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Xenopus mutant reveals necessity of rax for specifying the eye field which otherwise forms tissue with telencephalic and diencephalic character.

ABSTRACT: The retinal anterior homeobox (rax) gene encodes a transcription factor necessary for vertebrate eye development. rax transcription is initiated at the end of gastrulation in Xenopus, and is a key part of the regulatory network specifying anterior neural plate and retina. We describe here a Xenopus tropicalis rax mutant, the first mutant analyzed in detail from a reverse genetic screen. As in other vertebrates, this nonsense mutation results in eyeless animals, and is lethal peri-metamorphosis. Tissue normally fated to form retina in these mutants instead forms tissue with characteristics of diencephalon and telencephalon. This implies that a key role of rax, in addition to defining the eye field, is in preventing alternative forebrain identities. Our data highlight that brain and retina regions are not determined by the mid-gastrula stage but are by the neural plate stage. An RNA-Seq analysis and in situ hybridization assays for early gene expression in the mutant revealed that several key eye field transcription factors (e.g. pax6, lhx2 and six6) are not dependent on rax activity through neurulation. However, these analyses identified other genes either up- or down-regulated in mutant presumptive retinal tissue. Two neural patterning genes of particular interest that appear up-regulated in the rax mutant RNA-seq analysis are hesx1 and fezf2. These genes were not previously known to be regulated by rax. The normal function of rax is to partially repress their expression by an indirect mechanism in the presumptive retina region in wildtype embryos, thus accounting for the apparent up-regulation in the rax mutant. Knock-down experiments using antisense morpholino oligonucleotides directed against hesx1 and fezf2 show that failure to repress these two genes contributes to transformation of presumptive retinal tissue into non-retinal forebrain identities in the rax mutant.

SUBMITTER: Fish MB

PROVIDER: S-EPMC4267880 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Xenopus mutant reveals necessity of rax for specifying the eye field which otherwise forms tissue with telencephalic and diencephalic character.

Fish Margaret B MB Nakayama Takuya T Fisher Marilyn M Hirsch Nicolas N Cox Amanda A Reeder Rollin R Carruthers Samantha S Hall Amanda A Stemple Derek L DL Grainger Robert M RM

Developmental biology 20140916 2

The retinal anterior homeobox (rax) gene encodes a transcription factor necessary for vertebrate eye development. rax transcription is initiated at the end of gastrulation in Xenopus, and is a key part of the regulatory network specifying anterior neural plate and retina. We describe here a Xenopus tropicalis rax mutant, the first mutant analyzed in detail from a reverse genetic screen. As in other vertebrates, this nonsense mutation results in eyeless animals, and is lethal peri-metamorphosis. ...[more]

PMID: 25224223

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Project description:In human holoprosencephaly (HPE), the forebrain does not separate fully into two hemispheres. Further, the border between the telencephalon and diencephalon, the telencephalic/diencephalic junction (TDJ), is often indistinct, and the ventricular system can be blocked at the third ventricle, creating a forebrain 'holosphere'. Mice deficient in Sonic Hedgehog (Shh) have previously been described to show HPE and associated cyclopia. Here we report that the third ventricle is blocked in Shh null mutants, similar to human HPE, and that characteristic telencephalic and diencephalic signaling centers, the cortical hem and zona limitans intrathalamica (ZLI), are merged, obliterating the TDJ. The resulting forebrain holosphere comprises Foxg1-positive telencephalic- and Foxg1-negative diencephalic territories. Loss of one functional copy of Gli3 in Shh nulls rescues ventricular collapse and substantially restores the TDJ. Characteristic regional gene expression patterns are rescued on the telencephalic side of the TDJ but not in the diencephalon. Further analysis of compound Shh;Gli3 mutants revealed an unexpected type of signaling center deregulation. In Shh;Gli3 mutants, adjacent rings of Fgf8 and Wnt3a expression are induced in the diencephalon at the ZLI, reminiscent of the Fgf8/Wnt1-expressing isthmic organizer. Neither Shh nor Gli3 single mutants show this forebrain double ring of Fgf/Wnt expression; thus both Shh and Gli3 are independently required to suppress it. Adjacent tissue is not respecified to a midbrain/hindbrain fate, but shows overgrowth, consistent with ectopic mitogen expression. Our observations indicate that the separation of the telencephalon and diencephalon depends on interactions between Shh and Gli3, and, moreover, demonstrate that both Shh and Gli3 suppress a potential Fgf/Wnt signaling source in the forebrain. That optional signaling centers are actively repressed in normal development is a striking new insight into the processes of vertebrate brain development.

Dataset Information

Xenopus mutant reveals necessity of rax for specifying the eye field which otherwise forms tissue with telencephalic and diencephalic character.

Publications

Xenopus mutant reveals necessity of rax for specifying the eye field which otherwise forms tissue with telencephalic and diencephalic character.

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