title: Distinct Impact of Two Keratin Mutations Causing Epidermolysis Bullosa Simplex on Keratinocyte Adhesion and Stiffness.
abstract: Keratin filaments constitute the major component of the epidermal cytoskeleton from heterodimers of type I and type II keratin subunits. Missense mutations in keratin 5 or keratin 14, highly expressed in the basal epidermis, cause the severe skin blistering disease epidermolysis bullosa simplex (EBS) in humans by rendering the keratin cytoskeleton sensitive to mechanical stress; yet, the mechanisms by which individual mutations cause cell fragility are incompletely understood. Here, we compared the K14p.Arg125Pro with the K5p.Glu477Asp mutation, both giving rise to severe generalized EBS, by stable expression in keratin-free keratinocytes. This revealed distinctly different effects on keratin cytoskeletal organization, in agreement with in vivo observations, thus validating the cell system. Although the K14p.Arg125Pro mutation led to impaired desmosomes, downregulation of desmosomal proteins, and weakened epithelial sheet integrity upon shear stress, the K5p.Glu477Asp mutation did not impair these functions, although causing EBS with squamous cell carcinoma in vivo. Atomic force microscopy demonstrated that K14 mutant cells were even less resistant against deformation compared with keratin-free keratinocytes. Thus, a keratin mutation causing EBS compromises cell stiffness to a greater extent than the lack of keratins. Finally, re-expression of K14 in K14 mutant cells did not rescue the above defects. Collectively, our findings have implications for EBS therapy approaches.
authors.affiliation: Institute of Biology and Translational Center for Regenerative Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Melanie
authors.initials: M
authors.lastname: Homberg
authors.affiliation: Institute of Complex Systems, ICS-7: Biomechanics, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: Ramms
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Nicole
authors.initials: N
authors.lastname: Schwarz
authors.affiliation: Institute of Complex Systems, ICS-7: Biomechanics, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Dreissen
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Rudolf E
authors.initials: RE
authors.lastname: Leube
authors.affiliation: Institute of Complex Systems, ICS-7: Biomechanics, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Merkel
authors.affiliation: Institute of Complex Systems, ICS-7: Biomechanics, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Hoffmann
authors.affiliation: Institute of Biology and Translational Center for Regenerative Medicine, University of Leipzig, Leipzig, Germany. Electronic address: thomas.magin@trm.uni-leipzig.de.
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Magin
mesh_terms: D002448:Cell Adhesion / Q000235:genetics; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D018450:Disease Progression; D016110:Epidermolysis Bullosa Simplex / Q000235:genetics* / Q000473:pathology; D006801:Humans; D007382:Intermediate Filaments / Q000235:genetics; D053547:Keratin-14 / Q000235:genetics*; D053555:Keratin-5 / Q000235:genetics*; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D020125:Mutation, Missense; D012494:Sampling Studies; D012720:Severity of Illness Index; D012867:Skin / Q000473:pathology*; D018709:Statistics, Nonparametric
chemical_list: C507540:KRT14 protein, human; C503940:KRT5 protein, human; D053547:Keratin-14; D053555:Keratin-5