Fig. 5

Mapping of the Gag interaction domains in RPL7. a Scheme of Flag–RPL7 and its mutants: Flag–RPL7 mutants used in the present study are presented with a grey rectangle (RPL7 sequence conserved in the constructs) and with a black line (RPL7 sequence deleted in the constructs). Letters correspond to each construct and are used in b. On the right, (+) and (−) report the ability of each construct to be co-precipitated with Gag-eGFP. b Identification of Flag–RPL7 interaction with Gag-eGFP by immunoprecipitation experiments. HeLa cells were either not transfected (lane 1), or co-transfected with plasmids expressing eGFP and Flag–RPL7 (a, lane 2), Gag-eGFP alone (lane 3) or Gag-eGFP combined with Flag–RPL7 A (lane 4) or Flag–RPL7 B (lane 5), Flag–RPL7 C (lane 6), Flag–RPL7 D (lane 7), Flag–RPL7 E (lane 8), Flag–RPL7 F (lane 9), Flag–RPL7 G (lane 10). IP was performed with 1 mg of total protein and with mouse anti-eGFP antibody. 20 µg of total protein (input) and resuspended beads were resolved by 10 % SDS-PAGE and analyzed by immunoblotting using antibody against Flag followed by anti-mouse HRP conjugate. Membranes were stripped and re-blotted using mouse anti-eGFP antibody followed by anti-mouse HRP conjugate. Lane 1′ corresponds to beads without antibody. Asterisk residual signal of the secondary anti-mouse interacting with primary anti-Flag antibody