title: 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.
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Margaret B
authors.initials: MB
authors.lastname: Fish
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Takuya
authors.initials: T
authors.lastname: Nakayama
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Marilyn
authors.initials: M
authors.lastname: Fisher
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Hirsch
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Amanda
authors.initials: A
authors.lastname: Cox
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Rollin
authors.initials: R
authors.lastname: Reeder
authors.affiliation: Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, UK.
authors.forename: Samantha
authors.initials: S
authors.lastname: Carruthers
authors.affiliation: Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, UK.
authors.forename: Amanda
authors.initials: A
authors.lastname: Hall
authors.affiliation: Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, UK.
authors.forename: Derek L
authors.initials: DL
authors.lastname: Stemple
authors.affiliation: Department of Biology, University of Virginia, Charlottesville, VA 22904, USA.
authors.forename: Robert M
authors.initials: RM
authors.lastname: Grainger
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics; D005123:Eye / Q000196:embryology*; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D017403:In Situ Hybridization; D008856:Microscopy, Fluorescence; D009024:Morphogenesis / Q000235:genetics / Q000502:physiology*; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D016548:Prosencephalon / Q000196:embryology; D017423:Sequence Analysis, RNA; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus / Q000196:embryology* / Q000235:genetics; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*; D016335:Zinc Fingers / Q000235:genetics
chemical_list: D017931:DNA Primers; D005136:Eye Proteins; D018398:Homeodomain Proteins; C524416:Rax protein, Xenopus; D014157:Transcription Factors; D029867:Xenopus Proteins
keywords: Forebrain patterning; Retina formation; Xenopus tropicalis