Abrogation of N-glycosylation in MICA018 results in impaired cell surface expression.
A, JTag9 cells were transfected with an empty plasmid (pUC18) or plasmids encoding GFP-Myc-tagged MICA018 fusion proteins (MICA018 WT and MICA018 5× N-mut). Gating was carried out on viable and GFP-positive cells (gate 2), and the grid was set according to pUC18 transfected cells. Left panel shows representative results from five independent experiments presented as dot plots. The upper row shows GFP intensity, and the lower row shows Myc surface staining (percentages inserted upper right). The right panels show representative Western blots of transfected cells detecting GFP and ERK. B, JTag9 cells were transfected with plasmids encoding indicated GFP-Myc-tagged MICA018 fusion proteins. Gating was carried out on viable and GFP-positive cells, and the grid was set according to pUC18 transfected cells. Representative results from three independent experiments are presented as dot plots showing Myc surface staining (percentages inserted upper right). FSC, forward-side scatter.