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Fate and plasticity of the epidermis in response to congenital activation of BRAF.

Krishnaswami SR, Kumar S, Ordoukhanian P, Yu BD - J. Invest. Dermatol. (2014)

Bottom Line: Germline and somatic mutations in RAS and its downstream effectors are found in several congenital conditions affecting the skin.However, restoration of epidermal differentiation was non-cell autonomous and required dermal tissue to be present in tissue recombination studies.These studies indicate that early activation of the RAF signaling pathway in the ectoderm has effects on specific steps of epidermal differentiation, which may be amenable to treatment with currently available pharmacologic inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Division of Dermatology, Department of Medicine, Institute for Genomic Medicine, Stem Cell Program, University of California, San Diego, La Jolla, California, USA.

ABSTRACT
Germline and somatic mutations in RAS and its downstream effectors are found in several congenital conditions affecting the skin. Here we demonstrate that activation of BRAF in the embryonic mouse ectoderm triggers both craniofacial and skin defects, including hyperproliferation, loss of spinous and granular keratinocyte differentiation, and cleft palate. RNA sequencing of the epidermis confirmed these findings but unexpectedly revealed evidence of continued epidermal maturation, expression of >80% of epidermal differentiation complex genes, and formation of a hydrophobic barrier. Spinous and granular differentiation were restored by pharmacologic inhibition of MAPK/ERK kinase or BRAF. However, restoration of epidermal differentiation was non-cell autonomous and required dermal tissue to be present in tissue recombination studies. These studies indicate that early activation of the RAF signaling pathway in the ectoderm has effects on specific steps of epidermal differentiation, which may be amenable to treatment with currently available pharmacologic inhibitors.

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Reversion of BrafV600E-explants is not cell autonomous, requiring a dermal signal(a) Schematic of preparation of explants, epidermis-only vs. epidermis with dermis reattached. (b) Spinous K10 expression in epidermis-only explants vs. epidermis re-attached to dermis from wildtype and K14-cre; BrafV600E E17.5 embryos treated with BRAF inhibitor, PLX4720. Right panels demonstrate single channel of K10 expression. (c) Granular LOR expression was examined in wildtype and BrafV600E epidermis, demonstrating the requirement for dermal reattachment for proper re-activation of LOR protein expression. Right panels demonstrate single channel of LOR expression. Scale bar, 20 µm.
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Figure 6: Reversion of BrafV600E-explants is not cell autonomous, requiring a dermal signal(a) Schematic of preparation of explants, epidermis-only vs. epidermis with dermis reattached. (b) Spinous K10 expression in epidermis-only explants vs. epidermis re-attached to dermis from wildtype and K14-cre; BrafV600E E17.5 embryos treated with BRAF inhibitor, PLX4720. Right panels demonstrate single channel of K10 expression. (c) Granular LOR expression was examined in wildtype and BrafV600E epidermis, demonstrating the requirement for dermal reattachment for proper re-activation of LOR protein expression. Right panels demonstrate single channel of LOR expression. Scale bar, 20 µm.

Mentions: Because re-activation of spinous and granular keratinocyte identities appeared to be non-cell autonomous, we further tested whether surrounding tissues contribute to the epidermal response to BRAF inhibition. Previous tissue recombination studies have suggested an important role for the dermis in the histogenesis of the epidermis (Sengel, 1976). Therefore, we tested the response of epidermal sheets from wildtype and K14-cre; BrafV600E mouse embryos to PLX4720 after removal of the dermis (Fig. 6). Neither K10 nor LOR expression were restored by PLX4720 in K14-cre; BrafV600E epidermal sheets (Fig. 6a, b). As a control, we also tested the response of epidermal sheets after re-attached to dermis (Fig. 6a, b). Re-attachment of the epidermis to dermis restored the capacity of the epidermis to respond to BRAF inhibition and demonstrates the dermis directly or indirectly provides cues necessary for specification of spinous and granular keratinocyte identity.


Fate and plasticity of the epidermis in response to congenital activation of BRAF.

Krishnaswami SR, Kumar S, Ordoukhanian P, Yu BD - J. Invest. Dermatol. (2014)

Reversion of BrafV600E-explants is not cell autonomous, requiring a dermal signal(a) Schematic of preparation of explants, epidermis-only vs. epidermis with dermis reattached. (b) Spinous K10 expression in epidermis-only explants vs. epidermis re-attached to dermis from wildtype and K14-cre; BrafV600E E17.5 embryos treated with BRAF inhibitor, PLX4720. Right panels demonstrate single channel of K10 expression. (c) Granular LOR expression was examined in wildtype and BrafV600E epidermis, demonstrating the requirement for dermal reattachment for proper re-activation of LOR protein expression. Right panels demonstrate single channel of LOR expression. Scale bar, 20 µm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4289449&req=5

Figure 6: Reversion of BrafV600E-explants is not cell autonomous, requiring a dermal signal(a) Schematic of preparation of explants, epidermis-only vs. epidermis with dermis reattached. (b) Spinous K10 expression in epidermis-only explants vs. epidermis re-attached to dermis from wildtype and K14-cre; BrafV600E E17.5 embryos treated with BRAF inhibitor, PLX4720. Right panels demonstrate single channel of K10 expression. (c) Granular LOR expression was examined in wildtype and BrafV600E epidermis, demonstrating the requirement for dermal reattachment for proper re-activation of LOR protein expression. Right panels demonstrate single channel of LOR expression. Scale bar, 20 µm.
Mentions: Because re-activation of spinous and granular keratinocyte identities appeared to be non-cell autonomous, we further tested whether surrounding tissues contribute to the epidermal response to BRAF inhibition. Previous tissue recombination studies have suggested an important role for the dermis in the histogenesis of the epidermis (Sengel, 1976). Therefore, we tested the response of epidermal sheets from wildtype and K14-cre; BrafV600E mouse embryos to PLX4720 after removal of the dermis (Fig. 6). Neither K10 nor LOR expression were restored by PLX4720 in K14-cre; BrafV600E epidermal sheets (Fig. 6a, b). As a control, we also tested the response of epidermal sheets after re-attached to dermis (Fig. 6a, b). Re-attachment of the epidermis to dermis restored the capacity of the epidermis to respond to BRAF inhibition and demonstrates the dermis directly or indirectly provides cues necessary for specification of spinous and granular keratinocyte identity.

Bottom Line: Germline and somatic mutations in RAS and its downstream effectors are found in several congenital conditions affecting the skin.However, restoration of epidermal differentiation was non-cell autonomous and required dermal tissue to be present in tissue recombination studies.These studies indicate that early activation of the RAF signaling pathway in the ectoderm has effects on specific steps of epidermal differentiation, which may be amenable to treatment with currently available pharmacologic inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Division of Dermatology, Department of Medicine, Institute for Genomic Medicine, Stem Cell Program, University of California, San Diego, La Jolla, California, USA.

ABSTRACT
Germline and somatic mutations in RAS and its downstream effectors are found in several congenital conditions affecting the skin. Here we demonstrate that activation of BRAF in the embryonic mouse ectoderm triggers both craniofacial and skin defects, including hyperproliferation, loss of spinous and granular keratinocyte differentiation, and cleft palate. RNA sequencing of the epidermis confirmed these findings but unexpectedly revealed evidence of continued epidermal maturation, expression of >80% of epidermal differentiation complex genes, and formation of a hydrophobic barrier. Spinous and granular differentiation were restored by pharmacologic inhibition of MAPK/ERK kinase or BRAF. However, restoration of epidermal differentiation was non-cell autonomous and required dermal tissue to be present in tissue recombination studies. These studies indicate that early activation of the RAF signaling pathway in the ectoderm has effects on specific steps of epidermal differentiation, which may be amenable to treatment with currently available pharmacologic inhibitors.

Show MeSH
Related in: MedlinePlus