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Mutant p63 causes defective expansion of ectodermal progenitor cells and impaired FGF signalling in AEC syndrome.

Ferone G, Thomason HA, Antonini D, De Rosa L, Hu B, Gemei M, Zhou H, Ambrosio R, Rice DP, Acampora D, van Bokhoven H, Del Vecchio L, Koster MI, Tadini G, Spencer-Dene B, Dixon M, Dixon J, Missero C - EMBO Mol Med (2012)

Bottom Line: The AEC mutation exerts a selective dominant-negative function on wild-type p63 by affecting progenitor cell expansion during ectodermal development leading to a defective epidermal stem cell compartment.These phenotypes are associated with impairment of fibroblast growth factor (FGF) signalling resulting from reduced expression of Fgfr2 and Fgfr3, direct p63 target genes.Restoring Fgfr2b expression in p63(+/L514F) epithelial cells by treatment with FGF7 reactivates downstream mitogen-activated protein kinase signalling and cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Fondazione IRCCS SDN, Napoli, Italy.

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p63+/L514F epidermis is hypoplastic compared to the controlH&E staining reveals hypoplasia of p63+/L514F epidermis at the indicated times of development. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm.Quantification of epidermal thickness (µm) in +/+ and p63+/L514F skin at birth (*p-value = 0.0009; n = 6). Data are represented as mean ± SD.Immunofluorescence analysis for p63, Krt14 (basal layer), Krt10 (spinous layer), and loricrin (Lor, granular layer) at P0 reveals appropriate expression of these differentiation markers in mutant epidermis. Scale bar: 20 µm.Immunoblotting of total cell extracts from neonatal epidermis using antibodies against the indicated differentiation markers, Krt1 (spinous layer), the upper spinous layer marker involucrin (Ivl), the adherens junction component E-cadherin (Cdh1) and αTubulin (Tub) as loading control.BrdU staining of the epidermis reveals reduced proliferation in mutant epidermis during development. Left panels: BrdU positive cells are in red; nuclei are stained with DAPI. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm. Right panel: BrdU positive cells are expressed as percentage of the total number of basal cells (E13.5 *p-value = 0.0151; n = 7. E16.5 *p-value = 0.0271; n = 7). Data are represented as mean ± SD.Left panel: immunofluorescence for active caspase 3 at P0 showing low levels of apoptosis in p63+/L514F epidermis compared to wild-type. Arrows indicate apoptotic cells. Scale bar: 70 µm. Right panel: quantification of active caspase 3 positive cells. Data are represented as mean ± SD (p-value = 0.0010; n = 16).
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fig03: p63+/L514F epidermis is hypoplastic compared to the controlH&E staining reveals hypoplasia of p63+/L514F epidermis at the indicated times of development. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm.Quantification of epidermal thickness (µm) in +/+ and p63+/L514F skin at birth (*p-value = 0.0009; n = 6). Data are represented as mean ± SD.Immunofluorescence analysis for p63, Krt14 (basal layer), Krt10 (spinous layer), and loricrin (Lor, granular layer) at P0 reveals appropriate expression of these differentiation markers in mutant epidermis. Scale bar: 20 µm.Immunoblotting of total cell extracts from neonatal epidermis using antibodies against the indicated differentiation markers, Krt1 (spinous layer), the upper spinous layer marker involucrin (Ivl), the adherens junction component E-cadherin (Cdh1) and αTubulin (Tub) as loading control.BrdU staining of the epidermis reveals reduced proliferation in mutant epidermis during development. Left panels: BrdU positive cells are in red; nuclei are stained with DAPI. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm. Right panel: BrdU positive cells are expressed as percentage of the total number of basal cells (E13.5 *p-value = 0.0151; n = 7. E16.5 *p-value = 0.0271; n = 7). Data are represented as mean ± SD.Left panel: immunofluorescence for active caspase 3 at P0 showing low levels of apoptosis in p63+/L514F epidermis compared to wild-type. Arrows indicate apoptotic cells. Scale bar: 70 µm. Right panel: quantification of active caspase 3 positive cells. Data are represented as mean ± SD (p-value = 0.0010; n = 16).

Mentions: Severe hypoplasia was also observed in p63+/L514F epidermis (40% thinner than wild-type) during embryonic development and at birth, with a reduction in the number of nuclei and a more flattened appearance of cells in all layers (Fig 3A and B). In contrast, no hypoplasia was observed in epidermis of mice heterozygous for a mutation in p63 (p63+/−) (Supporting Information Fig S4A).


Mutant p63 causes defective expansion of ectodermal progenitor cells and impaired FGF signalling in AEC syndrome.

Ferone G, Thomason HA, Antonini D, De Rosa L, Hu B, Gemei M, Zhou H, Ambrosio R, Rice DP, Acampora D, van Bokhoven H, Del Vecchio L, Koster MI, Tadini G, Spencer-Dene B, Dixon M, Dixon J, Missero C - EMBO Mol Med (2012)

p63+/L514F epidermis is hypoplastic compared to the controlH&E staining reveals hypoplasia of p63+/L514F epidermis at the indicated times of development. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm.Quantification of epidermal thickness (µm) in +/+ and p63+/L514F skin at birth (*p-value = 0.0009; n = 6). Data are represented as mean ± SD.Immunofluorescence analysis for p63, Krt14 (basal layer), Krt10 (spinous layer), and loricrin (Lor, granular layer) at P0 reveals appropriate expression of these differentiation markers in mutant epidermis. Scale bar: 20 µm.Immunoblotting of total cell extracts from neonatal epidermis using antibodies against the indicated differentiation markers, Krt1 (spinous layer), the upper spinous layer marker involucrin (Ivl), the adherens junction component E-cadherin (Cdh1) and αTubulin (Tub) as loading control.BrdU staining of the epidermis reveals reduced proliferation in mutant epidermis during development. Left panels: BrdU positive cells are in red; nuclei are stained with DAPI. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm. Right panel: BrdU positive cells are expressed as percentage of the total number of basal cells (E13.5 *p-value = 0.0151; n = 7. E16.5 *p-value = 0.0271; n = 7). Data are represented as mean ± SD.Left panel: immunofluorescence for active caspase 3 at P0 showing low levels of apoptosis in p63+/L514F epidermis compared to wild-type. Arrows indicate apoptotic cells. Scale bar: 70 µm. Right panel: quantification of active caspase 3 positive cells. Data are represented as mean ± SD (p-value = 0.0010; n = 16).
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fig03: p63+/L514F epidermis is hypoplastic compared to the controlH&E staining reveals hypoplasia of p63+/L514F epidermis at the indicated times of development. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm.Quantification of epidermal thickness (µm) in +/+ and p63+/L514F skin at birth (*p-value = 0.0009; n = 6). Data are represented as mean ± SD.Immunofluorescence analysis for p63, Krt14 (basal layer), Krt10 (spinous layer), and loricrin (Lor, granular layer) at P0 reveals appropriate expression of these differentiation markers in mutant epidermis. Scale bar: 20 µm.Immunoblotting of total cell extracts from neonatal epidermis using antibodies against the indicated differentiation markers, Krt1 (spinous layer), the upper spinous layer marker involucrin (Ivl), the adherens junction component E-cadherin (Cdh1) and αTubulin (Tub) as loading control.BrdU staining of the epidermis reveals reduced proliferation in mutant epidermis during development. Left panels: BrdU positive cells are in red; nuclei are stained with DAPI. Dashed lines indicate the border between epidermis (top) and dermis (bottom). Scale bar: 50 µm. Right panel: BrdU positive cells are expressed as percentage of the total number of basal cells (E13.5 *p-value = 0.0151; n = 7. E16.5 *p-value = 0.0271; n = 7). Data are represented as mean ± SD.Left panel: immunofluorescence for active caspase 3 at P0 showing low levels of apoptosis in p63+/L514F epidermis compared to wild-type. Arrows indicate apoptotic cells. Scale bar: 70 µm. Right panel: quantification of active caspase 3 positive cells. Data are represented as mean ± SD (p-value = 0.0010; n = 16).
Mentions: Severe hypoplasia was also observed in p63+/L514F epidermis (40% thinner than wild-type) during embryonic development and at birth, with a reduction in the number of nuclei and a more flattened appearance of cells in all layers (Fig 3A and B). In contrast, no hypoplasia was observed in epidermis of mice heterozygous for a mutation in p63 (p63+/−) (Supporting Information Fig S4A).

Bottom Line: The AEC mutation exerts a selective dominant-negative function on wild-type p63 by affecting progenitor cell expansion during ectodermal development leading to a defective epidermal stem cell compartment.These phenotypes are associated with impairment of fibroblast growth factor (FGF) signalling resulting from reduced expression of Fgfr2 and Fgfr3, direct p63 target genes.Restoring Fgfr2b expression in p63(+/L514F) epithelial cells by treatment with FGF7 reactivates downstream mitogen-activated protein kinase signalling and cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Fondazione IRCCS SDN, Napoli, Italy.

Show MeSH
Related in: MedlinePlus