Inhibition of Ca2+ flux is protective against PV-IgG-induced pathogenic effects in vitro
PV-IgG caused loss of keratinocyte adhesion in dispase-based dissociation assays. Treatment with inhibitors against PI4K ,PLC , IP3R or CRAC added 1 h before PV-IgG effectively blocked cell monolayer fragmentation after 2 h and 24 h. After 24 h, inhibition of PLC was the most effective and even reduced the effect of AK23 (Figure 2a-d/S2a-d).
The role of Ca2+ signalling for PV-IgG-mediated effects on DSG 1 and DSG 3 localization, keratin retraction and reorganization of cortical actin was assessed by immunostaining and F-actin staining. After 24 h of PV-IgG treatment, DSG 1 andDSG 3 immunostaining was fragmented at cell borders and partially relocated to the cytosol. Treatment with inhibitors for PI4K ,PLC , IP3R or CRAC, ameliorated pathogenic effects (Figure 3a/S3a-c).
Retraction of keratin filaments from the cell borders was visible after 24 h of PV-IgG treatment. Treatment with the inhibitors ameliorated the effect (Figure 3b/S3d). Similarly, cortical F-actin also showed defects especially close to intercellular gaps after PV-IgG treatment, which was ameliorated by the respective inhibitors (Figure 3/3S). F-actin staining was more pronounced following inhibition of PLC compared to controls (Figure 3/S3) indicating that PLC basal activity might regulate actin remodelling.