Inducible expression of filaggrin increases keratinocyte susceptibility to apoptotic cell death

Abstract

Filaggrin is an intermediate filament associated protein that aids the packing of keratin filaments during terminal differentiation of keratinocytes. Premature aggregation of keratin filaments is prevented by filaggrin expression as the inactive precursor, profilaggrin, which is localized in keratohyalin granules in vivo. We have previously shown that filaggrin constructs, when transiently transfected into epithelial cells, lead to a collapsed keratin cytoskeletal network and dysmorphic nuclei with features of apoptosis. The apparent transfection rate is low with filaggrin constructs, supporting their disruptive role but hindering further study. To bypass this problem, we generated stable keratinocyte cell lines that express mature human filaggrin using a tetracyclineinducible promoter system. We found that cell lines expressing filaggrin, but not control cell lines, exhibited increased sensitivity to multiple apoptotic stimuli as measured by morphologic and biochemical criteria. None of the cell lines showed an increase in endogenous expression of filaggrin in response to the same stimuli. Filaggrin expression alone was insufficient to induce apoptosis in these keratinocyte cell lines. We conclude that filaggrin, due to its keratin binding ability, primes cells for apoptosis. Because filaggrin is expressed at a level of the epidermis where keratinocytes are in transition between the nucleated granular and the anucleate cornified layers, we hypothesize that filaggrin aids in the terminal differentiation process by facilitating apoptotic machinery. Cell Death and Differentiation (2000) 7, 566 ± 573.

AKT1-mediated Lamin A/C degradation is required for nuclear degradation and normal epidermal terminal differentiation

“…Filaggrin has been shown to be associated with nuclear degradation and keratin intermediate filament aggregation. 28,29 Studies have reported that keratohyalin granules with profilaggrin aggregates localise to the perinuclear region sometimes forming deep invaginations at the areas of association. 28,29 This is thought to facilitate entry of cleaved N-terminal domain of profilaggrin into the nucleus, where it favours nuclear degradation.…”

“…28,29 Studies have reported that keratohyalin granules with profilaggrin aggregates localise to the perinuclear region sometimes forming deep invaginations at the areas of association. 28,29 This is thought to facilitate entry of cleaved N-terminal domain of profilaggrin into the nucleus, where it favours nuclear degradation. 28 One potential explanation for the altered processing of filaggrin, partially supported by our data, is that filaggrin interacts with nuclear lamins and therefore the persistence of Lamin A/C increases its association with filaggrin intermediates at the nuclear lamina, preventing its processing to more mature forms.…”

Effect of topical magnesium application on epidermal integrity and barrier function

“…These cytokines are also expressed through similar pathways under magnesium deficient conditions (Miyazawa M et al 2008;Lee et al 2011). This TNF-a increase acts as an apoptotic signal, in response to which the keratinocytes are primed by FLG to aid its terminal differentiation, leading to the formation of corneocytes (Raj et al 2006;Kuechle et al 2000). We found that when excised skin was pre-treated with magnesium or calcium salts and subsequently stimulated with DNCB and TS, the FLG levels are always lower than that of the unstimulated salt treated skin (except in the case of 1.9M MgCl 2 stimulated with DNCB, section 6.3.3).…”

“…As mentioned earlier, the epidermal cells undergo continuous differentiation leading to terminal differentiation where stratum granulosum cells are anucleated to form stratum corneum. The role of FLG is to prepare the cells to undergo terminal differentiation (Kuechle et al 2000). In earlier studies FLG has been studied extensively for its loss-of-function related diseases characterised by malformation of the barrier layer (Bussmann et al 2011).…”

“…. This TNF-a increase acts as an apoptotic signal, in response to which the keratinocytes are primed by FLG to aid its terminal differentiation, leading to the formation of corneocytes (Raj et al 2006;Kuechle et al 2000;Eckhart et al 2000).…”