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Abstract Xom (also known as Xvent-2) is a homeobox-containing gene expressed throughout the early gastrula of the Xenopus embryo with the exception of the organizer. Activation of Xom is an immediate-early response to BMP signaling, and overexpression of Xom, like overexpression of BMP family members, causes ventralization of the embryo. In this paper we first show that Xom is a transcriptional repressor and we then define its preferred DNA-binding site. Overexpression of wild-type Xom and a dominant-negative form suggests that Xom functions by repressing transcription of goosecoid, and analysis of the goosecoid promoter reveals a site which is required for Xom-mediated repression of goosecoid promoter reporter constructs. Together, these results suggest that Xom causes down-regulation of goosecoid in a direct fashion and that this accounts, at least in part, for the ability of Xom to cause ventralization of the Xenopus embryo.
FIG. 2. Schematic representation of the consensus binding sequence derived from a PCR-based target site selection analysis (Pollock and
Treisman, 1990) performed with in vitro translated Xom protein. Sequences containing two (or more) TAAT motifs in antiparallel
orientation (75% of the 111 sequences analyzed) were aligned with reference to the TAAT motifs. The percentage frequency with which
a nucleotide (G, A, T, or C) appeared in the same position was scored. Of these sequences, most had six or seven nucleotides separating the
two TAAT antiparallel motifs (30 and 25%, respectively). A consensus binding sequence was established which contains two antiparallel
core motifs separated by six nucleotides and flanked by the indicated preferred nucleotides.
FIG. 5. Overexpression of Xom causes down-regulation of goosecoid expression while a XomVP16 construct causes ectopic activation. (A) Goosecoid expression in control embryos at early gastrula stage 10.5. (B) Goosecoid expression in embryos injected in both blastomeres of the two-cell stage embryo with 4 ng XomA RNA. Note down-regulation of goosecoid expression. (C, D) Two adjacent blastomeres of Xenopus embryos at the four-cell stage embryos were coinjected with 4 ng of XomVP16 RNA together with fluoresceinextran lineage tracer. They were analyzed at early gastrula stage 10.5 by whole mount in situ hybridization to reveal goosecoid expression and whole mount antibody staining to show fluoresceinextran. (C) Vegetal view of an injected embryo with dorsal to the top. Expression of goosecoid in the injected side of the embryo is greatly expanded. (D) Ventral view of another injected embryo with the animal pole to the top. Ectopic expression of goosecoid occurs ventrally and laterally in the marginal zone and extends into the animal pole region. Ectopic expression of goosecoid was observed in all 15 embryos injected with RNA encoding XomVP16. Control embryos injected only with the fluoresceinextran lineage marker had normal goosecoid expression and resembled embryos shown in (A).