Characterization of disease mutations targeting the DP N terminus. (A) Five disease mutations within SRs of the DP N terminus hotspot were selected for analysis. (B) DPII-GFP was created by splicing out base pairs 3,584–5,380 from a construct of DPI-GFP. (C) Immunolocalization of GFP-tagged WT and mutated DPII-GFP constructs expressed by retroviral transduction in SCC9 cells, in the context of endogenous DP. Bar, 10 µm. (D) Densitometric quantification of immunoprecipitated/total GFP for WT DP-NTP-GFP and N287K-DP-NTP-GFP coimmunoprecipitated with plakophilin-1 (PKP1-FLAG) or plakoglobin (Pg-Myc) from HEKs, used for robust protein expression (n = 2 independent experiments for each IP). (E) Dispase assay of adhesive strength for cells expressing WT or mutant DPII-GFP constructs. (E, left) The number of fragments observed. Blots show expression of DP constructs (GFP) in SCC9s. GAPDH was used as a loading control, and EB1 expression levels were equivalent across samples (n = triplicate wells from one of two independent experiments). Error bars indicate SEM.