title: Directed expression of a chimeric type II keratin partially rescues keratin 5-null mice.
abstract: The crucial role of structural support fulfilled by keratin intermediate filaments (IFs) in surface epithelia likely requires that they be organized into cross-linked networks. For IFs comprised of keratins 5 and 14 (K5 and K14), which occur in basal keratinocytes of the epidermis, formation of cross-linked bundles is, in part, self-driven through cis-acting determinants. Here, we targeted the expression of a bundling-competent KRT5/KRT8 chimeric cDNA (KRT8bc) or bundling-deficient wild type KRT8 as a control to the epidermal basal layer of Krt5-null mice to assess the functional importance of keratin IF self-organization in vivo. Such targeted expression of K8bc rescued Krt5-null mice with a 47% frequency, whereas K8 completely failed to do so. This outcome correlated with lower than expected levels of K8bc and especially K8 mRNA and protein in the epidermis of E18.5 replacement embryos. Ex vivo culture of embryonic skin keratinocytes confirmed the ability of K8bc to form IFs in the absence of K5. Additionally, electron microscopy analysis of E18.5 embryonic skin revealed that the striking defects observed in keratin IF bundling, cytoarchitecture, and mitochondria are partially restored by K8bc expression. As young adults, viable KRT8bc replacement mice develop alopecia and chronic skin lesions, indicating that the skin epithelia are not completely normal. These findings are consistent with a contribution of self-mediated organization of keratin IFs to structural support and cytoarchitecture in basal layer keratinocytes of the epidermis and underscore the importance of context-dependent regulation for keratin genes and proteins in vivo.
authors.affiliation: From the Training Program in Cellular and Molecular Medicine and Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, The Johns Hopkins University, Baltimore, Maryland 21205.
authors.forename: David M
authors.initials: DM
authors.lastname: Alvarado
authors.affiliation: From the Training Program in Cellular and Molecular Medicine and Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, The Johns Hopkins University, Baltimore, Maryland 21205Departments of Biological Chemistry and Dermatology, School of Medicine and coulombe@jhu.edu.
authors.forename: Pierre A
authors.initials: PA
authors.lastname: Coulombe
mesh_terms: D000505:Alopecia / Q000196:embryology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D004622:Embryo, Mammalian / Q000378:metabolism / Q000473:pathology; D015870:Gene Expression; D040181:Genetic Diseases, X-Linked / Q000196:embryology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D053569:Keratin-15; D053555:Keratin-5 / Q000235:genetics / Q000378:metabolism*; D053533:Keratin-8 / Q000096:biosynthesis* / Q000235:genetics; D015603:Keratinocytes / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D018345:Mice, Knockout; D041681:NIH 3T3 Cells; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012867:Skin / Q000196:embryology / Q000473:pathology
chemical_list: D053569:Keratin-15; D053555:Keratin-5; D053533:Keratin-8; C492053:Krt15 protein, mouse; C486595:Krt8 protein, mouse; D011993:Recombinant Fusion Proteins
keywords: Blistering; Cell Fragility; Epidermis; Epidermolysis Bullosa Simplex; Genodermatosis; Intermediate Filament; Keratin; Mitochondria; Skin; Transgenic Mice