Fig. 1

HCN1, HCN2, and HCN4 channels, but not the HCN3 channel, are S-palmitoylated in HEK cells. Representative blots showing S-palmitoylation of HCN2 and Kv1.5, but not Kir2.1 (a), and of HCN1, HCN2, and HCN4, but not HCN3 (b). FLAG-tagged channels were expressed in HEK cells, and were subjected to the ABE assay. Upper panels show biotin-labeled proteins probed with streptavidin-HRP. Lower panels show total proteins probed with anti-FLAG antibody and HRP-conjugated secondary antibody. For each sample, signal intensity of biotin-labeled proteins in the HA(+) lane was corrected for the expression level and the background signal shown in HA(−) lane. For each blot, corrected values of biotin-labeled intensities obtained from three independent experiments were normalized to the mean value of that of HCN2, and are shown below each blot (n = 3). **p < 0.01; ***p < 0.001; ns not significant versus HCN2 (one-way ANOVA followed by Dunnett’s test)