title: Transient expression of epidermal filaggrin in cultured cells causes collapse of intermediate filament networks with alteration of cell shape and nuclear integrity.
abstract: Filaggrin is an intermediate filament-associated protein (IFAP) that aggregates epidermal keratin filaments in vitro and is thought to perform a similar function during terminal differentiation in vivo. To test this function in living cells, we transiently expressed constructs encoding human filaggrin in both simple epithelial cells (COS-7) and rat keratinocytes. Scanning laser confocal microscopy showed that filaggrin-positive cells had collapsed keratin and vimentin intermediate filament (IF) networks, and that filaggrin partially co-localized with the IF networks. Filaggrin was also detected diffusely in the cytoplasm and nucleus. In contrast, when profilaggrin-like constructs, containing five filaggrin domains separated by the linker sequences, were expressed in cultured cells, immunoreactive granules formed. This finding is reminiscent of the insoluble nature of native profilaggrin that accumulates in keratohyalin granules in vivo, suggesting that the linker peptides (present in profilaggrin but not filaggrin) are important for granule formation. Cells expressing filaggrin also displayed disruption of the nucleus and the nuclear envelope; they rounded up and lost attachment to the substratum, in contrast to control cells over-expressing beta-galactosidase. This functional test of filaggrin in living cells supports its role in the reorganization and packing of keratin IF in epidermal differentiation. Moreover, the observed effects on cell morphology and nuclear integrity suggest that filaggrin may contribute to the form of apoptosis associated with terminal differentiation in epidermis.
authors.affiliation: Department of Oral Biology, University of Washington, Seattle 98195-7132, USA.
authors.forename: B A
authors.initials: BA
authors.lastname: Dale
authors.forename: R B
authors.initials: RB
authors.lastname: Presland
authors.forename: S P
authors.initials: SP
authors.lastname: Lewis
authors.forename: R A
authors.initials: RA
authors.lastname: Underwood
authors.forename: P
authors.initials: P
authors.lastname: Fleckman
mesh_terms: D000818:Animals; D016923:Cell Death / Q000187:drug effects; D002460:Cell Line / Q000166:cytology; D002467:Cell Nucleus / Q000187:drug effects*; D048429:Cell Size / Q000187:drug effects*; D002522:Chlorocebus aethiops; D000078404:Epidermal Cells; D004817:Epidermis / Q000737:chemistry; D000091344:Filaggrin Proteins; D006801:Humans; D007381:Intermediate Filament Proteins / Q000096:biosynthesis* / Q000502:physiology; D007382:Intermediate Filaments / Q000648:ultrastructure*; D015603:Keratinocytes / Q000737:chemistry; D007668:Kidney / Q000166:cytology; D010455:Peptides / Q000737:chemistry; D017434:Protein Structure, Tertiary; D051381:Rats
chemical_list: C000717249:FLG protein, human; D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D010455:Peptides