Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
???displayArticle.abstract???
Little is known of the control of gene expression in the animal hemisphere of the Xenopus embryo. Here we show that expression of FoxI1e, a gene essential for normal ectoderm formation, is expressed regionally within the animal hemisphere, in a highly dynamic fashion. In situ hybridization shows that FoxI1e is expressed in a wave-like fashion that is initiated on the dorsal side of the animal hemisphere, extends across to the ventral side by the mid-gastrula stage, and is then turned off in the dorsal ectoderm, the neural plate, at the neurula stage. It is confined to the inner layers of cells in the animal cap, and is expressed in a mosaic fashion throughout. We show that this dynamic pattern of expression is controlled by both short- and long-range signals. Notch signaling controls both the mosaic, and dorsal/ventral changes in expression, and is controlled, in turn, by Vg1 signaling from the vegetal mass. FoxI1e expression is also regulated by nodal signaling downstream of VegT. Canonical Wnt signaling contributes only to late changes in the FoxI1e expression pattern. These results provide new insights into the roles of vegetally localized mRNAs in controlling zygotic genes expressed in the animal hemisphere by long-range signaling. They also provide novel insights into the role of Notch signaling at the earliest stages of vertebrate development.
Fig. 1. FoxI1e is expressed in a dorsal to ventral wave in the blastula and gastrula stages. (A) In situ hybridization for FoxI1e shows initial staining at stage 9.5 on one side of the embryo, and then spreading across the embryo. Its expression is always mosaic. (B) Embryos injected in the two dorsal, animal blastomeres at the 4-cell stage with 50 pg of β-Gal mRNA were stained with Red-Gal before in situ hybridization, showing the initial expression is on the dorsal side of the embryo. (C) Embryos were dissected into dorsal and ventral halves at stage 10 and frozen for real-time PCR. FoxI1e expression is enriched on the dorsal side at stage 10. Results are normalized to ODC expression levels. (D) Stage 11 embryos were stained for FoxI1e and sectioned. Staining with Wheat Germ Agglutinin defines a small population of FoxI1e-positive cells between the sensorial and epithelial layers of the ectoderm. Scale bars represent 200 μm, unless otherwise noted.
Fig. 3. VegT and Nodal signaling act at long-range to affect FoxI1e expression in the animal cap. (A) In situ hybridization in VegT-depleted and CerS-injected embryos shows FoxI1e is most upregulated in the animal cap rather than the vegetal mass at stage 10. Dorsal is the to the right in the bisected uninjected embryo. The VegT and CerS embryos did not dorsal axes. (B) Vegetal masses stripped of all mesoderm contamination dissected from control and CerS-injected embryos confirm that the vast majority of increase in FoxI1e expression is derived from non-endodermal tissue. Scale bars represent 200 μm.
Fig. 4. Vg1 is a long-range inhibitor of FoxI1e expression. (A, B) Depletion of Vg1 results in a 5-fold increase in FoxI1e expression at stage 10, resulting largely from an increase in expression in non-endodermal tissues. (C) Co-depletion of VegT and Vg1 does not increase the expression of FoxI1e in the vegetal mass over either one alone, indicating the presence of an unidentified inhibitor in the vegetal mass, or the absence of an activator. Dorsal is to the right in the bisected embryos shown. Scale bars represent 200 μm.
Fig. 5. Notch signaling is responsible for the initial dorsal restriction and mosaic expression of FoxI1e. (A) Maternal Xotch mRNA is depleted to 20–25% with 15 ng of thioate-modified DNA oligo. The level of Xotch remains low in the blastula and gastrula. (B) FoxI1e expression is 2- to 3-fold upregulated by depletion of Xotch. In situ hybridization shows an expansion of FoxI1e expression (C). (D) Injection of the constitutively active Notch Intracellular Domain (NICD) causes downregulation of FoxI1e relative to controls, and injection of the dominant negative construct Su(H)-DBM causes an upregulation of FoxI1e (E). (F) NICD upregulates the Notch target ESR-1, and Su(H)-DBM downregulates it. (G) In situ hybridization for FoxI1e comparing control and Su(H)-DBM injected embryos at stage 9.5 indicates that more, and in the most severe cases, all of the sensorial-layer animal cap cells express FoxI1e. Dorsal is to the right in all embryos shown. Scale bars represent 200 μm.
Fig. 6. Maternal Vg1 activates Notch signaling in the blastula to control FoxI1e expression. (A) Vg1-depleted embryos were injected with 50 or 500 pg of NICD mRNA at the 2-cell stage. NICD rescued the increase in FoxI1e expression caused by Vg1 depletion. (B) These results were confirmed by in situ hybridization for FoxI1e at stage 10, which shows an upregulation of FoxI1e in Vg1-depleted embryos, and a reversal of this upregulation by subsequent injection with NICD. The control embryo is oriented with dorsal to the right. Depletion of Vg1 results in a delay of gastrulation, and so the orientations of both the Vg1-depleted and the NICD-rescued embryos are indeterminate. Scale bars represent 200 μm. (C) Real-time PCR at stage 10 shows that the Notch target ESR-1 is downregulated in Vg1-depleted embryos relative to controls, indicating that Notch signaling depends on Vg1 at this stage. (D) 200 pg of Vg1 mRNA was unable to rescue the increase in FoxI1e expression induced by loss of Notch signaling by injection of 500 pg Su(H)-DBM mRNA.
Fig. 2. Wnt-dependent dorsal axis formation controls late, but not early, FoxI1e expression. Embryos injected with 40 ng β-Cat MO had reduced levels of direct targets Siamois and Xnr3 at stage 10 (A), and embryos injected with 50 pg β-Cat mRNA had increased levels (B). In control explants, the level of FoxI1e was higher in dorsal halves than ventral halves (C). The total level and distribution of FoxI1e was unchanged by β-Cat MO or mRNA (D–F). (G) Levels of FoxI1e mRNA at stages 9.5, 10, and 14 (compared to ODC mRNA levels at each stage) in embryos injected with either 100 pg BMP4 mRNA (upper panels), or 10, 40, or 160 pg of noggin mRNA (lower panels). Neither BMP4 overexpression, nor inhibition using Noggin, consistently affected the level of FoxI1e expression at the late blastula stage (stage 9.5). By the early gastrula stage (stage 10), BMP4 overexpression increased FoxI1e expression, but Noggin still had little effect. Expression of FoxI1e in the early neurula (stage 14) was increased by BMP4 overexpression and completely ablated by Noggin. This indicates that the early expression of FoxI1e is BMP-independent, but that the restriction of FoxI1e to the epidermis at neurulation is BMP-dependent. (H) Overall level of FoxI1e expression at stage 14 is unaffected by β-Cat MO. Embryos injected with 40 ng β-Cat MO at the 2-cell stage were injected with β-Gal at the 32-cell stage in the A1 blastomere. Red-gal staining, and FoxI1e in situ hybridization were carried out at stage 14. The anterior (A), posterior (P), dorsal (D), and ventral (V) regions of the embryo are marked in the control embryo, with the derivatives of the dorsal animal blastomere (A1), marked by the yellow arrowhead in the neural plate. Embryos lacking β-Cat (right panel) lack axes altogether. The red-gal positive cells (arrowed) mark the derivatives of the A1 blastomere. FoxI1e expression in β-Cat-depleted embryos persists in that clone of cells, indicating the β-Cat dependence of restriction of FoxI1e from the prospective CNS. The reddish cast toward the posterior end of the uninjected embryo is residual maternal pigment, not affected by bleaching. Scale bars represent 200 μm.
Abe,
Activin-like signaling activates Notch signaling during mesodermal induction.
2004,
Pubmed
,
Xenbase
Abe,
Notch signaling modulates the nuclear localization of carboxy-terminal-phosphorylated smad2 and controls the competence of ectodermal cells for activin A.
2005,
Pubmed
,
Xenbase
Agius,
Endodermal Nodal-related signals and mesoderm induction in Xenopus.
2000,
Pubmed
,
Xenbase
Baker,
Wnt signaling in Xenopus embryos inhibits bmp4 expression and activates neural development.
1999,
Pubmed
,
Xenbase
Birsoy,
Vg 1 is an essential signaling molecule in Xenopus development.
2006,
Pubmed
,
Xenbase
Chalmers,
Grainyhead-like 3, a transcription factor identified in a microarray screen, promotes the specification of the superficial layer of the embryonic epidermis.
2006,
Pubmed
,
Xenbase
Chalmers,
Oriented cell divisions asymmetrically segregate aPKC and generate cell fate diversity in the early Xenopus embryo.
2003,
Pubmed
,
Xenbase
Coffman,
Expression of an extracellular deletion of Xotch diverts cell fate in Xenopus embryos.
1993,
Pubmed
,
Xenbase
Deblandre,
A two-step mechanism generates the spacing pattern of the ciliated cells in the skin of Xenopus embryos.
1999,
Pubmed
,
Xenbase
Dupont,
Germ-layer specification and control of cell growth by Ectodermin, a Smad4 ubiquitin ligase.
2005,
Pubmed
,
Xenbase
Harland,
In situ hybridization: an improved whole-mount method for Xenopus embryos.
1991,
Pubmed
,
Xenbase
Hawley,
Disruption of BMP signals in embryonic Xenopus ectoderm leads to direct neural induction.
1995,
Pubmed
,
Xenbase
Heasman,
Beta-catenin signaling activity dissected in the early Xenopus embryo: a novel antisense approach.
2000,
Pubmed
,
Xenbase
Heasman,
Fates and states of determination of single vegetal pole blastomeres of X. laevis.
1984,
Pubmed
,
Xenbase
Heasman,
Fertilization of cultured Xenopus oocytes and use in studies of maternally inherited molecules.
1991,
Pubmed
,
Xenbase
Houston,
The Xenopus LIM-homeodomain protein Xlim5 regulates the differential adhesion properties of early ectoderm cells.
2003,
Pubmed
,
Xenbase
Kofron,
The role of maternal axin in patterning the Xenopus embryo.
2001,
Pubmed
,
Xenbase
Luo,
Transcription factor AP-2 is an essential and direct regulator of epidermal development in Xenopus.
2002,
Pubmed
,
Xenbase
Melton,
Translocation of a localized maternal mRNA to the vegetal pole of Xenopus oocytes.
,
Pubmed
,
Xenbase
Mir,
FoxI1e activates ectoderm formation and controls cell position in the Xenopus blastula.
2007,
Pubmed
,
Xenbase
Snape,
Changes in states of commitment of single animal pole blastomeres of Xenopus laevis.
1987,
Pubmed
,
Xenbase
Suri,
Xema, a foxi-class gene expressed in the gastrula stage Xenopus ectoderm, is required for the suppression of mesendoderm.
2005,
Pubmed
,
Xenbase
Toyama,
The LIM class homeobox gene lim5: implied role in CNS patterning in Xenopus and zebrafish.
1995,
Pubmed
,
Xenbase
Yun,
Negative regulation of Activin/Nodal signaling by SRF during Xenopus gastrulation.
2007,
Pubmed
,
Xenbase
Zhang,
The role of maternal VegT in establishing the primary germ layers in Xenopus embryos.
1998,
Pubmed
,
Xenbase
Zhang,
Xenopus VegT RNA is localized to the vegetal cortex during oogenesis and encodes a novel T-box transcription factor involved in mesodermal patterning.
1996,
Pubmed
,
Xenbase
Zimmerman,
The Spemann organizer signal noggin binds and inactivates bone morphogenetic protein 4.
1996,
Pubmed
,
Xenbase
