates from IFO1267_pRKL1-4/2-4 cells (yEGFP3-K2ORF6p, K1ORF4p-HA).