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Analysis of the spatial pattern of expression of embryo-specific epidermal cytokeratin genes in Xenopus laevis shows earliest activity in the animal pole cells of stage-9 blastulae. These genes are transcribed predominantly in the epithelial or outer ectoderm, to a lesser extent in the sensorial or inner ectoderm, and at low levels if at all in other regions of the embryo. In the early gastrula the entire ectoderm, including preneural and preepidermal regions, expresses cytokeratin mRNAs; accumulation of these mRNAs in preneural cells is terminated after contact is made with involuting chordamesoderm. On the basis of this and earlier work (Sargent et al. 1986) we suggest that the pattern of expression of cytokeratin genes in frog embryogenesis is based on prelocalized components modulated by the inductive influence of involuting chordamesoderm. The cytokeratin proteins are deposited in the form of filamentous networks in both layers of the epidermis. In the epithelial layer, a much denser mesh of filaments is facing the outside of the embryo. This polarity is established at the onset of the polymerization of these filaments. Thus, intraembryonic and intracellular localization of keratin gene expression and protein deposition is established at the onset of activation of these genes.
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???displayArticle.link???Genes Dev
Figure 1. In situ hybridization of DG81 to sections of stage-40
tadpole. Hybridization is almost exclusively to the epidermis.
The ectoderm-derived neural tube (N) does not show any hybridization.
(A} Dark-field illumination; (B) bright-field illumination.
Figure 2. Localization of cytokeratin mRNA in dissected regions
of Xenopus embryos. (A) Northern blot analysis of RNA
form the animal and vegetal third of 7-, 8-, and 9-hr-old embryos
using nick-translated DG70 DNA as a probe. The cytokeratin
RNA is present in the animal pole cells from 7 hr (stage
9) on. (B) Northern blot analysis of the RNA extracted from
epithelial and sensorial ectoderm of stage-10 embryos using the
same probe as in A. The DG70 gene is predominantly expressed
in the epithelial ectoderm.
Figure 3. In situ hybridization of DG76 to a section of the
blastocoel roof of stage-10 1/2 embryo. The hybridization is to
the epithelial layer of ectoderm. Outside of embryo at top and
blastocoel cavity at bottom of figure. (A) Dark-field illumination;
IB) bright-field illumination.
Figure 4. In situ hybridization of DG76 to
sections of stage-10 1/2 embryos. Boxed areas
in A and D indicate the regions enlarged in B,
C and E, F, respectively. Dark-field (B) and
phase-contrast (C) show hybridization of
DG76 to the animal region of a stage-10 1/2
embryo. The entire epithelial ectoderm
shows hybridization. Dark-field (E) and
phase-contrast (F) show the hybridization to
the preneural (dorsal) region. Arrow shows
where ectoderm is in contact with the
leading edge of chordamesoderm.
Figure 5. In situ hybridization of DG76 to sections
of stage-12 Xenopus embryos⢠Planes of
sections of B, C and E, F are indicated in A and
D, respectively. Arrows in A and D indicate the
direction of chordamesodermal involution.
Dark-field (B) and phase-contrast (C) show hybridization
of DG76 to the extending epidermis;
the plane of this section does not cross the neuroectoderm.
E and F show a cross section; note
the lack of hybridization in the neuroectoderm.
The apparent signal in the chordamesoderm
(arrows) is due to trapped air bubbles and not to
silver grains.
Figure 6. In situ hybridization of DG76 to a section of stage-14
neurula. Neural plate is at top. The absence of cytokeratin RNA
in the neural plate region is obvious. (A) Dark-field microscopy;
{B) phase-contrast. -
Figure 7. Immunoperoxidase staining of a frozen
section of a stage-25 embryo using cytokeratin antibodies.
Staining is limited to the epidermis of the
embryo. (A) Immunoperoxidase staining; (BI phasecontrast.
Figure 8. Indirect immunofluorescence of
frozen section and dissociated cells with cytokeratin
antibodies. Two epidermal cell
layers are decorated by these antibodies. The
epithelial layer shows a concentration of antibodies
in the region facing the outside of the
embryo. (A)Immunofluorescence. {B)Phasecontrast.
{C) Indirect immunofluorescence of
a single dissociated epidermal cell from
stage-18 epithelial ectoderm; the epidermal
cytokeratins are assembled into a network of
filaments with well-defined intracellular polarity.
(D) Indirect immunofluorescence of a
dissociated cell from stage- i 1 embryo; the polarity
of the filament network is already apparent.
(£) Indirect immunofluoresccnce of
partially dissociated epidermis of stage-12
embryos as viewed from outside the embryo.
