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Focal activation of a mutant allele defines the role of stem cells in mosaic skin disorders.

Arin MJ, Longley MA, Wang XJ, Roop DR - J. Cell Biol. (2001)

Bottom Line: This genetic model allows activation of a somatic K10 mutation in epidermal stem cells in a spatially and temporally controlled manner using an inducible Cre recombinase.Our results indicate that lack of selective pressure against certain mutations in epidermal stem cells leads to mosaic phenotypes.This finding has important implications for the development of new strategies for somatic gene therapy of dominant genodermatoses.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.

ABSTRACT
Stem cells are crucial for the formation and maintenance of tissues and organs. The role of stem cells in the pathogenesis of mosaic skin disorders remains unclear. To study the molecular and cellular basis of mosaicism, we established a mouse model for the autosomal-dominant skin blistering disorder, epidermolytic hyperkeratosis (MIM 113800), which is caused by mutations in either keratin K1 or K10. This genetic model allows activation of a somatic K10 mutation in epidermal stem cells in a spatially and temporally controlled manner using an inducible Cre recombinase. Our results indicate that lack of selective pressure against certain mutations in epidermal stem cells leads to mosaic phenotypes. This finding has important implications for the development of new strategies for somatic gene therapy of dominant genodermatoses.

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Gross appearance and skin morphology of mutneo/mutneo mice. (A) mutneo/mutneo pup shortly after birth with severe blistering and erosions. (B) Histological analysis of a skin biopsy from A shows a split in the suprabasal layer of the epidermis. Hematoxylin and eosin, bar = 50 μm. (C) Immunolabeling shows abundant mutant K10 (yellow) throughout the disintegrating suprabasal layer against the red K14 background stain. Bars = 50 μm.
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Figure 4: Gross appearance and skin morphology of mutneo/mutneo mice. (A) mutneo/mutneo pup shortly after birth with severe blistering and erosions. (B) Histological analysis of a skin biopsy from A shows a split in the suprabasal layer of the epidermis. Hematoxylin and eosin, bar = 50 μm. (C) Immunolabeling shows abundant mutant K10 (yellow) throughout the disintegrating suprabasal layer against the red K14 background stain. Bars = 50 μm.

Mentions: An interesting aspect of the mouse models generated in this study (+/mutneo, +/mutloxP) is the correlation of a mild phenotype with reduced expression of the mutant EHK allele. Heterozygous mice that contain the point mutation and the neo cassette (+/mutneo) exhibit a very mild scaling phenotype due to suppressed expression levels of the mutant allele. RNA analysis revealed that the mRNA from the mutant allele was significantly reduced to 35–40% of the levels of the wild-type allele (data not shown). To confirm that mutant K10 mRNA was efficiently translated into protein, we mated heterozygous +/mutneo mice to obtain mice that were homozygous for the mutant allele (mutneo/mutneo). These mice showed a very severe phenotype at birth with extensive blistering and erosions, and died shortly thereafter (Fig. 4 A). Skin biopsies showed a complete disintegration of the stratum spinosum in lesional areas (Fig. 4 B). Immunofluorescence microscopy revealed abundant K10 expression in the suprabasal layers of the epidermis (Fig. 4 C). As the wild-type K10 allele is not present in this mouse, all of the K10 protein detected is expressed from the mutant alleles. Therefore, the mutant K10 mRNA is translated into protein, but the presence of wild-type K10 in heterozygous +/mutneo mice is sufficient to overcome the effects of the reduced level of mutant K10.


Focal activation of a mutant allele defines the role of stem cells in mosaic skin disorders.

Arin MJ, Longley MA, Wang XJ, Roop DR - J. Cell Biol. (2001)

Gross appearance and skin morphology of mutneo/mutneo mice. (A) mutneo/mutneo pup shortly after birth with severe blistering and erosions. (B) Histological analysis of a skin biopsy from A shows a split in the suprabasal layer of the epidermis. Hematoxylin and eosin, bar = 50 μm. (C) Immunolabeling shows abundant mutant K10 (yellow) throughout the disintegrating suprabasal layer against the red K14 background stain. Bars = 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2195990&req=5

Figure 4: Gross appearance and skin morphology of mutneo/mutneo mice. (A) mutneo/mutneo pup shortly after birth with severe blistering and erosions. (B) Histological analysis of a skin biopsy from A shows a split in the suprabasal layer of the epidermis. Hematoxylin and eosin, bar = 50 μm. (C) Immunolabeling shows abundant mutant K10 (yellow) throughout the disintegrating suprabasal layer against the red K14 background stain. Bars = 50 μm.
Mentions: An interesting aspect of the mouse models generated in this study (+/mutneo, +/mutloxP) is the correlation of a mild phenotype with reduced expression of the mutant EHK allele. Heterozygous mice that contain the point mutation and the neo cassette (+/mutneo) exhibit a very mild scaling phenotype due to suppressed expression levels of the mutant allele. RNA analysis revealed that the mRNA from the mutant allele was significantly reduced to 35–40% of the levels of the wild-type allele (data not shown). To confirm that mutant K10 mRNA was efficiently translated into protein, we mated heterozygous +/mutneo mice to obtain mice that were homozygous for the mutant allele (mutneo/mutneo). These mice showed a very severe phenotype at birth with extensive blistering and erosions, and died shortly thereafter (Fig. 4 A). Skin biopsies showed a complete disintegration of the stratum spinosum in lesional areas (Fig. 4 B). Immunofluorescence microscopy revealed abundant K10 expression in the suprabasal layers of the epidermis (Fig. 4 C). As the wild-type K10 allele is not present in this mouse, all of the K10 protein detected is expressed from the mutant alleles. Therefore, the mutant K10 mRNA is translated into protein, but the presence of wild-type K10 in heterozygous +/mutneo mice is sufficient to overcome the effects of the reduced level of mutant K10.

Bottom Line: This genetic model allows activation of a somatic K10 mutation in epidermal stem cells in a spatially and temporally controlled manner using an inducible Cre recombinase.Our results indicate that lack of selective pressure against certain mutations in epidermal stem cells leads to mosaic phenotypes.This finding has important implications for the development of new strategies for somatic gene therapy of dominant genodermatoses.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.

ABSTRACT
Stem cells are crucial for the formation and maintenance of tissues and organs. The role of stem cells in the pathogenesis of mosaic skin disorders remains unclear. To study the molecular and cellular basis of mosaicism, we established a mouse model for the autosomal-dominant skin blistering disorder, epidermolytic hyperkeratosis (MIM 113800), which is caused by mutations in either keratin K1 or K10. This genetic model allows activation of a somatic K10 mutation in epidermal stem cells in a spatially and temporally controlled manner using an inducible Cre recombinase. Our results indicate that lack of selective pressure against certain mutations in epidermal stem cells leads to mosaic phenotypes. This finding has important implications for the development of new strategies for somatic gene therapy of dominant genodermatoses.

Show MeSH
Related in: MedlinePlus