Identification of cellular proteins interacting with the tyrosine-phosphorylated ORF5. (Left) Glutathione-Sepharose beads containing GST (lane 1), GST-ORF5 (lane 2), or tyrosine-phosphorylated GST-ORF5 (GST-pORF5) (lane 3) were resolved by SDS-PAGE, followed by immunoblotting (IB) with an antiphosphotyrosine antibody (αpY). (Right) Glutathione-Sepharose beads containing GST (lane 1), GST-ORF5 (lane 2), or phosphorylated GST-ORF5 (GST-pORF5) (lane 3) were mixed with lysates of Jurkat T cells. ORF5 binding proteins were resolved by SDS-PAGE and visualized by Coomassie blue staining. Arrows indicate the PI3 kinase catalytic subunit (p110), PI3 kinase regulatory subunit (p85), SLP-76 (p75), Lck, and Fyn (p55). Numbers in parentheses indicate the frequency of peptides of each cellular protein obtained from mass spectrometry.