XB-ART-37157Dev Dyn. March 1, 2008; 237 (3): 768-79.
Here, we report the localization within embryonic tissues of xWnt6 protein; together with the temporal and spatial expression of Xenopus laevis Wnt6 mRNA. Wnt6 expression in Xenopus embryos is low until later stages of neurulation, when it is predominantly found in the surface ectoderm. Wnt6 expression increases during early organogenesis in the epidermis overlaying several developing organs, including the eye, heart, and pronephros. At later stages of development, Wnt6 mRNA and protein generally localize in epithelial tissues and specifically within the epithelial tissues of these developing organs. Wnt6 localization correlates closely with sites of both epithelial to mesenchymal transformations and mesenchymal to epithelial transformations. Xenopus Wnt6 sequence and its expression pattern are highly conserved with other vertebrates. Xenopus embryos, therefore, provide an excellent model system for investigating the function of vertebrate Wnt6 in organ development and regulation of tissue architecture.
PubMed ID: 18224714
Article link: Dev Dyn.
Grant support: Biotechnology and Biological Sciences Research Council , Wellcome Trust
Genes referenced: hist1h4d mapk1 myc rho tbx2 tnnt2 wnt4 wnt6
Antibodies referenced: Tnnt2 Ab1
Article Images: [+] show captions
|Figure 2. xWnt6 mRNA is expressed during Xenopus organogenesis. A: Quantitative reverse transcriptase-polymerase chain reaction analysis shows the relative expression levels of Wnt6 mRNA. Wnt6 expression is normalized to histone H4 levels. B-K: xWnt6 mRNA localization determined by whole-mount in situ hybridization using a digoxigenin-labeled xWnt6-specific LNA probe. L-P: Control embryos hybridized with a digoxigenin-labeled nonspecific LNA control probe. Numerical figures indicate the developmental stage. Note, xWnt6 mRNA initially localizes to animal pole and neural fold tissues, then more generally throughout the ectoderm and later in epithelial tissues and in the somites, the developing eye and pronephros. ba, branchial arches; c, cloaca; ea, eye anlage; g, gills; h, heart; hm, hypaxial muscles; nf, neural folds; pa, pronephric anlage; pt, pronephric tubules; s, somites.|
|Figure 3. xWnt6 mRNA is localized within epithelial tissues and developing organs. A-C,E-G,I-K,M-O,Q-S,U-W,Y,Z: Cross-sections of embryos hybridized with a digoxigenin-labeled xWnt6-specific LNA probe. D,H,L,P,T,X,AA: Cross-sections of embryos hybridized with a digoxigenin-labeled nonspecific LNA control probe. Numerical figures indicate developmental stage. ba, branchial arches; ce, corneal epithelium; e, eye; ec, ectoderm; ev, eye vesicle, f, fin; h, heart; hg, hindgut; i, intestine; iev, invaginating eye vesicle; l, lens; lp, lens placode; nf, neural folds; p, pharynx; pt, pronephric tubules; s, somite; vf, ventral fin; vp, velar plate. Scale bars = 200 mu m.|
|Figure 4. Wnt6 protein is expressed at different stages of development and is localized to the epidermis overlaying developing organs and at later stages to the pronephric tubules. A,B: Western blots showing both endogenous xWnt6 expression as well as overexpression of xWnt6 and mWnt6 in lanes two and three at 3 hr after injection (A) and 5 hr after injection (B). A,B middle panels: Immunodetection showing expression of myc-tagged proteins xWnt4 and xWnt5A at 2 hr and 5 hr after injection, respectively. A,B bottom panels: Immunodetection of ERK2, demonstrating relatively equal protein loading in each lane. C: Western blot showing xWnt6 protein expression at different stages of development. C bottom panel: Immunodetection of ERK2, demonstrating equal protein loading in each lane. D-F,H,I: Whole-mount immunocytochemistry staining using an anti-Wnt6 antibody. G: Control embryo stained without primary antibody and with secondary antibody alone. I: Insert showing close-up of xWnt6 protein expression in the pronephric tubules at stage 40. Numerical figures indicate developmental stage. e, eye; h, heart, p, pronephros; pt, pronephric tubules.|
|Figure 5. xWnt6 protein is localized within the ectoderm and epithelial tissues and within the developing heart, eye, and pronephros. A-C,E-G,I-K,M-O,Q-S,U-W: Immunofluorescence staining performed on cryosectioned embryos using an anti-Wnt6 antibody. D,H,L,P,T,X: Control sections stained without primary antibody and with secondary antibody alone. Y,Z,AA,BB,CC,DD,EE,FF: Higher magnification immunofluorescence staining performed on cryosectioned embryos using an anti-Wnt6 antibody (green) and an anti-troponin T antibody (red). GG,HH: Confocal optical frontal sections showing Wnt6 (green) and troponin T staining (red) in isolated Xenopus hearts. II,JJ: Immunofluorescence staining performed on cryosectioned embryos using an anti-Wnt6 antibody (green) and an anti-rhodopsin antibody (red). KK,LL,MM: Confocal three-dimensional images showing Wnt6 staining of the proximal tubules. The location of the sections is indicated for each developmental stage. Numerical figures indicate developmental stage. a, atrium; ba, branchial arches; ce, corneal epithelium; e, eye; ec, ectoderm, ee, endocardial endothelium; en, endocardium; ev, eye vesicle, f, fin; fav, future arterial valve; fg, foregut; g, gut; hs, head somite; i, intestine; l, lens; ld, liver diverculum; m, myocardium; mg, midgut; n, neuroceol; ose, ocular surface ectoderm; p, pericardium; pa, pronephric anlage; ph, pharynx; pt, pronephric tubules; r, rods; ros, rods outer segment; s, somite; vp, velar plate. Scale bars = 200 mu m in A-X, 100 mu m in Y-MM.|
|Tnnt2 (troponin T type 2 (cardiac)) gene expression in Xenopus laevis embryos, NF stage 42, as assayed by immunohistochemistry (red channel). Frontal Section, dorsal up. Red channel Lateral view: anterior left, dorsal up. Image copyrighted by Wiley-Liss , 2008 Image extracted from XB-IMG-42163 and originally published in: Lavery DL et al. (2008)|