Extracellular epimorphin impairs expression and processing of profilaggrin in HaCaT keratinocytes

The expression and processing of filaggrin, a filament-associated protein in the skin epidermis, is closely associated with keratinocyte cornification. The large precursor profilaggrin (Pro-FLG) is initially detected at the granular layer in keratohyalin granules, subsequently processed into 10 to 12 filaggrin monomers (mFLGs) for keratin assembly, and ultimately degraded into smaller peptides that behave as natural moisturizing factor (NMF) at the outermost epidermis. We previously reported that epimorphin (EPM) extruded upon external stimuli severely perturbs epidermal terminal differentiation. Using HaCaT keratinocytes with inducible expression and recombinant EPM and FLG, we investigated the effect of extracellular EPM on the expression profile of filaggrin. As expression and processing of Pro-FLG in primary keratinocytes are accompanied with apoptotic cell death, we employed HaCaT keratinocytes that grow and express filaggrin mRNA in standard culture medium. In response to ectopic stimulation with extracellular EPM, Pro-FLG expression decreased with elimination of keratohyalin granules in the cells, with filaggrin mRNA remained constant and profilaggrin processing was not accelerated. Additionally, using a recombinant form of mFLG engineered for intracellular localization, we found that extracellular EPM hindered proteolytic cleavage of mFLG for production of NMF. Taken together, extracellularly extruded EPM, an epidermal cornification blocker, not only decreases Pro-FLG expression but also reduces the production of NMF in HaCaT keratinocytes.