Sequential roles for Otx2 in visceral endoderm and neuroectoderm for forebrain and midbrain induction and specification

The homeobox gene Otx2 is a mouse cognate of the Drosophila orthodenticle gene, which is required for development of the brain, rostral to rhombomere three. We have investigated the mechanisms involved in this neural function and specifically the requirement for Otx2 in the visceral endoderm and the neuroectoderm using chimeric analysis in mice and explant recombination assay. Analyses of chimeric embryos composed of more than 90% of Otx2−/− ES cells identified an essential function for Otx2 in the visceral endoderm for induction of the forebrain and midbrain. The chimeric studies also demonstrated that an anterior neural plate can form without expressing Otx2. However, in the absence of Otx2, expression of important regulatory genes, such as Hesx1/Rpx, Six3, Pax2, Wnt1 and En, fail to be initiated or maintained in the neural plate. Using explant-recombination assay, we could further demonstrate that Otx2 is required in the neuroectodem for expression of En. Altogether, these results demonstrate that Otx2 is first required in the visceral endoderm for the induction, and subsequently in the neuroectoderm for the specification of forebrain and midbrain territories.

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Overexpression of the forebrain-specific homeobox gene six3 induces rostral forebrain enlargement in zebrafish

Development ◽

10.1242/dev.125.15.2973 ◽

1998 ◽

Vol 125 (15) ◽

pp. 2973-2982 ◽

Cited By ~ 9

Author(s):

M. Kobayashi ◽

R. Toyama ◽

H. Takeda ◽

I.B. Dawid ◽

K. Kawakami

Keyword(s):

Homeobox Gene ◽

Cloning And Expression ◽

Optic Stalk ◽

Sine Oculis ◽

Murine Homolog ◽

Enhanced Expression ◽

Rostral Region ◽

The Brain ◽

Early Gastrula ◽

Anterior Neural Plate

The Drosophila homeobox gene sine oculis is expressed in the rostral region of the embryo in early development and is essential for eye and brain formation. Its murine homolog, Six3, is expressed in the anterior neural plate and eye anlage, and may have crucial functions in eye and brain development. In this study, we describe the cloning and expression of zebrafish six3, the apparent ortholog of the mouse Six3 gene. Zebrafish six3 transcripts are first seen in hypoblast cells in early gastrula embryos and are found in the anterior axial mesendoderm through gastrulation. six3 expression in the head ectoderm begins at late gastrula. Throughout the segmentation period, six3 is expressed in the rostral region of the prospective forebrain. Overexpression of six3 in zebrafish embryos induced enlargement of the rostral forebrain, enhanced expression of pax2 in the optic stalk and led to a general disorganization of the brain. Disruption of either the Six domain or the homeodomain abolish these effects, implying that these domains are essential for six3 gene function. Our results suggest that the vertebrate Six3 genes are involved in the formation of the rostral forebrain.

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Cell autonomous and non-cell autonomous functions of Otx2 in patterning the rostral brain

Development ◽

10.1242/dev.126.19.4295 ◽

1999 ◽

Vol 126 (19) ◽

pp. 4295-4304 ◽

Cited By ~ 9

Author(s):

M. Rhinn ◽

A. Dierich ◽

M. Le Meur ◽

S. Ang

Keyword(s):

Cell Adhesion ◽

Target Genes ◽

Homeobox Gene ◽

Visceral Endoderm ◽

Wild Type ◽

Candidate Target ◽

Important Regulator ◽

Mutant Cells ◽

The Brain ◽

Brain Patterning

Previous studies have shown that the homeobox gene Otx2 is required first in the visceral endoderm for induction of forebrain and midbrain, and subsequently in the neurectoderm for its regional specification. Here, we demonstrate that Otx2 functions both cell autonomously and non-cell autonomously in neurectoderm cells of the forebrain and midbrain to regulate expression of region-specific homeobox and cell adhesion genes. Using chimeras containing both Otx2 mutant and wild-type cells in the brain, we observe a reduction or loss of expression of Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2 in mutant cells, whereas expression of En2 and Six3 is rescued by surrounding wild-type cells. Forebrain Otx2 mutant cells subsequently undergo apoptosis. Altogether, this study demonstrates that Otx2 is an important regulator of brain patterning and morphogenesis, through its regulation of candidate target genes such as Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2.

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Head induction in the chick by primitive endoderm of mammalian, but not avian origin

Development ◽

10.1242/dev.126.4.815 ◽

1999 ◽

Vol 126 (4) ◽

pp. 815-825 ◽

Cited By ~ 5

Author(s):

H. Knoetgen ◽

C. Viebahn ◽

M. Kessel

Keyword(s):

Lower Layer ◽

Homeobox Gene ◽

Primitive Streak ◽

Neural Plate ◽

Visceral Endoderm ◽

Avian Origin ◽

Bmp Antagonist ◽

Rabbit Embryos ◽

Inductive Potential ◽

Anterior Visceral Endoderm

Different types of endoderm, including primitive, definitive and mesendoderm, play a role in the induction and patterning of the vertebrate head. We have studied the formation of the anterior neural plate in chick embryos using the homeobox gene GANF as a marker. GANF is first expressed after mesendoderm ingression from Hensen's node. We found that, after transplantation, neither the avian hypoblast nor the anterior definitive endoderm is capable of GANF induction, whereas the mesendoderm (young head process, prechordal plate) exhibits a strong inductive potential. GANF induction cannot be separated from the formation of a proper neural plate, which requires an intact lower layer and the presence of the prechordal mesendoderm. It is inhibited by BMP4 and promoted by the presence of the BMP antagonist Noggin. In order to investigate the inductive potential of the mammalian visceral endoderm, we used rabbit embryos which, in contrast to mouse embryos, allow the morphological recognition of the prospective anterior pole in the living, pre-primitive-streak embryo. The anterior visceral endoderm from such rabbit embryos induced neuralization and independent, ectopic GANF expression domains in the area pellucida or the area opaca of chick hosts. Thus, the signals for head induction reside in the anterior visceral endoderm of mammals whereas, in birds and amphibia, they reside in the prechordal mesendoderm, indicating a heterochronic shift of the head inductive capacity during the evolution of mammalia.

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Expression of Pax-3 is initiated in the early neural plate by posteriorizing signals produced by the organizer and by posterior non-axial mesoderm

Development ◽

10.1242/dev.124.10.2075 ◽

1997 ◽

Vol 124 (10) ◽

pp. 2075-2085 ◽

Cited By ~ 17

Author(s):

A.G. Bang ◽

N. Papalopulu ◽

C. Kintner ◽

M.D. Goulding

Keyword(s):

Retinoic Acid ◽

Neural Tube ◽

Retinoic Acid Receptor ◽

Homeobox Gene ◽

Neural Plate ◽

Transcript Expression ◽

Fgf Receptor ◽

Acid Receptor ◽

Anterior Neural Plate ◽

Lines Of Evidence

Pax-3 is a paired-type homeobox gene that is specifically expressed in the dorsal and posterior neural tube. We have investigated inductive interactions that initiate Pax-3 transcript expression in the early neural plate. We present several lines of evidence that support a model where Pax-3 expression is initiated by signals that posteriorize the neuraxis, and then secondarily restricted dorsally in response to dorsal-ventral patterning signals. First, in chick and Xenopus gastrulae the onset of Pax-3 expression occurs in regions fated to become posterior CNS. Second, Hensen's node and posterior non-axial mesoderm which underlies the neural plate induce Pax-3 expression when combined with presumptive anterior neural plate explants. In contrast, presumptive anterior neural plate explants are not competent to express Pax-3 in response to dorsalizing signals from epidermal-ectoderm. Third, in a heterospecies explant recombinant assay with Xenopus animal caps (ectoderm) as a responding tissue, late, but not early, Hensen's node induces Pax-3 expression. Chick posterior non-axial mesoderm also induces Pax-3, provided that the animal caps are neuralized by treatment with noggin. Finally we show that the putative posteriorizing factors, retinoic acid and bFGF, induce Pax-3 in neuralized animal caps. However, blocking experiments with a dominant-inhibitory FGF receptor and a dominant-inhibitory retinoic acid receptor suggest that Pax-3 inductive activities arising from Hensen's node and posterior non-axial mesoderm do not strictly depend on FGF or retinoic acid.

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Ectopic expression of a genomic fragment containing a homeobox causes neural defects in the axolotl

Biochemistry and Cell Biology ◽

10.1139/o91-056 ◽

1991 ◽

Vol 69 (5-6) ◽

pp. 366-374 ◽

Cited By ~ 2

Author(s):

Mary Whiteley ◽

John B. Armstrong

Keyword(s):

Pattern Formation ◽

Dna Recognition ◽

Ectopic Expression ◽

Neural Plate ◽

Ambystoma Mexicanum ◽

Genomic Fragment ◽

Severe Reduction ◽

Recognition Helix ◽

The Brain ◽

Anterior Neural Plate

An axolotl (Ambystoma mexicanum) genomic fragment containing the Ahoxl homeobox was placed under the control of the mouse hsp68 promoter, which seems to function constitutively in the axolotl. The resulting construct was injected into fertilized axolotl eggs to see if it would perturb development. Of the injected embryos, 20% showed severe reduction of the anterior neural plate. Later in development, these embryos had small heads, no eyes, and appeared to lack the normal regionalization of the brain. An additional 35% of the embryos were less severely affected, but had reduced or missing eyes. Control embryos, including ones injected with a construct missing the DNA recognition helix of the homeobox, developed normally.Key words: axolotl, homeobox, neural defects, pattern formation.

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Role of Xrx1 in Xenopus eye and anterior brain development

Development ◽

10.1242/dev.126.11.2451 ◽

1999 ◽

Vol 126 (11) ◽

pp. 2451-2460 ◽

Cited By ~ 10

Author(s):

M. Andreazzoli ◽

G. Gestri ◽

D. Angeloni ◽

E. Menna ◽

G. Barsacchi

Keyword(s):

Brain Development ◽

Transcriptional Activation ◽

Homeobox Gene ◽

Brain Regions ◽

Neural Plate ◽

Loss Of Function ◽

Expression Domain ◽

Severe Impairment ◽

Development Analysis ◽

Anterior Neural Plate

The anteriormost part of the neural plate is fated to give rise to the retina and anterior brain regions. In Xenopus, this territory is initially included within the expression domain of the bicoid-class homeobox gene Xotx2 but very soon, at the beginning of neurulation, it becomes devoid of Xotx2 transcripts in spatiotemporal concomitance with the transcriptional activation of the paired-like homeobox gene Xrx1. By use of gain- and loss-of-function approaches, we have studied the role played by Xrx1 in the anterior neural plate and its interactions with other anterior homeobox genes. We find that, at early neurula stage Xrx1 is able to repress Xotx2 expression, thus first defining the retina-diencephalon territory in the anterior neural plate. Overexpression studies indicate that Xrx1 possesses a proliferative activity that is coupled with the specification of anterior fate. Expression of a Xrx1 dominant repressor construct (Xrx1-EnR) results in a severe impairment of eye and anterior brain development. Analysis of several brain markers in early Xrx1-EnR-injected embryos reveals that anterior deletions are preceded by a reduction of anterior gene expression domains in the neural plate. Accordingly, expression of anterior markers is abolished or decreased in animal caps coinjected with the neural inducer chordin and the Xrx1-EnR construct. The lack of expansion of mid-hindbrain markers, and the increase of apoptosis in the anterior neural plate after Xrx1-EnR injection, indicate that anterior deletions result from an early loss of anterior neural plate territories rather than posteriorization of the neuroectoderm. Altogether, these data suggest that Xrx1 plays a role in assigning anterior and proliferative properties to the rostralmost part of the neural plate, thus being required for eye and anterior brain development.

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Visceral endoderm-restricted translation of Otx1 mediates recovery of Otx2 requirements for specification of anterior neural plate and normal gastrulation

Development ◽

10.1242/dev.125.24.5091 ◽

1998 ◽

Vol 125 (24) ◽

pp. 5091-5104 ◽

Cited By ~ 16

Author(s):

D. Acampora ◽

V. Avantaggiato ◽

F. Tuorto ◽

P. Briata ◽

G. Corte ◽

...

Keyword(s):

Transcriptional Control ◽

Expression Patterns ◽

Epileptic Seizures ◽

Primitive Streak ◽

Neural Plate ◽

Visceral Endoderm ◽

Temporal Expression ◽

Biochemical Activity ◽

The Difference ◽

Anterior Neural Plate

Otx1 and Otx2, two murine homologs of the Drosophila orthodenticle (otd) gene, contribute to brain morphogenesis. In particular Otx1 null mice are viable and show spontaneous epileptic seizures and abnormalities affecting the dorsal telencephalic cortex. Otx2 null mice die early in development and fail in specification of the rostral neuroectoderm and proper gastrulation. In order to determine whether Otx1(−/−)and Otx2(−/−) highly divergent phenotypes reflect differences in temporal expression or biochemical activity of OTX1 and OTX2 proteins, the Otx2-coding sequence was replaced by a human Otx1 full-coding cDNA. Homozygous mutant embryos recovered anterior neural plate and proper gastrulation but failed to maintain forebrain-midbrain identities, displaying a headless phenotype from 9 days post coitum (d.p.c.) onwards. Unexpectedly, in spite of the RNA distribution in both visceral endoderm (VE) and epiblast, the hOTX1 protein was synthesized only in the VE. This VE-restricted translation was sufficient to recover Otx2 requirements for specification of the anterior neural plate and proper organization of the primitive streak, thus providing evidence that the difference between Otx1 and Otx2 null mice phenotypes originates from their divergent expression patterns. Moreover, our data lead us to hypothesize that the differential post-transcriptional control existing between VE and epiblast cells may potentially contribute to fundamental regulatory mechanisms required for head specification.

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A Xenopus homebox gene defines dorsal-ventral domains in the developing brain

Development ◽

10.1242/dev.118.1.193 ◽

1993 ◽

Vol 118 (1) ◽

pp. 193-202 ◽

Cited By ~ 9

Author(s):

M.S. Saha ◽

R.B. Michel ◽

K.M. Gulding ◽

R.M. Grainger

Keyword(s):

Initial Step ◽

Neural Plate ◽

Developing Brain ◽

Vertebrate Development ◽

Tissue Interactions ◽

Temporal Localization ◽

Distinguishing Features ◽

The Brain ◽

Anterior Neural Plate ◽

Anterior Posterior

One of the distinguishing features of vertebrate development is the elaboration of the anterior neural plate into forebrain and midbrain, yet little is known about the early tissue interactions that regulate pattern formation in this region or the genes that mediate these interactions. As an initial step toward analyzing the process of regionalization in the anterior-most region of the brain, we have screened an anterior neural cDNA library for homeobox clones and have identified one which we have called XeNK-2 (Xenopus NK-2) because of its homology to the NK-2 family of homeobox genes. From neurula stages, when XeNK-2 is first detectable, through hatching stages, XeNK-2 mRNA is expressed primarily in the anterior region of the brain. By swimming tadpole stages, XeNK-2 expression resolves into a set of bands positioned at the forebrain-midbrain and the midbrain-hindbrain boundaries, after which XeNK-2 transcripts are no longer detectable. In addition to localized expression along the anterior-posterior axis, XeNK-2 may also play a role in the process of regionalization along the dorsal-ventral axis of the developing brain. At all stages examined, XeNK-2 mRNA is restricted to a pair of stripes that are bilaterally symmetrical in the ventral-lateral region of the brain. To begin to identify the tissue interactions that are required for the proper spatial and temporal localization of XeNK-2, we have performed a series of explant experiments. Consistent with earlier work showing that the A/P axis is not fixed at mid-gastrula stages, we show that XeNK-2 expression is activated when assayed in gastrula stage explants taken from any region along the entire A/P axis and that the tissue interactions necessary to localize XeNK-2 along the A/P axis are not completed until later neurula stages.

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