GAS5 binds to eIF4E through RNA binding motifs.
(A) RNA binding motifs, which are italic and bold, in the eIF4E protein were predicted with 2 web-based tools, BindN and PPRInt. The motif, W56, W102 and E103 for m7G binding is bold and underlined. N-terminally located sequence is motif-1 and the C-terminal one is motif-2. (B-C) GAS5 RNA was detected by RT-PCR after RNA-IP assay using HA antibody in the cells transfected with (B) RNA binding deletion mutants (HA-eIF4EDel1, HA-eIF4EDel2 and HA-eIF4EDel1&2) and (C) cap binding mutant (HA-eIF4Ecap mutant). GAPDH was used as a control.