Figure 1.

Deletion of the Robo1 Ig1 domain prevents Slit–Robo1 interaction in cultured Drosophila cells. Drosophila S2R+ cells were transfected with the indicated HA-tagged UAS-Robo transgenes, incubated with Slit-conditioned media, and then stained with anti-HA (magenta) and anti-Slit (green) antibodies. Slit does not bind to mock-transfected cells that do not express transgenic Robo1 (A), but binds robustly to cells expressing a full-length Robo1 transgene (B). The level of Slit binding correlates with the level of Robo1 expression, as cells expressing lower levels of Robo1 also exhibit weaker Slit binding (arrowhead). Cells expressing transgenic Robo1^ΔIg1 exhibit similar levels of Slit binding to control cells (C). (A–C) Confocal max projections through the entire cells; (D–F) single confocal Z-slices through the cells indicated with arrows in (A–C). Robo1^ΔIg1 is properly localized at the plasma membrane, similar to Robo1 (compare HA in E and F), indicating that deletion of Ig1 does not disrupt expression of Robo1 at the cell surface. Schematics of the tested Robo receptor variants are shown at left.