CAP1 expression is developmentally regulated in Xenopus

We have cloned and characterized a Xenopus member of the cyclase associated protein (CAP) gene family. xCAP1 is expressed as a maternal transcript, but is up-regulated prior to gastrulation and subsequently localizes to head mesenchyme, lens, otic vesicle, and trunk mesoderm including the pronephros. At different stages, the gone also appears to differentiate surface from deep (sensorial) ectoderm. As in Drosophila, Xenopus CAP1 is expressed in the developing eye, specifically in the differentiating lens. However, in distinction to Drosophila, Xenopus CAP1 does not express in periodically arrayed neural bands. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved

Conservation of Pitx1 expression during amphibian limb morphogenesis

In contrast to the pattern of limb emergence in mammals, chicks, and the newt N. viridescens, embryos such as Xenopus laevis and Eleutherodactylus coqui initiate pelvic limb buds before they develop pectoral ones. We studied the expression of Pitx1 in X. laevis and E. coqui to determine if this paired-like homeodomain transcription factor directs differentiation specifically of the hindlinib, or if it directs the second pair of limbs to form, namely the forelimbs. We also undertook to determine if embryonic expression patterns were recapitulated during the regeneration of an amputated limb bud. Pitx1 is expressed in hindlimbs in both X. laevis and E. coqui, and expression is similar in both developing and regenerating limb buds. Expression in hindlimbs is restricted to regions of proliferating mesenchyme

xArx2: An Aristaless Homolog That Regulates Brain Regionalization During Development in Xenopus laevis

The aristaless-related gene, Arx, plays a fundamental role in patterning the brain in humans and mice. Arx mutants exhibit lissencephaly among other anomalies. We have cloned a Xenopus aristaless homolog that appears to define specific regions of the developing forebrain. xArx2 is transcribed in blastula through neurula stages, and comes to be restricted to the ventra and lateral telencephalon, lateral diencephalon, neural floor plate of the anterior spinal cord, and somites. In this respect, Arx2 expresses in regions similar to Arx with the exception of the somites. Overexpression enlarges the telencephalon, and interference by means of antisense morpholino-mediated translation knockdown reduces growth of this area. Overexpression and inhibition studies demonstrate that misregulation of xArx2 imposes dire consequences upon patterns of differentiation not only in the forebrain where the gene normally expresses, but also in more caudal brain territories and derivatives as well. This suggests that evolutionary changes that expanded Arx-expression from ventral to dorsal prosencephalon might be one of the determinants that marked development and expansion of the telencephalon. genesis 47:19-31, 2009. (C) 2008 Wiley-Liss, Inc

Expression of CAP2 during early Xenopus embryogenesis

We have cloned and characterized a second member of the Xenopus CAP (cyclase associated protein) gene family. xCAP2 demonstrates greater restriction of expression than its homolog, xCAP1, and is differentially expressed throughout early embryogenesis. Although present as a maternal transcript, CAP2 comes to be expressed in the anterior-most mesoderm/endoderm during late gastrulation, in paraxial mesoderm during late neurula stages, and later expresses in lens, cardiac primordia, somites, otic vesicles, retina,and in the optic and craniofacial musculature. The gene is also expressed in the leading edge of myotome

xPitx1 plays a role in specifying cement gland and head during early Xenopus development

Xenopus Pitx1 is a homeobox gene whose family members are structurally and functionary conserved in organisms as diverse as Drosophila, chick, mouse, human, and frog. Present as a maternal transcript, the gene is zygotically expressed during gastrulation in a dorsal streak of cells. This streak restricts to a small circular domain underlying the center of presumptive neural plate. Shortly thereafter, a crescent of expression develops at the border of anterior neural ectoderm, and as the central plate domain diminishes, the crescent coalesces to define the presumptive cement gland. Expression remains high throughout cement gland development, and subsequently expands to include ectodermal cells involved in stomodeal invagination. During early organogenesis, expression ensues in developing eye, posterior lateral mesoderm, and first branchial arch derivatives. Ectopic expression of xPitx1 causes head deformities including enlarged cement gland, ectopic cement glands, and posterior deformities or, in extreme cases, inhibition of recognizable structures posterior to the cement gland. Expression of markers such as XCG-1, xOtx2, xPax6, neural beta tubulin, and xTwist suggest that increases in cement gland and lower mandibular size are likely at the expense of other head tissues. Paradoxically, overexpression is sufficient to partially rescue embryos that are axially perturbed by ultraviolet irradiation or retinoic acid administration. Ectopic expression of xPitx1 in ectodermal explants directly promotes cement gland development as there was no evidence that mesodermal or neural tissue was present in explants. genesis 29:18-90, 2001. (C) 2001 Wiley-Liss, Inc

Adenine Nucleotide Translocase expression undergoes dynamic regulation during gastrulation in Xenopus laevis (Accession # AF231347)

We report the isolation and characterization of the Xenopus homolog to human T1 ANT (adenine nucleotide translocase) The 1290 nucleotide sequence contains initiation and termination signals, and encodes a conceptual protein of 298 amino acids. The sequence shares high amino acid identity with the mammalian adenine translocases. The transcript is present in unfertilized eggs, and it is expressed at higher levels during formation of the antero-posterior dorsal axis in embryos. Although low levels are expressed constitutively except in endodermal cells, ANT expression is dynamically regulated during neurulation. At this stage expression in ectoderm rapidly diminishes as the neural folds form, and then ANT expression increases slightly in mesoderm. At the culmination of neurulation, the neural tube briefly expresses ANT, and thereafter its expression predominates in the somitic mesoderm and also the chordoneural hinge. In addition, ANT expression is particularly high in the prosencephalon, the mesencephalon, the branchial arches, eye, and the otic vesicle. Treatment of embryos with retinoic acid has the effect of diminishing constitutive expression of ANT, but microinjection studies demonstrate that immediate and local repression cannot be induced in dorsal structures

Microarray-based identification of Pitx3 targets during Xenopus embryogenesis

Background: Unexpected phenotypes resulting from morpholino-mediated translational knockdown of Pitx3 in Xenopus laevis required further investigation regarding the genetic networks in which the gene might play a role. Microarray analysis was, therefore, used to assess global transcriptional changes downstream of Pitx3. Results: From the large data set generated, selected candidate genes were confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) and in situ hybridization. Conclusions: We have identified four genes as likely direct targets of Pitx3 action: Pax6, beta Crystallin-b1 (Crybb1), Hes7.1, and Hes4. Four others show equivocal promise worthy of consideration: Vent2, and Ripply2 (aka Ledgerline or Stripy), eFGF and RXRa. We also describe the expression pattern of additional and novel genes that are Pitx3-sensitive but that are unlikely to be direct targets

Lens and Retina Formation Require Expression of Pitx3 in Xenopus Pre-lens Ectoderm

Pitx3 is expressed in tissues fated to contribute to eye development, namely, neurula stage ectoderm and prechordal mesoderm, then presumptive lens ectoderm, placode, and finally lens. Pitx3 overexpression alters lens, optic cup, optic nerve, and diencephalon development. Many of the induced anomalies are attributable to midline deficits; however, as assessed by molecular markers, ectopic Pitx3 appears to temporarily enlarge the lens field. These changes are usually insufficient to generate either ectopic lenses to enlarge the eye that eventually differentiates. Conversely, use of a repressor chimera or of antisense morpholinos alters early expression of marker genes, and later inhibits lens development, thereby abrogating retinal induction. Reciprocal grafting experiments using wild-type and morpholino-treated tissues demonstrate that Pitx3 expression in the presumptive lens ectoderm is required for lens formation. Contradictory to recent assertions that retina can form in the absence of a lens, the expression of Pitx3 in the presumptive lens ectoderm. is critical for retina development

Xenopus pitx3 target genes lhx1 and xnr5 are identified using a novel three-fluor flow cytometry–based analysis of promoter activation and repression

Background: Pitx3 plays a well understood role in directing development of lens, muscle fiber, and dopaminergic neurons; however, in Xenopus laevis, it may also play a role in early gastrulation and somitogenesis. Potential downstream targets of pitx3 possess multiple binding motifs that would not be readily accessible by conventional promoter analysis. Results: We isolated and characterized pitx3 target genes lhx1 and xnr5 using a novel three-fluor flow cytometry tool that was designed to dissect promoters with multiple binding sites for the same transcription factor. This approach was calibrated using a known pitx3 target gene, Tyrosine hydroxylase. Conclusions: We demonstrate how flow cytometry can be used to detect gene regulatory changes with exquisite precision on a cell-by-cell basis, and establish that in HEK293 cells, pitx3 directly activates lhx1 and represses xnr5

Mouth development

WIREs Developmental Biology published by Wiley Periodicals, Inc. A mouth is present in all animals, and comprises an opening from the outside into the oral cavity and the beginnings of the digestive tract to allow eating. This review focuses on the earliest steps in mouth formation. In the first half, we conclude that the mouth arose once during evolution. In all animals, the mouth forms from ectoderm and endoderm. A direct association of oral ectoderm and digestive endoderm is present even in triploblastic animals, and in chordates, this region is known as the extreme anterior domain (EAD). Further support for a single origin of the mouth is a conserved set of genes that form a ‘mouth gene program’ including foxA and otx2. In the second half of this review, we discuss steps involved in vertebrate mouth formation, using the frog Xenopus as a model. The vertebrate mouth derives from oral ectoderm from the anterior neural ridge, pharyngeal endoderm and cranial neural crest (NC). Vertebrates form a mouth by breaking through the body covering in a precise sequence including specification of EAD ectoderm and endoderm as well as NC, formation of a ‘pre-mouth array,’ basement membrane dissolution, stomodeum formation, and buccopharyngeal membrane perforation. In Xenopus, the EAD is also a craniofacial organizer that guides NC, while reciprocally, the NC signals to the EAD to elicit its morphogenesis into a pre-mouth array. Human mouth anomalies are prevalent and are affected by genetic and environmental factors, with understanding guided in part by use of animal models.National Institute of Dental and Craniofacial Research (U.S.) (Grant RO1 DE021109