[show abstract][hide abstract] ABSTRACT: During early embryogenesis, the neural plate is specified along the anterior-posterior (AP) axis by the action of graded patterning signals. In particular, the attenuation of canonical Wnt signals plays a central role in the determination of the anterior brain region. Here, we show that the extracellular matrix (ECM) protein Del1, expressed in the anterior neural plate, is essential for forebrain development in the Xenopus embryo. Overexpression of Del1 expands the forebrain domain and promotes the formation of head structures, such as the eye, in a Chordin-induced secondary axis. Conversely, the inhibition of Del1 function by a morpholino oligonucleotide (MO) represses forebrain development. Del1 also augments the expression of forebrain markers in neuralized animal cap cells, whereas Del1-MO suppresses them. We previously reported that Del1 interferes with BMP signaling in the dorsal-ventral patterning of the gastrula marginal zone. By contrast, we demonstrate here that Del1 function in AP neural patterning is mediated mainly by the inhibition of canonical Wnt signaling. Wnt-induced posteriorization of the neural plate is counteracted by Del1, and the Del1-MO phenotype (posteriorization) is reversed by Dkk1. Topflash reporter assays show that Del1 suppresses luciferase activities induced by Wnt1 and beta-catenin. This inhibitory effect of Del1 on canonical Wnt signaling, but not on BMP signaling, requires the Ror2 pathway, which is implicated in non-canonical Wnt signaling. These findings indicate that the ECM protein Del1 promotes forebrain development by creating a local environment that attenuates the cellular response to posteriorizing Wnt signals via a unique pathway.
Development 10/2010; 137(19):3293-302. · 6.60 Impact Factor
[show abstract][hide abstract] ABSTRACT: Here, we demonstrate self-organized formation of apico-basally polarized cortical tissues from ESCs using an efficient three-dimensional aggregation culture (SFEBq culture). The generated cortical neurons are functional, transplantable, and capable of forming proper long-range connections in vivo and in vitro. The regional identity of the generated pallial tissues can be selectively controlled (into olfactory bulb, rostral and caudal cortices, hem, and choroid plexus) by secreted patterning factors such as Fgf, Wnt, and BMP. In addition, the in vivo-mimicking birth order of distinct cortical neurons permits the selective generation of particular layer-specific neurons by timed induction of cell-cycle exit. Importantly, cortical tissues generated from mouse and human ESCs form a self-organized structure that includes four distinct zones (ventricular, early and late cortical-plate, and Cajal-Retzius cell zones) along the apico-basal direction. Thus, spatial and temporal aspects of early corticogenesis are recapitulated and can be manipulated in this ESC culture.
[show abstract][hide abstract] ABSTRACT: We show here that a secreted EGF-Discoidin-domain protein, Xenopus Del1 (xDel1), is an essential factor for dorsal development in the early Xenopus embryo. Knockdown of the xDel1 function causes obvious ventralization of the embryo. Conversely, overexpression of xDel1 expands dorsal-marker expression and suppresses ventral-marker expression in the gastrula embryo. Forced expression of xDel1 dorsalizes ventral marginal zone explants, whereas it weakly induces neural differentiation but not mesodermal differentiation in animal caps. The dorsalizing activity of xDel1 is dependent on the Discoidin domains and not on the RGD motif (which is implicated in its angiogenic activity) or EGF repeats. Luciferase assays show that xDel1 attenuates BMP-signaling reporter activity by interfering with the pathway downstream of the BMP receptor. Thus, xDel1 functions as a unique extracellular regulatory factor of DV patterning in early vertebrate embryogenesis.
[show abstract][hide abstract] ABSTRACT: In Xenopus, an asymmetric distribution of Wnt activity that follows cortical rotation in the fertilized egg leads to the dorsal-ventral (DV) axis establishment. However, how a clear DV polarity develops from the initial difference in Wnt activity still remains elusive. We report here that the Teashirt-class Zn-finger factor XTsh3 plays an essential role in dorsal determination by enhancing canonical Wnt signaling. Knockdown of the XTsh3 function causes ventralization in the Xenopus embryo. Both in vivo and in vitro studies show that XTsh3 substantially enhances Wnt signaling activity in a beta-catenin-dependent manner. XTsh3 cooperatively promotes the formation of a secondary axis on the ventral side when combined with weak Wnt activity, whereas XTsh3 alone has little axis-inducing ability. Furthermore, Wnt1 requires XTsh3 for its dorsalizing activity in vivo. Immunostaining and protein analyses indicate that XTsh3 is a nuclear protein that physically associates with beta-catenin and efficiently increases the level of beta-catenin in the nucleus. We discuss the role of XTsh3 as an essential amplifying factor of canonical Wnt signaling in embryonic dorsal determination.
The EMBO Journal 06/2007; 26(9):2350-60. · 9.82 Impact Factor
[show abstract][hide abstract] ABSTRACT: Here, we report in vitro generation of Math1+ cerebellar granule cell precursors and Purkinje cells from ES cells by using soluble patterning signals. When neural progenitors induced from ES cells in a serum-free suspension culture are subsequently treated with BMP4 and Wnt3a, a significant proportion of these neural cells become Math1+. The induced Math1+ cells are mitotically active and express markers characteristic of granule cell precursors (Pax6, Zic1, and Zipro1). After purification by FACS and coculture with postnatal cerebellar neurons, ES cell-derived Math1+ cells exhibit typical features of neurons of the external granule cell layer, including extensive motility and a T-shaped morphology. Interestingly, differentiation of L7+/Calbindin-D28K+ neurons (characteristic of Purkinje cells) is induced under similar culture conditions but exhibits a higher degree of enhancement by Fgf8 rather than by Wnt3a. This is the first report of in vitro recapitulation of early differentiation of cerebellar neurons by using the ES cell system.
[show abstract][hide abstract] ABSTRACT: During gastrulation in Xenopus, the head ectoderm is subdivided into the central nervous system (CNS) anlage (neural plate) and the non-CNS ectoderm (i.e. epidermis, placodes and neural crest). The winged-helix transcription factor Xfoxi1a is one of the earliest markers for the preplacodal region at the mid-neurula stage. Interestingly, before the establishment of the preplacodal region, Xfoxi1a expression is detected in the entire cephalic non-neural ectoderm at the mid- and late gastrula stages. The present study focuses on the role of Xfoxi1a particularly at the gastrula stages. The early Xfoxi1a expression in the anteroventral ectoderm is dependent on Bmp signals and suppressed by Wnt signals. Inhibition of Xfoxi1a activities by injection of antisense oligonucleotides leads to suppression of non-CNS ectodermal markers (e.g. keratin) and expansion of the anterior expression domain of the CNS marker Sox2. Conversely, misexpression of Xfoxi1a suppresses Sox2 and induces keratin in the anterior neural plate. In the animal cap, Xfoxi1a overexpression antagonizes the neuralizing activity of Chordin (Chd). Studies using an inducible Xfoxi1a construct (GR-Xfoxi1a) show that the ventralizing function of Xfoxi1a is confined to the gastrula stage. Thus, Xfoxi1a is an essential regulator of ventral specification of the early head ectoderm during gastrulation.
Development 10/2005; 132(17):3885-94. · 6.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: We have isolated a novel secreted dorsalizing factor of the neural tube, Xenopus Tiarin, which belongs to the olfactomedin-related family. Tiarin expression starts at the late gastrula stage in the nonneural ectoderm adjacent to the anterior neural plate. Overexpression of Tiarin in the embryo causes expansion of dorsal neural markers and suppression of ventral markers. In the eye-forming field, Tiarin overexpression induces the retinal markers and represses optic stalk markers. Tiarin directly dorsalizes neural tissues in the absence of mesodermal tissues and antagonizes the ventralizing activity of Sonic hedghog (Shh). Unlike BMP4, another dorsalizing factor, Tiarin does not display antineuralizing activity on the ectoderm or mesoderm-ventralizing activity. These findings show that Tiarin is a novel patterning signal candidate acting in the specification of the dorsal neural tube.