1. Impaired Trafficking of the Desmoplakins in Cultured Darier's Disease Keratinocytes 
Jittima Dhitavat, Christian Cobbold, Natalie Leslie, Susan Burge, Alain Hovnanian 
Journal of Investigative Dermatology 
Volume 121, Issue 6, Pages (December 2003)
DOI: /j x
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Thapsigargin inhibits the trafficking of desmosomal proteins to the cell surface. NHK cells were incubated in low-calcium medium (A,D,G) or incubated in the presence (C,F,I) or absence of thapsigargin (B,E,H) for 30 min. Medium was removed, and cells were incubated in normal-calcium medium for an additional 4 h to induce cell differentiation (B,C,E,F,H,I). Cells were fixed in methanol and processed for indirect immunofluorescence and confocal microscopy. Bound mouse anti-Dp, anti-Dsg, and guinea pig anti-Dsc were detected using fluorescein isothiocyanate-conjugated antibody for the corresponding species. (J) Histogram showing quantification of the relative levels of plasma membrane protein in response to different conditions.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Desmosomal proteins colocalize with the ER membrane protein calnexin. NHK cells incubated under low-calcium conditions were fixed in methanol and processed for indirect immunofluorescence and confocal microscopy. Bound mouse anti-Dp, anti-Dsg, and guinea pig anti-Dsc (green) and anti-rabbit calnexin (red) were detected using fluorescein isothiocyanate-conjugated anti-mouse/guinea pig and Texas Red-conjugated anti-rabbit secondary antibodies.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Desmosomal proteins form insoluble aggregates in the presence of thapsigargin. NHK cells were incubated in low-calcium medium or incubated in the absence or presence of thapsigargin for 30 min, medium was removed, and cells were incubated in normal-calcium medium for an additional 4 h. Cells were lyzed in mild detergent, and lysates were centrifuged at 10,000 g for 30 min to recover a pellet of insoluble protein aggregates (A) and a supernatant containing cytosol and solubilized membrane proteins (S). Samples were analyzed by SDS-PAGE and western blot using antibodies to Dp (A), PG (B), Dsg (C), and Dsc (D).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
The trafficking of Dp is inhibited in DD keratinocytes. DD cells were incubated in low-calcium medium (A,C,E) or incubated for 4 h in normal-calcium medium to induce cell differentiation (B,D,F). Cells were fixed in methanol and processed for indirect immunofluorescence and confocal microscopy. Bound mouse anti-Dp, anti-Dsg and guinea pig anti-Dsc were detected using fluorescein isothiocyanate-conjugated antibody for the corresponding species. (G) Histogram showing quantification of the relative levels of plasma membrane protein of DD keratinocytes in comparison with NHK in response to different calcium conditions.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Dp forms detergent-insoluble aggregates in DD keratinocytes. DD cells were incubated in low-calcium medium or incubated for 4 h in normal-calcium medium to induce cell differentiation. Cells were lyzed in mild detergent, and lysates were centrifuged at 10,000 g for 30 min to recover a pellet of insoluble protein aggregates (A) and a supernatant containing cytosol and solubilized membrane proteins (S). Samples were analyzed by SDS-PAGE and western blot using antibodies to Dp (A), PG (B), Dsg (C), and Dsc (D).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Dp associates with SERCA2 during differentiation. (A) NHK cells grown in low-calcium medium were pulse-labeled with [35S]methionine and cysteine and then chased in a normal-calcium medium for 4 h. Cells were lyzed and immunoprecipitated with the indicated antibody. (B) NHK cells were incubated in normal-calcium medium for 4 h to induce cell differentiation. Cells were lyzed and immunoprecipitated with anti-SERCA2 antibody. SERCA2 immunoprecipitates were analyzed by SDS-PAGE and western blot using an anti-Dp antibody (lane 2). Lane 1, Western blot of total cell extract.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions