FIGURE 3.

SNEV^Prp19-Pso4 physically and directly interacts with Blom7α. A, GST pulldown experiments using bacterially expressed His₆-tagged Blom7 and GST-SNEV^Prp19-Pso4. His₆-Blom7 is detected by Western blotting (IB) only when GST-SNEV^Prp19-Pso4 is added to the beads. B, in vitro transcription coupled translation of HA-tagged Blom7 was incubated with insect cell expressed His_6-SNEV^Prp19-Pso4. Precipitation was performed using Ni²⁺-NTA-agarose beads and is dependent on the presence of His_6-SNEV^Prp19-Pso4 as detected by anti-HA antibody. C, immunoprecipitation of SNEV^Prp19-Pso4 using beads covalently coupled to anti-Blom7 antibodies and detection on Western blots using anti-SNEV^Prp19-Pso4 antibody. Whereas rabbit IgG coupled beads do not precipitate SNEV^Prp19-Pso4, anti-Blom7 antibodies result in detection of SNEV^Prp19-Pso4. D, Western blot using 20 μg of HeLa cell lysate was separated by SDS-PAGE and detected using Blom7 antibody.