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Neural patterning occurs soon after neural induction during early development. In Xenopus, several caudalizing factors transform anterior neural to posterior neural tissue at the open neural plate stages, while other factors are responsible for setting up mediolateral polarity which becomes the dorsoventral (D-V) axis after neural tube closure. Many Wnt ligands are expressed in the neural tube in distinct anteroposterior (A-P) and D-V domains, implying a function in neural patterning. Here we report the cloning of a full-length Xenopus Wnt7B gene. Xwnt7B induces neural crest markers Xslug and Xtwist in ectodermal explants coinjected with neural inducer noggin and in ectodermal cells neuralized by dissociation. In vivo, Xwnt7B expands the Xtwist expression domain when injected in the animal pole. Our results suggest that Wnt members are involved in dorsoventral patterning of the neural tube.
FIG. 1. Sequence and expression of Xwnt7B. (A) Sequence alignment of Xenopus Wnt7B with mouse Wnt7 genes. (B) Spatial expression of
Xwnt7B assayed by whole-mount in situ hybridization. Gastrula embryos are viewed fromanimal (panel A) or vegetal (panel B) sides, showing
even distribution of RNA in the animal half and marginal zone at this stage. Panel C is a control embryo hybridized with a Xwnt7B sense
probe. Panel D shows Xwnt7B expression in the dorsal neural tube and the epidermal layer after neural tube closure, and panel E is a tailbudembryo stained with a Xwnt7B sense probe. Panel F shows Xwnt7B expression in the dorsal neural tube and skin in a tailbud stage embryo.
Panel G shows expression of Xwnt7B in the dorsal midline of the neural tube and a segmented pattern of staining underneath the skin.
Panel H shows dorsal neural as well as epidermal staining in a transverse section of a tailbudembryo. Panels I and J show specific staining
of the midbrain–hindbrain boundary and the dorsal hindbrain of an embryo hybridized with Xwnt1 antisense probe. In panels D to J (except
panel H), the heads of the embryos are on the right side; panel I is a dorsal view while the others are lateral views.
FIG. 2. Neural crest induction by Xwnt7B in ectodermal explants. (A) Xwnt7B induces dorsal and posterior neural markers in noggininjected
animal explants, while Xwnt8 does not. Animal caps are explanted at blastula stage 9, and RNA is extracted at midgastrula stage
11 or neurula stage 17. The amounts of RNA used for injection are 50 pg Xwnt7B, 50 pg Xwnt8, and 200 pg noggin, in isolation or
combination (lanes 2 to 6, respectively). Lane 1 shows control uninjected explants, while whole embryo controls with or without reverse
transcriptase (RT) are shown in lanes 7 and 8. (B) Neural patterning at different doses of Xwnt7B. Noggin RNA (0.2 ng) is coinjected with
10, 50, 100, and 500 pg of Xwnt7B RNA (lanes 3 to 6). Neural crest markers and Krox20 can be induced at a low dose of Xwnt7B (10 pg),
while other caudal markers cannot be induced efficiently. (C) Differences in caudalizing activity of Xwnt7B and Xwnt3A. Lanes 3 and 4
are animal explants injected with 0.2 ng noggin and 50 pg Xwnt3A or 50 pg Xwnt7B, respectively. Xwnt3A is a stronger caudalizing factor
than Xwnt7B. (D) Xwnt7B induces neural crest markers in dissociated animal caps. Lane 1 is uninjected control caps, lane 2 is intact
animal caps injected with 100 pg Xwnt7B RNA, and lanes 3 (uninjected) and 4 (injected with 100 pg Xwnt7B RNA) are explants dissociated
in calcium- and magnesium-free medium for 4 h before reaggregation and incubation to midneurula stage.
FIG. 3. Expansion of expression domain of neural crest marker Xtwist by Xwnt7B in vivo. Albino embryos are injected with 50 pg
Xwnt7B RNA into the animal pole of one of the blastomeres at the two-cell stage. Embryos are incubated to midneurula stages 16 to 17
before the expression of neural crest marker Xtwist is examined by in situ hybridization. The top panel shows uninjected control embryos,
while Xwnt7B-injected embryos are shown in the bottom panel. Injected embryos show the expanded expression domain of Xtwist
extending both ventrally and posteriorly from the original location.
