Zaraisky AG et al. (1992), A novel homeobox gene expressed in the anterior...

XB-ART-23529

Dev Biol 1992 Aug 01;1522:373-82. doi: 10.1016/0012-1606(92)90144-6.

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A novel homeobox gene expressed in the anterior neural plate of the Xenopus embryo.

Zaraisky AG , Lukyanov SA , Vasiliev OL , Smirnov YV , Belyavsky AV , Kazanskaya OV .

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To obtain gene sequences controlling the early steps of amphibian neurogenesis, we have performed differential screening of a subtractive cDNA library prepared by a novel PCR-based method from a single presumptive neural plate of a Xenopus laevis late-gastrula embryo. As a result we have isolated a fragment of a novel homeobox gene (named XANF-1, for Xenopus anterior neural folds). This gene is expressed predominantly in the anterior part of the developing nervous system. Such preferential localization of XANF-1 mRNA is established from its initially homogenous distribution in ectoderm of early gastrula. This change in the expression pattern is conditioned by a differential influence of various mesoderm regions on ectoderm: anterior mesoderm activates XANF-1 expression in the overlying ectoderm, whereas posterior axial and ventral mesoderm areas inhibit it. The data obtained demonstrate for the first time that selection of genes for specific expression in the CNS of the early vertebrate embryo is affected not only by chordamesoderm (a neural inductor) but also by ventral mesoderm.

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Species referenced: Xenopus laevis
Genes referenced: hesx1

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	FIG. 1. Subtractive cloning of neuroectoderm-specific sequences. (A) A scheme of amplified cDNA synthesis. (B) Preparation of the suhtracted neural cDNA library enriched with neuroectoderm-specific sequences and isolation of specific clones by differential screening.
	Flc. 2. Primary structnw olâ the cloned fragment of X,kNF-1 cDNA and its putative I)rotein. (A) Nucleotide sequence of the cDNA insert and deduced amino acid sequcnco of the protein. The homeodomain is underlined. The polyadenylation signal is indicated by dotted line. Sequence data have been assigned Accession No. XfOO$jS in the GeneBank (EMBL). (B) Alignment of the amino acid sequences ofâ the homeodomains from SANEâ-1, Mis. 1 (Rosa, 198!1), Xhos-3 (Ruis i Altaba and Melton, 1$1X9), and I)aired (Frigerio c,f ol., 1986). Dashes indicate identify with the corresl,onding amino acids in the SASF-1 homcwdomain. The percentage of homology between XANF-1 and each of the other homeodomains is indicated to thus right of wch scquc~nw.

	FIG. 4. PCR analysis of regional XANF-1 expression in ~~~t~~~~ws embryos. Regions taken for the analysis are marked by dotted lines. (A) Diagram shelving the dissected regions of the embryo at the late gastrula stage (sagittal section): (1) presumptive ncuroectoderm, (2) axial mesoderm, (3) endoderm, (4) presumptive epidermis. (B) Diagram showing the dissected regions of the embryo at the early gastruia stage (sagittai section). Dissected regions of ectoderm are numbered counterclockwise from the dorsal biastopor lip (on the right ). (C) Diagram showing the dissected regions of the embryo at the midgastrula stag<% (sagittai section). Dissected regions of ectodrrm arc numbered counterclockwise from the dorsal blastopor lip (on the right) to the ventral blastopor lip (on the left). (I)) The embryo at the midneurula stage as seen from the dorsal side (anterior end of the neural plate is at the top). Neural folds are sho\vn as a keyhole-shaped band located bctwwn two solid lines. Dissected regions containing the neural plate and adjacent epidermal cells are numbered in the anterior- posterior direction. (El The same view of the whole midneurula embryo as in C. Dissected regions are (1) presumptive cement gland region, (2) frontal anterior neural fold, (3) the bottom of neural plate, C-1) lateral anterior neural fold. and (5) presumptive anterior epidcrmis.
	FIG. 5. Effect of mesoderm on XANF-1 expression in ectodermal explants. Cut portions of ectoderm and mesoderm (shaded): (1) presumptive ventral ectoderm, (2) anterior presumptive neuroectoderm, (3) posterior presumptive neuroectoderm, (4) ventral mesoderm, (5) anterior axial mesoderm, and (6) posterior axial mesoderm. Prior to PCR the explants were cultured to the stage equal to that of midneurula (see Materials and Methods and text for details). Lane 1, anterior presumptive ncuroectoderm combined with posterior axial mesoderm; lane 2, anterior presumptive neuroectoderm combined with ventral mesoderm; lane 3, anterior presumptive neuroectoderm; lane 4, posterior presumptive neuroectodcrm combined with anterior axial mesoderm; lane 5, presumptive ventral ectoderm combined with anterior axial mesoderm; lane 6, anterior axial mesoderm; lane 7, posterior presumptive neuroectoderm; and lane 8, presumptive ventral ectoderm.