Distinct expression profiles of transcriptional coactivators for thyroid hormone receptors during Xenopus laevis metamorphosis

The biological effects of thyroid hormone (T3) are mediated by the thyroid hormone receptor (TR). Amphibian metamorphosis is one of the most dramatic processes that are dependent on T3. T3 regulates a series of orchestrated developmental changes, which ultimately result in the conversion of an aquatic herbivorous tadpole to a terrestrial carnivorous frog. T3 is presumed to bind to TRs, which in turn recruit coactivators, leading to gene activation. The best-studied coactivators belong to the p160 or SRC family. Members of this family include SRC1/NCoA-1, SRC2/TIF2/GRIP1, and SRC3/pCIP/ACTR/AIB-1/RAC-3/TRAM-1. These SRCs interact directly with liganded TR and function as adapter molecules to recruit other coactivators such as p300/CBP. Here, we studied the expression patterns of these coactivators during various stages of development. Amongst the coactivators cloned in Xenopus laevis, SRC3 was found to be dramatically upregulated during natural and T3-induced metamorphosis, and SRC2 and p300 are expressed throughout postembryonic development with little change in their expression levels. These results support the view that these coactivators participate in gene regulation by TR during metamorphosis.     

Distinct expression profiles of transcriptional coactivators for thyroid hormone receptors during Xenopus laevis metamorphosis

The biological effects of thyroid hormone (T3) are mediated by the thyroid hormone receptor (TR). Amphibian metamorphosis is one of the most dramatic processes that are dependent on T3. T3 regulates a series of orchestrated developmental changes, which ultimately result in the conversion of an aquatic herbivorous tadpole to a terrestrial carnivorous frog. T3 is presumed to bind to TRs, which in turn recruit coactivators, leading to gene activation. The best-studied coactivators belong to the p160 or SRC family. Members of this family include SRC1/ NCoA-1, SRC2/TIF2/GRIP1, and SRC3/pCIP/ACTR/AIB-l/RAC-3/TRAM-1. These SRCs interact directly with liganded TR and function as adapter molecules to recruit other coactivators such as p300/CBP. Here, we studied the expression patterns of these coactivators during various stages of development. Amongst the coactivators cloned in Xenopus laevis, SRC3 was found to be dramatically upregulated during natural and T3-induced metamorphosis, and SRC2 and p300 are express     

Some effects of X-irradiation on the activity of the thyroid and pituitary glands of Xenopus laevis (Amphibia) during metamorphosis

A histological study of irradiated thyroid and pituitary glands of metamorphosing Xenopus tadpoles was made. Both glands shrink, reaching a minimum nine hours after irradiation and recover by 24 hours. It is suggested that sudden release of hormones may explain a reported hastening of metamorphosis in irradiated tadpoles.  

Summary:

Xenopus larvae received 2500 rads of whole body X-rays.
Nine hours after irradiation both pituitary and thyroid volumes were significantly less than the controls. From 24 hours after, the volumes had returned to control levels.
The mitotic incidence in irradiated thyroids fell within five hours to one tenth of the control value and remained suppressed for at least three days.     

The distribution of E-cadherin during Xenopus laevis development

A vast amount of experimental evidence suggests that cell surface molecules involved in cell-to-cell and/or cell-to-substrate interactions participate in the control of basic events in morphogenesis. E-cadherin is a cell adhesion molecule directly implicated in the control of Ca2(+)-dependent interactions between epithelial cells. We report here the patterns of expression of E-cadherin in developmental stages of Xenopus laevis ranging from early embryo to adult using immunofluorescence microscopy. Although its distribution shares some similarities with those of L-CAM in the chicken and E-cadherin/Uvomorulin in the mouse, the distribution of E-cadherin in Xenopus presents several peculiar and unique features. In early stages of Xenopus development, E-cadherin is not expressed. The molecule is first detectable in the ectoderm of late gastrulas (stage 13-13.5 NF). At this time both the external and the sensory layer of the nonneural ectoderm accumulate high levels of E-cadherin while the ectoderm overlying the neural plate and regions of the involuting marginal zone (IMZ) not yet internalized by the movements of gastrulation are E-cadherin-negative. Unlike most other species, endodermal cells express no or very low levels of E-cadherin up to stage 20 NF. Endodermal cells become strongly E-cadherin-positive only when a well-differentiated epithelium forms in the gut. No mesodermal structures are stained during early development. In the placodes, in contrast to other species, E-cadherin disappears very rapidly after placode thickening. During further embryonic development E-cadherin is present in the skin, the gut epithelium, the pancreas, many monostratified epithelia and most glands. Hepatocytes are stained weakly while most other tissues, including the pronephros, are negative. In the mesonephros, the Wolffian duct and some tubules are positive. During metamorphosis a profound restructuring of the body plan takes place under the control of thyroid hormones, which involves the degeneration and subsequent regeneration of several tissues such as the skin and the gut. All newly formed epithelia express high levels of E-cadherin. Surprisingly, degenerating epithelia of both skin and intestine maintain high levels of the protein even after starting to become disorganized and to degenerate. In the adult, staining is strong in the skin, the glands, the lungs, the gut epithelium and the pancreas, weak in the liver and absent from most other tissues. Our results show that the expression of E-cadherin in Xenopus is strongly correlated with the appearance of differentiated epithelia.     

Hypothalamic neurosecretion and metamorphosis in Xenopus laevis

Summary Tadpoles of Xenopus laevis were treated with propylthiouracil from the second half of prometamorphosis. Sagittal sections of the head region were stained a.o. with pseudoisocyanine. The goitrogen caused a degranulation of neurosecretory cells in the dorsal part of the preoptic region of the hypothalamus, suppressed the development of ventral neurosecretory cells and of the outer zone of the median eminence, stimulated the thyrotropic cells in the adenohypophysis, caused a hypertrophy of the thyroids, and impaired metamorphosis. Returning the animals to tap water had reciprocal effects and restored the normal activity of the hypothalamus, adenohypophysis and thyroid glands. It is concluded that thyroid hormones exert a morphogenetic influence upon hypothalamic centres and the outer zone of the median eminence and that a negative feed back relation exists between the thyroids on the one hand and the dorsal neurosecretory cells and the thyrotropic cells on the other.The author thanks Prof. Dr. P. G. W. J. van Oordt for his active interest and helpful advice, and Miss Tineke Aafjes for technical assistance.     

Hypothalamic neurosecretion and metamorphosis in Xenopus laevis

Summary Normal and propylthiouracil (PTU) treated Xenopus laevis tadpoles were fixed during all stages of metamorphosis and sagittal sections were stained with aldehyde fuchsin (AF) or pseudoisocyanine (PIC). Whereas AF positive neurosecretory material could only be demonstrated in the preoptic nucleus from late prometamorphosis, increasing amounts of PIC positive material were found in cells of the dorsal part of the preoptic region from early premetamorphosis. The development of these cells correlated with that of the thyrotropic cells and the thyroids. Likewise, signs of hyperactivity in thyroids and thyrotropic cells of PTU treated larvae were accompanied by a depletion of the dorsal PIC positive cells. In the ventral preoptic region PIC positive cells developed from late prometamorphosis in control larvae, but failed to do so in PTU treated animals. It is argued that the differentiation of the PIC positive cells is largely dependent on thyroid hormones; that the dorsal PIC positive cells may produce a thyrotropin releasing factor; and that the function of these dorsal cells is inhibited by thyroid hormones.The authors thank Dr. P. G. W. J. van Oordt for his active interest and helpful advices, Miss Tineke Aafjes for technical assistance and Mr. H. van Kooten for making the photographs.     

Asymmetric growth and development of the Xenopus laevis retina during metamorphosis is controlled by type III deiodinase

During the metamorphosis of the Xenopus laevis retina, thyroid hormone (TH) preferentially induces ventral ciliary marginal zone (CMZ) cells to both increase their proliferation and give rise to ipsilaterally projecting ganglion cells. Here we show that dorsal CMZ cells express type III deiodinase (D3), an enzyme that inactivates TH. The dorsal CMZ cells can be induced to proliferate if deiodinase activity is inhibited. D3 or dominant-negative thyroid hormone receptor transgenes inhibit both TH-induced proliferation of the ventral CMZ cells and the formation of the ipsilateral projection. Thus, the localized expression of D3 in the dorsal CMZ cells accounts for the asymmetric growth of the frog retina.     

Comparison of induction during development between Xenopus tropicalis and Xenopus laevis

Several in vitro systems exist for the induction of animal caps using growth factors such as activin. In this paper, we compared the competence of activin-treated animal cap cells dissected from the late blastulae of Xenopus tropicalis and Xenopus laevis. The resultant tissue explants from both species differentiated into mesodermal and endodermal tissues in a dose-dependent manner. In addition, RT-PCR analysis revealed that organizer and mesoderm markers were expressed in a similar temporal and dose-dependent manner in tissues from both organisms. These results indicate that animal cap cells from Xenopus tropicalis have the same competence in response to activin as those from Xenopus laevis.     

Ultrastructural changes in the intestinal connective tissue of Xenopus laevis during metamorphosis

Ultrastructural changes in the intestinal connective tissue of Xenopus laevis during metamorphosis have been studied. Throughout the larval period to stage 60, the connective tissue consists of a few immature fibroblasts surrounded by a sparse extracellular matrix: few collagen fibrils are visible except close to the thin basal lamina. At the beginning of the transition from larval to adult epithelial form around stage 60, extensive changes are observed in connective tissue. The cells become more numerous and different types appear as the collagen fibrils increase in number and density. Through gaps in the thickened and extensively folded basal lamina, frequent contacts between epithelial and connective tissue cells are established. Thereafter, with the progression of fold formation, the connective tissue cells become oriented according to their position relative to the fold structure. The basal lamina beneath the adult epithelium becomes thin after stage 62, while that beneath the larval epithelium remains thick. Upon the completion of metamorphosis, the connective tissue consists mainly of typical fibroblasts with definite orientation and numerous collagen fibrils. These observations indicate that developmental changes in the connective tissue, especially in the region close to the epithelium, are closely related spatiotemporarily to the transition from the larval to the adult epithelial form. This suggests that tissue interactions between the connective tissue and the epithelium play important roles in controlling the epithelial degeneration, proliferation, and differentiation during metamorphic climax.     

Spatio-temporal expression profile of stem cell-associated gene LGR5 in the intestine during thyroid hormone-dependent metamorphosis in Xenopus laevis

Background

The intestinal epithelium undergoes constant self-renewal throughout adult life across vertebrates. This is accomplished through the proliferation and subsequent differentiation of the adult stem cells. This self-renewal system is established in the so-called postembryonic developmental period in mammals when endogenous thyroid hormone (T3) levels are high.

Methodology/Principal Findings

The T3-dependent metamorphosis in anurans like Xenopus laevis resembles the mammalian postembryonic development and offers a unique opportunity to study how the adult stem cells are developed. The tadpole intestine is predominantly a monolayer of larval epithelial cells. During metamorphosis, the larval epithelial cells undergo apoptosis and, concurrently, adult epithelial stem/progenitor cells develop de novo, rapidly proliferate, and then differentiate to establish a trough-crest axis of the epithelial fold, resembling the crypt-villus axis in the adult mammalian intestine. The leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) is a well-established stem cell marker in the adult mouse intestinal crypt. Here we have cloned and analyzed the spatiotemporal expression profile of LGR5 gene during frog metamorphosis. We show that the two duplicated LGR5 genes in Xenopus laevis and the LGR5 gene in Xenopus tropicalis are highly homologous to the LGR5 in other vertebrates. The expression of LGR5 is induced in the limb, tail, and intestine by T3 during metamorphosis. More importantly, LGR5 mRNA is localized to the developing adult epithelial stem cells of the intestine.

Conclusions/Significance

These results suggest that LGR5-expressing cells are the stem/progenitor cells of the adult intestine and that LGR5 plays a role in the development and/or maintenance of the adult intestinal stem cells during postembryonic development in vertebrates.     

Short-term effects of thyroid hormones during development: Focus on signal transduction

Extranuclear or nongenomic effects of thyroid hormones are mediated by receptors located at the plasma membrane or inside cells, and are independent of protein synthesis. Recently the αVβ3 integrin was identified as a cell membrane receptor for thyroid hormones, and a wide variety of nongenomic effects have now been shown to be induced through binding of thyroid hormones to this receptor. However, also other thyroid hormone receptors can produce nongenomic effects, including the cytoplasmic TRα and TRβ receptors and probably also a G protein-coupled membrane receptor, and increasing importance is now given to thyroid hormone metabolites like 3,5-diiodothyronine and reverse T₃ that can mimick some nongenomic effects of T₃ and T₄. Signal transduction from the αVβ3 integrin may proceed through at least three independent pathways (protein kinase C, Src or mitogen-activated kinases) but the details are still unknown. Thyroid hormones induce nongenomic effects on at least three important Na⁺-dependent transport systems, the Na⁺/K⁺-ATPase, the Na⁺/H⁺ exchanger, and amino acid transport System A, leading to a mitogenic response in embryo cells; but modulation of the same transport systems may have different roles in other cells and at different developmental stages. It seems that thyroid hormones in many cases can modulate nongenomically the same targets affected by the nuclear receptors through long-term mechanisms. Recent results on nongenomic effects confirm the old theory that the primary role of thyroid hormones is to keep the steady-state level of functioning of the cell, but more and more mechanisms are discovered by which this goal can be achieved.     

Cell intercalation during notochord development in Xenopus laevis

Morphometric data from scanning electron micrographs (SEM) of cells in intact embryos and high-resolution time-lapse recordings of cell behavior in cultured explants were used to analyze the cellular events underlying the morphogenesis of the notochord during gastrulation and neurulation of Xenopus laevis. The notochord becomes longer, narrower, and thicker as it changes its shape and arrangement and as more cells are added at the posterior end. The events of notochord development fall into three phases. In the first phase, occurring in the late gastrula, the cells of the notochord become distinct from those of the somitic mesoderm on either side. Boundaries form between the two tissues, as motile activity at the boundary is replaced by stabilizing lamelliform protrusions in the plane of the boundary. In the second phase, spanning the late gastrula and early neurula, cell intercalation causes the notochord to narrow, thicken, and lengthen. Its cells elongate and align mediolaterally as they rearrange. Both protrusive activity and its effectiveness are biased: the anterioposterior (AP) margins of the cells advance and retract but produce much less translocation than the more active left and right ends. The cell surfaces composing the lateral boundaries of the notochord remain inactive. In the last phase, lasting from the mid- to late neurula stage, the increasingly flattened cells spread at all their interior margins, transforming the notochord into a cylindrical structure resembling a stack of pizza slices. The notochord is also lengthened by the addition of cells to its posterior end from the circumblastoporal ring of mesoderm. Our results show that directional cell movements underlie cell intercalation and raise specific questions about the cell polarity, contact behavior, and mechanics underlying these movements. They also demonstrate that the notochord is built by several distinct but carefully coordinated processes, each working within a well-defined geometric and mechanical environment.