Anti-desmoglein 3 (Dsg3) Monoclonal Antibodies Deplete Desmosomes of Dsg3 and Differ in Their Dsg3-depleting Activities Related to Pathogenicity

Yukari Yamamoto,Yumi Aoyama,En Shu,Kazuyuki Tsunoda,Masayuki Amagai,Yasuo Kitajima

doi:10.1074/jbc.m607963200

Yukari Yamamoto, Yumi Aoyama + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m607963200

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Abstract

Pemphigus vulgaris (PV) is an autoimmune blistering disease, characterized by the loss of cell-cell adhesion between epidermal keratinocytes and the presence of autoantibody against desmoglein 3 (Dsg3), which provides adhesive integrity to desmosomes between adjacent keratinocytes. We have previously shown that PV-IgG purified from patients depletes desmosomes of Dsg3. However, PV-IgG contains not only antibodies against a variety of different epitopes of Dsg3 but also against other unknown antigens. Therefore, we examined whether the Dsg3-depleting activity of PV-IgG is generated specifically by anti-Dsg3 activity in a human squamous cell carcinoma cell line (DJM-1) and normal human keratinocytes by using four different pathogenic and nonpathogenic monoclonal antibodies against Dsg3. We demonstrate that these monoclonal antibodies deplete cells and desmosomes of Dsg3, as PV-IgG does. Individual monoclonal anti-Dsg3 antibodies display characteristic limits to their Dsg3-depleting activity, which correlates with their pathogenic activities. In combination, these antibodies exert a cumulative or synergistic effect, which may explain the potent Dsg3-depleting capability of PV-IgG, which is polyclonal. Finally, although Dsg3-depletion activity correlated with AK-monoclonal antibody pathogenicity in mouse models, the residual level of Dsg3, when below approximately 50%, does not correlate with the adhesive strength index in the present study. This may suggest that although the Dsg3 depletion is not indicative for adhesive strength, the level of Dsg3 can be used as a read-out of pathogenic changes within the cell and that the Dsg3 depletion from desmosomes plays an important role in skin fragility or susceptibility to blister formation in PV patients.

Highlights

Characterized by suprabasal acantholysis in the epidermis and autoantibodies against desmoglein 3 (Dsg3) [1,2,3,4], and pemphigus foliaceus, characterized by superficial acantholysis in the granular cell layer of the epidermis and autoantibodies against Dsg1 [5, 6]
AK23 monoclonal antibody (mAb) co-localized with Dsg3 in a punctate, linear manner at cell-cell contacts at 30 min (Fig. 1A), whereas AK23 mAb and Dsg3 disappeared from the cell and cell borders by 24 h after stimulation (Fig. 1B)
Pathogenic Anti-Dsg3 Monoclonal Antibody AK23 Depletes Cells of Dsg3, but Did Not Affect the Levels of DPK and Dsg2: Semi-quantitative Immunoblot Analysis Normalized to PG, ␤-Catenin, and ␣-Tubulin—Because we have shown that stimulation with Pemphigus vulgaris (PV)-IgG for 24 h causes a prominent reduction of Dsg3 from DJM-1 cells [18], we confirmed this result semiquantitatively and examined whether the pathogenic anti-Dsg3 monoclonal antibody, AK23, depletes cells of Dsg3, by semiquantitative immunoblot analysis of total cell lysates of cells exposed to AK23 mAb (0.5 mg/ml) and PV-IgG (0.5 mg/ml)

Summary

Introduction

Characterized by suprabasal acantholysis (the loss of cell-cell adhesion between keratinocytes) in the epidermis and autoantibodies against Dsg3 [1,2,3,4], and pemphigus foliaceus, characterized by superficial acantholysis in the granular cell layer of the epidermis and autoantibodies against Dsg1 [5, 6].

Results

Conclusion