Figure 6

Cellular localization of Abi1, HSPC300, and the Scar homology domain. Transfected Cos cells were stained with a monoclonal anti-Myc antibody and polyclonal anti-HA antibodies to detect over-expressed proteins. (A) Cos7 cells were co-transfected with (i) HA-HSPC300, (ii) HA-Abi1, (iii) Myc-Scar1 SHD, (iv) Myc-Scar2 SHD, or (v) Myc-Scar3 SHD to examine localization of these proteins. HSPC300, and all three SHDs localise in a diffuse cytoplasmic pool with some enrichment at protrusive edges of cells. Abi1 is not detected at the edges of cells, but appears in vesicle-like spots throughout the cytoplasm. (B) Cos7 cells were co-transfected with Myc-Scar1 SHD (i) and HA-HSPC300 (ii) shown as red and green respectively in a merged image (iii). Scar1-SHD and HSPC300 exhibit the same diffuse staining throughout the cytoplasm but also co-localize at protruding edges of cells. (C) Cos7 cells were co-transfected with Myc-Scar1 SHD (i) and HA-Abi1 (ii). (iii) Shows a merge image with myc-Scar1 SHD in red and HA-Abi1 in green. Scar1 SHD and Abi1 both colocalize to punctate spots similar to those seen for Abi1 alone. (D) Cos7 cells co-transfected with Myc-Scar2 SHD (i) and HA-HSPC300 (ii) shown as red and green respectively in a merge image (iii). Both Scar2 SHD and HSPC300 exhibit peri-nuclear staining and localization to protrusive edges of cells. (E) Cos7 cells co-transfected with Myc-Scar2 SHD (i) and HA-Abi1 (ii). (iii) Scar2 SHD (red) and Abi1 (green) show co-localization to cytoplasmic spots and the edges of cells. (F) Cos7 cells co-transfected with (i) Myc-Scar3 SHD (red) and (ii) HA-HSPC300 (green), in a merged image (iii). HSPC300 and Scar3 SHD show diffuse cytoplasmic staining with enrichment at protrusive edges of cells. (G) Cos7 cells co-transfected with (i) Myc-Scar3 SHD and (ii) HA-Abi1. (iii) In a merge image Abi1 (green) and Scar3 SHD (red) co-localize to punctate cytoplasmic spots and to the edges of lamellipodia. Scale bars in all panels are 20 μm.