Dorsal induction from dorsal vegetal cells in Xenopus occurs after mid-blastula transition

In a screening for activin-responsive genes, we isolated a Xenopus lefty/antivin-related gene, called Xantivin (Xatv). In the animal cap assay, the expression of Xatv was induced by activin signaling, and in the embryo, by nodal-related genes. Overexpression of Xatv in the marginal zone caused suppression of mesoderm formation and gastrulation defects, and inhibited the secondary axis formation induced by Xnr1 and Xactivin, suggesting that Xatv acted as a feedback inhibitor of activin signaling. However, in the animal cap, Xatv failed to antagonize Xnr1 and Xactivin. This result suggested that Xatv has different responses in the marginal zone and in the animal region, and antagonizes to a higher degree activin signaling in the marginal zone.     

The Drosophila zygotic lethal gene shuttle craft is required maternally for proper embryonic development

 The Drosophila gene shuttle craft (stc) is expressed zygotically in the embryonic central nervous system (CNS) where it is required to maintain the proper morphology of motoneuronal axon nerve routes following their migration from the ventral cord. Here, we report that a prominent maternal source of STC protein is also present throughout both oogenesis and embryogenesis. To determine whether this maternal component is required in the ovary and/or embryo, we used the Drosophila autosomal dominant female sterile technique to generate germ-line clones that lacked the stc maternal function. Our results demonstrate that a maternally derived source of STC protein is required during embryogenesis but not oogenesis. In contrast to the zygotic phenotype, the primary defect in embryos derived from stc germ-line clones affects segmentation by causing disruptions and deletions in distinct thoracic (T1–T3) and abdominal (A4–A8) segments. These localized defects are responsible for additional phenotypes observed later in development which include gaps in the ventral nerve cord and deletions of denticle belts in the cuticle. An additional phenotype occurring in all other neuromeric segments consists of the misguided migration of motoneuronal axons as they project out of the ventral nerve cord. Thus, the stc zygotic function is required later in development and cannot correct the segmentation and subsequent CNS abnormalities associated with loss of its earlier acting maternally derived activity. Received: 12 March 1998 / Accepted: 9 April 1998     

Xenopus TACC3/maskin is not required for microtubule stability but is required for anchoring microtubules at the centrosome

Members of the transforming acidic coiled coil (TACC) protein family are emerging as important mitotic spindle assembly proteins in a variety of organisms. The molecular details of how TACC proteins function are unknown, but TACC proteins have been proposed to recruit microtubule-stabilizing proteins of the tumor overexpressed gene (TOG) family to the centrosome and to facilitate their loading onto newly emerging microtubules. Using Xenopus egg extracts and in vitro assays, we show that the Xenopus TACC protein maskin is required for centrosome function beyond recruiting the Xenopus TOG protein XMAP215. The conserved C-terminal TACC domain of maskin is both necessary and sufficient to restore centrosome function in maskin-depleted extracts, and we provide evidence that the N terminus of maskin inhibits the function of the TACC domain. Time-lapse video microscopy reveals that microtubule dynamics in Xenopus egg extracts are unaffected by maskin depletion. Our results provide direct experimental evidence of a role for maskin in centrosome function and suggest that maskin is required for microtubule anchoring at the centrosome.     

Zygotic Wnt activity is required for Brachyury expression in the early Xenopus laevis embryo

The canonical, beta-catenin-dependent Wnt pathway is a crucial player in the early events of Xenopus development. Dorsal axis formation and mesoderm patterning are accepted effects of this pathway, but the regulation of expression of genes involved in mesoderm specification is not. This conclusion is based largely on the inability of the Wnt pathway to induce mesoderm in animal cap explants. Using injections of inhibitors of canonical Wnt signaling, we demonstrate that expression of the general mesodermal marker Brachyury (Xbra) requires a zygotic, ligand-dependent Wnt activity throughout the marginal zone. Analysis of the Xbra promoter reveals that putative TCF-binding sites mediate Wnt activation, the first sites in this well-studied promoter to which an activation role can be ascribed. However, established mesoderm inducers like eFGF and activin can bypass the Wnt requirement for Xbra expression. Another mesoderm promoting factor, VegT, activates Xbra in a Wnt-dependent manner. We also show that the activin/nodal signaling is necessary for ectopic Xbra induction by the Wnt pathway, but not by VegT. Our data significantly change the understanding of Brachyury regulation in Xenopus, implying the existence of an unknown zygotic Wnt ligand in Spemann's organizer. Since Brachyury is considered to have a major role in mesoderm formation, it is possible that Wnts might play a role in mesoderm specification, in addition to patterning.     

A novel gene, BENI is required for the convergent extension during Xenopus laevis gastrulation

Activin-like signaling plays an important role in early embryogenesis. Activin A, a TGF-beta family protein, induces mesodermal/endodermal tissues in animal cap assays. In a screen for genes expressed early after treatment with activin A, we isolated a novel gene, denoted as BENI (Brachyury Expression Nuclear Inhibitor). The BENI protein has a conserved domain at the N-terminus that contains a nuclear localization signal (NLS), and two other NLSs in the C-terminal domain. BENI mRNA was localized to the animal hemisphere at the gastrula stages and to ectoderm except for neural regions at stage 17; expression persisted until the tadpole stage. The overexpression of BENI caused gastrulation defects and inhibition of elongation of activin-treated animal caps with reduction of Xbra expression. Moreover, whole-mount in situ hybridization revealed reduced expression of Xbra in BENI mRNA-injected regions of gastrula embryos. Functional knockdown of BENI using an antisense morpholino oligonucleotide also resulted in an abnormal phenotype of embryos curling to the dorsal side, and excessive elongation of activin-treated animal caps without altered expression of mesodermal markers. These results suggested that BENI expression is regulated by activin-like signaling, and that this regulation is crucial for Xbra expression.     

Xenopus Dab2 is required for embryonic angiogenesis

Background  

The molecular mechanisms governing the formation of the embryonic vascular system remain poorly understood. Here, we show that Disabled-2 (Dab2), a cytosolic adaptor protein, has a pivotal role in the blood vessel formation in Xenopus early embryogenesis.     

The maternal gene product D7 is not required for early Xenopus development.

The Xenopus D7 gene codes for a novel protein whose expression is restricted to early development. D7 protein is synthesized for the first time during oocyte maturation (1988, Genes Dev. 2, 1296-1306). Injection of D7 RNA into the full-grown oocyte and its subsequent translation into D7 protein neither induced oocyte maturation nor affected the kinetics of hormone-induced maturation. Overexpression of D7 protein by 20-fold in the early Xenopus embryo by injection of D7 RNA into fertilized eggs did not affect subsequent development. Oocytes specifically lacking D7 mRNA were generated by oligodeoxynucleotide-mediated RNA destruction within the oocyte. Unfertilized eggs generated from such oocytes lacked detectable D7 protein, but nevertheless could be activated and fertilized. Embryos generated from such eggs, estimated to contain less than 5% of wildtype levels of D7 protein, developed normally up to the tailbud stage. Thus the D7 protein, the product of a maternal mRNA that is under strict translational repression in oocytes, appears not to be required for oocyte maturation, activation, fertilization or early embryonic development in Xenopus.     

Notch is required for outgrowth of the Xenopus tail bud

It has previously been shown that Notch, Delta and Lunatic Fringe are expressed together in the leading edge of the tail bud of the Xenopus embryo prior to outgrowth (Beck and Slack, 1998). It has also been shown that ectopic expression of a constitutive form of Notch, Notch-ICD, will provoke ectopic tail formation (Beck and Slack, 1999). Here we show that inhibition of Notch activity in vivo prevents outgrowth of the tail bud. This is achieved using inhibitors of the protease that carries out the ligand-induced intramembranous cleavage of Notch. Other protease inhibitors that do not inhibit Notch cleavage do not affect tail outgrowth.     

MBD2 is required for correct spatial gene expression in the gut

Gene expression in the gut is segmentally regulated, but little is known of the molecular origin of patterning. Analysis of gene expression in colons from mice lacking the methyl-CpG binding repressor MBD2 revealed frequent activation of genes that are normally only expressed in the exocrine pancreas and duodenum. Reduced DNA methylation activated the same gene set in the colon. No significant differences in DNA methylation between the colon and duodenum were detected, but MBD2 was significantly more abundant in the colon. The relevance of MBD2 concentration was tested in a human colon cancer cell line. Depletion of MBD2 was again found to activate exocrine pancreatic genes. Gene activation in this cell culture model was accompanied by loss of promoter-bound MBD2 and increased histone acetylation. The results suggest that modulation of MBD2 during gut development establishes a region-specific gene expression pattern that is essential for establishing correct segmental character.