XB-ART-5810Proc Natl Acad Sci U S A February 18, 2003; 100 (4): 1769-74.
Transgenic tadpoles that express a dominant negative thyroid hormone (TH) receptor specifically in their skin undergo normal metamorphosis, with one exception: they retain a larval epidermis over the developing adult epithelium. TH-induced death of the tadpole epidermis is inhibited by the dominant negative TH receptor whereas the TH-induced response of the neighboring fibroblasts and the cells that form the adult skin occur normally. Therefore death of the tadpole skin is a direct and cell autonomous target of TH, and its protection has no detectable influence on TH-induced changes of other cell types.
PubMed ID: 12560472
PMC ID: PMC149908
Article link: Proc Natl Acad Sci U S A
Species referenced: Xenopus
Genes referenced: casp3.2 dan4l krt12.5 krt62 krt78.6 thdl20
Article Images: [+] show captions
|Figure 1 Larval and adult-specific epidermal markers. Tadpoles and cross sections taken from dorsal skin (vertical lines) of premetamorphic stage NF 57 (A–E) and metamorphic climax stage NF 62 (A′–E′). Sections were stained with hematoxylin and eosin (B and B′), in situ hybridizations with antisense RNA probes (dark blue/purple) against gene 19 (C and C′), larval keratin (D and D′), and adult keratin (E and E′). Eosin stains the epithelial layers bright pink; black arrowheads (B) denote subepithelial fibroblasts, white arrowheads (B′) denote adult dermis. [Scale bars: 500 μM (A and A′) and 5 μM (B–E′).]|
|Figure 2 Larval epithelial cell death at metamorphic climax coincides with expression of active caspase-3. Active caspase-3 immunoreactivity detected in whole-mount processed skin from the body is rarely present in larval skin before metamorphosis (NF 57) (A), strongly present at the start of metamorphic climax (NF 60; arrows) (B), and rarely present at the peak of climax (NF 62) after the larval skin has been replaced by adult skin (C). Adult skin glands present at NF 62 are weakly autofluorescent. (D) Active caspase-3 immunoreactivity (green) in cross sections of NF 60 skin is confined to a vacuolated cell (v) in the larval epithelium; red, 4′,6-diamidino-2-phenylindole (DAPI) nuclear counterstain; arrowheads, collagen lamella; solid arrowheads, subepithelial fibroblasts (SEF). [Scale bars: 100 μM (A–C) and 5 μM (D).]|
|Figure 3 Larval keratin colocalizes with GFP expression in tadpoles transgenic for Ker:GFP. (A) Larval keratin immunoreactivity (red) in the body skin of a premetamorphic tadpole (NF 54) is confined to the epithelial layer. (B) The larval keratin promoter drives GFP expression (green) specifically in the epithelial layers of skin from the body. (C) Cross section of the tail showing expression of Ker:GFP (white). A and B are counterstained with 4′,6-diamidino-2-phenylindole (DAPI) (white). [Scale bars: 10 μM (A and B) and 100 μM (C).]|
|Figure 4 Targeted expression of a TRDN to the larval epithelium inhibits down-regulation of gene 19 and up-regulation of active caspase-3 after induction of premetamorphic tadpoles (NF 54) with 30 nM T3 for 48 h. (A–D) WT tadpoles. (A′–D′) Tadpoles transgenic for Ker:TRDN. (A and A′) Antisense RNA probes hybridized in tail cross sections show the presence of gene 19 in both WT and transgenic tadpoles before induction. Gene 19 is down-regulated by TH in the WT skin (B) but continues to be expressed in transgenic skin after T3 induction (B′). Collagenase-3 is up-regulated by TH in subepithelial fibroblasts in WT (C) and transgenic (C′) skin. cl, collagen lamella. After T3 induction, active caspase-3 immunoreactivity (arrowheads) is broadly expressed in WT skin (D) but has much less expression in transgenic skin (D′). Black spots in whole-mounted skin (D and D′) are melanophores. [Scale bars: 5 μM (A–C′) and 200 μM (D and D′).]|
|Figure 5 Froglets (NF 64) transgenic for Ker:TRDN retain larval epithelium at the end of metamorphosis. Vertical lines denote regions of skin sectioned from the body and tail of a WT (A) and Ker:TRDN (A′) froglet. (B) Gene 19 expression (dark blue/purple) visualized by in situ hybridization is absent in WT body and tail (F) skin but retained in transgenic body (B′) and tail (F′) skin (black dots are pigment clusters). * denotes adult skin glands. Larval keratin expression in body skin is absent in WT (C) but present in transgenic (C′) skin. Adult keratin is expressed in basal cells of both WT (D) and transgenic (D′) skin. Body skin labeled with both gene 19 and adult keratin show the retention of gene 19 expression in transgenic skin (E′), but not in control skin (E). Control (F) and transgenic (F′) tail tips hybridized with gene 19 probe identify retention of larval gene expression in the Ker:TRDN animal. [Scale bars: 500 μM (A and A′), 5 μM (B–E′), and 50 μM (F and F′).]|
References [+] :
Berry, The expression pattern of thyroid hormone response genes in the tadpole tail identifies multiple resorption programs. 1998, Pubmed, Xenbase