Figure 2.

ATP11A coexpression reduces B1 signaling activity.A, B1 schematics: depiction of B1FL on left, shown with full NTF, and B1ΔNT on right, lacking NTF up to site of predicted GAIN domain cleavage. B, coexpression with B1FL in HEK293T cells resulted in 54% reduction in B1FL activation of SRF-luciferase (mean ± SEM shown, ordinary one-way ANOVA with Tukey’s multiple comparisons test, p < 0.0001, n = 13, ROUT method used at 10% to remove two outliers). “IB" refers to “immunoblot” to indicate what antibody was used to detect the protein bands shown via Western blot. C, ATP11A coexpression with B1ΔNT resulted in no significant change in receptor activation of SRF-luciferase (mean ± SEM is shown, ordinary one-way ANOVA with Tukey’s multiple comparisons test, n = 11, ROUT method used at 10% to remove one outlier). D, ATP11A coexpression with B1FL did not significantly alter total cell lysate expression levels of receptor. Representative Western blot shown on left with quantification on right (normalized mean ± SEM is shown, unpaired t test, n = 13). E, ATP11A coexpression with B1ΔNT did not significantly alter total cell lysate expression levels of receptor. Representative Western blot shown on left with quantification on right (normalized mean ± SEM is shown, unpaired t test, n = 3). F, ATP11A coexpression with B1FL did not significantly alter receptor surface expression. Representative Western blot is shown on left with quantification on right (normalized mean ± SEM is shown, unpaired t test, n = 5). B1FL, full-length B1; GAIN, G protein–coupled receptor autoproteolysis–inducing domain; HEK293T, human embryonic kidney 293T cell line; NTF, N-terminal fragment.