Description

Constituvely active mutant Arf1-GFP (Arf1(Q71L)-GFP) (gray) blocks redistribution of Helix Pomatia Lectin (HPL) (green) from Golgi to ER in Src-deficient SYF fibroblasts cotransfected with active Src (SrcE-DsRed) (red). HPL binds various glycans but the Tn antigen in particular. The Tn antigen refers to terminal α-linked N-acetyl galactosamine residues (GalNAc) linked to Ser or Thr residues. HPL indicates distribution of GalNac-Ts. Active Src is enriched at the Golgi upon expression of Arf1(Q71L)-GFP . Cells were fixed for 10 min (4% paraformaldehyde) and permeabilized (0.2% Triton X-100) prior to staining with Hoechst (blue) and Alexa 647-conjugated-HPL. Cells were mounted onto glass slides using FluorSave (Merck) and imaged at room temperature using an inverted FluoView confocal microscope (model IX81; Olympus) with either a 60x objective (U Plan Super Apochromatic, UPLSAPO; NA 1.35) or 100x objective (UPLSAPO; NA 1.40) using Immersol oil. Microscope coupled with a CCD camera (model FVII). Images were acquired and processed using Olympus FV10-ASW software. Image corresponds to Fig 7E in J Cell Biol. 189: 843-858. 2010. Images in Fig 7 include CIL#s 13571, 13572, 13573, 13574, 13575, 13576, 13577.