Inhibition of protein phosphorylation, but not synthesis nor lysosomal degradation, prevent keratinocyte adhesion loss induced by pemphigus vulgaris serum

Impaired hnction of desmosomes is recognized as the main cause for the loss of cohesion among keratinocytes, or acantholysis, occurring in the autoimmune disease pemphigus vulgaris (PV). However, disruption of cell-to-cell interaction induced by PV serum is a complex phenomenon passing through phosphorylation events, endocytosis and lysosomal degradation of desmoglein (Dsg) 3, and protein synthesis, leading to desmosome disassembly. But it is still unclear whether these intracellular events are actually pathogenic or instead merely represent epiphenomena. Here we adopt a pharmacological approach to address these questions. Stawosporine (STS), cycloheximide (CHX), and monensin were used to inhibit protein phospho~lation, synthesis, and lysosomal degradation, respectively. Keratinocyte changes triggered by whole PV sera were studied by means of both morphological and functional assays. Ow results show that inhibition of kinase activity through STS prevents keratinocyte cytoskeleton reorganization, depletion of Dsg3, and loss of cell-ce11 adhesion induced by PV semm. Conversely, pretreatment with either monensin or CHX does not affect the progression towards acantholysis. These data were confirmed by functional analysis, revealing that inhibition of protein phosphorylation reduced PV-treated keratinocyte dissociation score by 75%, whereas block of protein synthesis and lysosomal degradation had no effects. Thus, phosphorylation events crucially orchestrate cellular response in experimental PV and could be pharmacologically targeted to prevent acantholysis. Whether further developed, tliese findings may open new avenues toward a better therapy of the disease. Key words: pho-yliorylation, degradation, desmoglein 3, pemphigus, kinases