Figure 1.
FynCA increases protein expression of Nav1.7 in HEK 293 cells stably expressing Nav1.7r channels (HEK-Nav1.7st cells). (a) Representative Western blot image of cells transfected with FynCA or FynDN with or without incubation of PP2 (a blocker of Src family kinases). The expression of Nav1.7 was detected using antisodium channel antibody (PanNav) and the sample loading variability was corrected by normalizing the intensity of Nav1.7 bands with that of GAPDH of the corresponding lane. FynCA increased the total protein expression of Nav1.7, which was almost completely eliminated by PP2. (b) The histograms of the Nav1.7 protein expression in cells expressing FynCA or FynDN, as well as the mean effect of PP2 on FynCA-induced upregulation of Nav1.7 channels. Data are presented as means ± SE, *p < 0.05 by one-way ANOVA or two-sample student t test. (c) Representative Western blot image of cell surface biotinylation experiment. The PanNav antibody revealed two distinguishable bands (labeled with filled and open arrowheads) from the biotinylated samples. This probably reflects the different glycosylation levels of Nav1.7 channels. FynCA elevated cell surface expression of Nav1.7, with the majority channels not fully glycosylated. (d) The histograms of total expression and surface expression of Nav1.7 in cells transfected with FynCA or FynDN. Na/K-ATPase and GAPDH were used to correct sample loading variability for biotinylated samples and total lysates, respectively. FynCA increased surface expression of Nav1.7 for ∼3.23 ± 0.80 folds (n = 6), which was relatively less than that of total Nav1.7 expression (7.80 ± 1.75 folds, n = 6). Data are presented as means ± SE, *p < 0.05 by two-sample student t test. GAPDH: glyceraldehyde phosphate dehydrogenase.