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Gastrulation movements in Xenopus laevis are becoming increasingly well characterised, however the molecular mechanisms involved are less clear. Active migration of the leading edge mesendoderm across the fibronectin-coated blastocoel roof is necessary for further development of tissues such as headmesoderm, heart, blood and liver. The zinc finger transcription factors GATA4 and GATA6 are expressed in this migratory tissue during gastrulation, but their role here is unknown. This study further characterises the expression of GATA4 and 6 during gastrulation, and investigates their function in migratory behaviour. Gain-of-function experiments with these GATA factors induce cell spreading, polarisation and migration in non-motile presumptive ectoderm cells. Expression of a dominant-interfering form of GATA6, which inhibits transactivation of GATA targets, severely impairs the ability of dorsal leading edge mesendoderm to spread and translocate on fibronectin. Mosaic inhibition of GATA activity indicates that GATA factors function cell autonomously to induce cell spreading and movement in dorsal mesendoderm. Knockdown of specific GATA factors using anti-sense morpholinos indicates that GATA4 and GATA6 both contribute to dorsal mesendoderm migration in vitro. GATA4 and GATA6 are known to be involved in cell-specification of mesoderm and endoderm-derived tissues, but this is the first description of an additional role for these factors in cell migration.
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16949798
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Fig. 1. GATA4 and GATA6 are expressed in migratory leading edge mesendoderm throughout gastrulation. (A–L) In situ hybridisation on embryos just prior to (stage 9), during (stages 10.25 and 12) and at the end of (stage 13) gastrulation. Serial sections were hybridised with probes for GATA6, GATA4 and the mesodermal marker Xbra. (M–O) In situ hybridisation showing coexpression of GATA6 with cerberus and Xhex (markers of leading edge mesendoderm) during gastrulation. The vegetal pole is to the bottom and dorsal is to the right in all panels except J–L, where dorsal is towards the top and anterior is left. Arrows in (A) and (B) denote suprablastoporal endoderm, where punctate staining is visible in a few cells. Arrow in (J) denotes archenteron roof. All sections are sagittal and 10 μm.
Fig. 3. Inhibition of GATA function disrupts gastrulation movements. (A) Induction of Sox17α by mdlGATA6 in animal caps is inhibited by myqGATA6EnR. Relative gene expression in animal caps at sib stage 12 measured by real-time RT-PCR. Fifty picograms of each RNA was injected into animal caps of one-cell embryos. This experiment was repeated twice giving similar results each time. (B) Transactivation of a GATA-driven luciferase reporter gene by endogenous factors in the marginal zone is inhibited by myqGATA6EnR. The marginal zone of each blastomere at the four-cell stage was injected with reporter DNA ±myqGATA6EnR mRNA. Luciferase activity is expressed as fold activation relative to background activity in uninjected embryos. This experiment was repeated twice giving similar results each time. (C and D) Inhibition of GATA function disrupts gastrulation movements and anterior development. (C) myqGATA6EnR-injected embryos display delayed gastrulation, poor head development and shortened anteriorposterior axes. Fluorescent image shows nlsGFP expression in a myqGATA6EnR-injected embryo (see bright-field image above) and demonstrates accurate targeting of the marginal zone. (D) Pooled embryo phenotype counts from three independent experiments. Only predominant defects were scored to avoid multiple counting of the same specimen. The phenotype observed varied between embryo batches, however gastrulation was always delayed and disturbed. (E) Morphology and gene expression in myqGATA6EnR-injected embryos. Following injection of myqGATA6EnR or EnR (control) RNA into each blastomere at the four cell stage, embryos were cultured to stage 11, sagitally sectioned and hybridised with anti-sense probes for the indicated genes. Morphology and gene expression are normal in EnR-injected embryos, but are disrupted by myqGATA6EnR-injection.
