Biomedical subjects

T Pieler

Publications and source records attributed to T Pieler.

At least 19 recordsLinked to original sources

PubMed2001

Molecular cloning and expression analysis of the Hedgehog receptors XPtc1 and XSmo in Xenopus laevis.

Inductive signaling mediated by secreted factors of the Hedgehog (Hh) gene family regulates cellular proliferation and differentiation in many embryonic tissues. Two transmembrane proteins associated in a complex, Patched (Ptc) and Smoothened (Smo), are indispensable for the reception of Hh signals (Cell 86 (1996) 221; Nature 382 (1996) 547; Nature 384 (1996) 176; Nature 384 (1996) 129). Here, we report on the identification of Ptc and Smo homologues from Xenopus and analyze their spatio-temporal expression during embryogenesis. The intracellular response to Hh signals involves upregulation of Ptc transcription (Genes Dev. 10 (1996) 301; J. Biol. Chem. 271 (1996) 12125). In accordance with its putative function as Shh target gene, XPtc1 expression during early stages of Xenopus embryogenesis is detected in mesodermal and neuroectodermal tissues proximal to the notochord, a known expression domain of Shh. Although the expression pattern of XPtc1 was similar to that of other vertebrates, expression domains specific to Xenopus could be detected in the hypochord, dorsal mesencephalon, otic vesicles and pituitary anlage. Unlike other vertebrate Ptc1 homologues, somitic expression of XPtc1 is confined to a central cell layer. In contrast to the tissue-specific expression characteristics of XPtc1, XSmo expression appears to be ubiquitously activated in early embryonic stages but condenses in the terminal regions of the embryo at tailbud stage. In many tissues and organs of the adult, XPtc1 and XSmo are found to display similar expression levels.

Amino Acid Sequence↗

PubMed2001

Increased XRALDH2 activity has a posteriorizing effect on the central nervous system of Xenopus embryos.

Retinoic acid (RA) metabolizing enzymes play important roles in RA signaling during vertebrate embryogenesis. We have previously reported on a RA degrading enzyme, XCYP26, which appears to be critical for the anteroposterior patterning of the central nervous system (EMBO J. 17 (1998) 7361). Here, we report on the sequence, expression and function of its counterpart, XRALDH2, a RA generating enzyme in Xenopus. During gastrulation and neurulation, XRALDH2 and XCYP26 show non-overlapping, complementary expression domains. Upon misexpression, XRALDH2 is found to reduce the forebrain territory and to posteriorize the molecular identity of midbrain and individual hindbrain rhombomeres in Xenopus embryos. Furthermore, ectopic XRALDH2, in combination with its substrate, all-trans-retinal (ATR), can mimic the RA phenotype to result in microcephalic embryos. Taken together, our data support the notion that XRALDH2 plays an important role in RA homeostasis by the creation of a critical RA concentration gradient along the anteroposterior axis of early embryos, which is essential for proper patterning of the central nervous system in Xenopus.

Aldehyde Dehydrogenase 1 Family↗

PubMed2001

Xpitx3: a member of the Rieg/Pitx gene family expressed during pituitary and lens formation in Xenopus laevis.

Xpitx3 is the Xenopus homologue of the mouse Pitx3 gene and belongs to the family of RIEG/PITX homeobox genes. Here, we report on the embryonic expression of Xpitx3. It is transcribed in the presumptive pituitary already at the open neural tube stage. During further development Xpitx3 is strongly transcribed in the pituitary Anlage, the lens placodes and head mesenchyme, respectively.

Amino Acid Sequence↗

PubMed2001

Cytosolic import factor- and Ran-independent nuclear transport of ribosomal protein L5.

Ribosomal protein L5 is a shuttling protein that, in Xenopus oocytes, is involved in the nucleocytoplasmic transport of 5S rRNA. As demonstrated earlier, L5 contains three independent nuclear import signals (NLSs), which function in oocytes as well as in somatic cells. Upon physical separation, these NLSs differ in respect to their capacity to bind to nuclear import factors in vitro and to mediate the nuclear import of a heterologous RNP in vivo. As reported in this communication, analysis of the in vitro nuclear import activity of these three NLSs reveals that they also differ in respect to their requirements for cytosolic import factors and Ran. Nuclear import mediated by the N-terminal and the central NLS depends on cytosolic import factor(s) and Ran, whereas import via the C-terminal NLS occurs independently from these factors. Thus, the presence of multiple NLSs in ribosomal protein L5 appears to allow for efficient nuclear transport via utilisation of multiple, mechanistically different import pathways.

Active Transport, Cell Nucleus↗

PubMed2000

Cloning and expression of xSix3, the Xenopus homologue of murine Six3.

The vertebrate Six family of transcription factor genes are homologues of the fruitfly gene sine oculis (so). Human, murine, avian and fish (medaka, zebrafish) homologues have recently been cloned. We report the cloning and developmental pattern of expression of xSix3, the Xenopus laevis homologue of Six3. In addition, we have compared all the known sequences of vertebrate Six3 genes. xSix3 is very homologous to Six3 in other vertebrates in terms of amino acid sequence. The reported developmental pattern of expression of Six3 in chick and mouse includes not only the developing eyes and the ventral diencephalic tissue between them, but also a large, sagittally-oriented telencephalic region. The distribution of xSix3, however, is virtually restricted to the eyes and ventral diencephalon, showing only a very small territory of expression in the telencephalon.

Amino Acid Sequence↗

PubMed2000

Xnkx-2.1: a homeobox gene expressed during early forebrain, lung and thyroid development in Xenopus laevis.

Nkx-2.1 is a member of the vertebrate Nkx family of homeobox genes; it was originally identified as a tissue-specific regulator of thyroglobulin and thyroperoxidase gene transcription. Here we report on the embryonic expression of Xnkx-2.1, which is expressed in the presumptive forebrain from early neurulation onwards. In tadpole stage embryos Xnkx-2.1 transcripts are primarily detected in ventral forebrain, lung buds and thyroid anlage. Therefore, Xnkx-2.1 may be part of the genetic network that controls the early development of these organs.

Amino Acid Sequence↗

PubMed2000

Expanded retina territory by midbrain transformation upon overexpression of Six6 (Optx2) in Xenopus embryos.

During vertebrate eye development, the expression of the homeobox gene Six6 is restricted to the neural retina and is initiated later than Rx and Pax6 in the presumptive retina field. We show here that overexpression of mouse Six6 in Xenopus embryos can induce transformation of competent tissue of the anterior neural plate into retinal tissue. In Six6 injected embryos, the molecular identity of the presumptive midbrain and rostral hindbrain regions was lost, as shown by the absence of XEn-2 and Xpax2 expression, being replaced by the ectopic expression of the retinal markers Xpax6 and Xrx. When allowed to grow further, Six6 injected embryos developed ectopic eye-like structures in the rostral brain and showed a transformation of the midbrain into retina. Similar results were obtained upon overexpression of Six3 or Xsix3, revealing a possible redundance of Six3 and Six6 activities. Taken together, results obtained suggest that during normal retina development, the relatively late expressed Six6 gene becomes part of a network of retinal homeobox genes that are linked together by positive feedback loops. Furthermore, our results demonstrate that the primitive neural ectoderm of the future midbrain and rostral hindbrain is competent to form retinal tissue.

Animals↗

PubMed2000

Structure and expression of Xenopus karyopherin-beta3: definition of a novel synexpression group related to ribosome biogenesis.

Karyopherin-beta3 is a nuclear transport receptor that appears to be involved in nuclear import of ribosomal proteins. Here we report on sequence and expression of karyopherin-beta3 in Xenopus. The differential distribution of karyopherin-beta3 mRNA during Xenopus embryogenesis is similar to that of several other protein import factors and of ribosomal proteins. These genes thus define a novel synexpression group in the context of ribosome biogenesis.

Amino Acid Sequence↗

PubMed2000

Planar signalling is not sufficient to generate a specific anterior/posterior neural pattern in pseudoexogastrula explants from Xenopus and Triturus.

Early observations on the morphology of total exogastrulae from urodeles (Axolotl) had provided evidence for essential vertical signalling mechanisms in the process of neural induction. Conversely, more recent studies with anurans (Xenopus laevis) making use of molecular markers for neural-specific gene expression appear to support the idea of planar signalling as providing sufficient information for neural differentiation along the anterior-posterior axis. In an attempt to resolve this apparent contradiction, we report on the comparative analysis of morphology and gene expression characteristics with explants prepared from both urodeles (Triturus alpestris) and anurans (Xenopus laevis). For this purpose, we have made use of a refined experimental protocol for the preparation of exogastrulae that is intended to combine the advantages of the Holtfreter type exogastrula and the Keller sandwich techniques, and which we refer to as pseudoexogastrula explants. Analysis of histology and expression of several neural and ectodermal marker genes in such explants suggests that neural differentiation is induced in both species, but only within the intermediate zone between ectoderm and endomesoderm. Therefore, experiments with Xenopus and Triturus explants described in this communication argue against planar signalling events as being sufficient to generate a specific anterior/posterior neural pattern.

Animals↗

PubMed1999

Vax1, a novel homeobox-containing gene, directs development of the basal forebrain and visual system.

The novel homeobox-containing gene Vax1, a member of the Emx/Not gene family, is specifically expressed in the developing basal forebrain and optic nerve. Here, we show that Vax1 is essential for normal development of these structures. Mice carrying a targeted mutation of Vax1 show dysgenesis of the optic nerve, coloboma, defects in the basal telencephalon, and lobar holoprosencephaly. With the help of molecular markers we determined that in the developing visual system, the absence of Vax1 results in a proximal expansion of the activity of Pax6 and Rx. This observation suggests that Vax1 may interfere negatively with the expression of Pax6 and Rx. In reciprocal gain-of-function experiments, injection of Xvax1 mRNA or Shh into Xenopus embryos primarily affects the brain at the level of the eye primordium. Consistent with the loss-of-function results, the injection of Xvax1 results in a down-regulation of Rx. Similarly, Shh injection expands the Vax1 and Pax2 territory at the expense of the Pax6 and Rx region. On the basis of these results, we propose a model for a molecular cascade involved in the establishment of structures of the visual system.

Abnormalities, Multiple↗

PubMed1999

Functional modules in ribosomal protein L5 for ribonucleoprotein complex formation and nucleocytoplasmic transport.

Ribosomal protein L5 forms a small, extraribosomal complex with 5 S ribosomal RNA, referred to as the 5 S ribonucleoprotein complex, which shuttles between nucleus and cytoplasm in Xenopus oocytes. Mapping elements in L5 that mediate nuclear protein import defines three separate such activities (L5-nuclear localization sequence (NLS)-1, -2, and -3), which are functional in both oocytes and somatic cells. RNA binding activity involves N-terminal as well as C-terminal elements of L5. In contrast to the full-length protein, none of the individual NLSs carrying L5 fragments are able to allow for the predominating accumulation in the nucleoli that is observed with the full-length protein. The separate L5-NLSs differ in respect to two activities. Firstly, only L5-NLS-1 and -3, not L5-NLS-2, are capable of promoting the nuclear transfer of a heterologous, covalently attached ribonucleoprotein complex. Secondly, only L5-NLS-1 is able to bind strongly to a variety of different import receptors; those that recognize L5-NLS-2 and -3 have yet to be identified.

Amino Acid Sequence↗

PubMed1999

Regulation of DNA binding activity and nuclear transport of B-Myb in Xenopus oocytes.

DNA binding activity and nuclear transport of B-Myb in Xenopus oocytes are negatively regulated. Two distinct sequence elements in the C-terminal portion of the protein are responsible for these different inhibitory activities. A C-terminal Xenopus B-Myb protein fragment inhibits the DNA binding activity of the N-terminal repeats in trans, indicating that intramolecular folding may result in masking of the DNA binding function. Xenopus B-Myb contains two separate nuclear localization signals (NLSs), which, in Xenopus oocytes, function only outside the context of the full-length protein. Fusion of an additional NLS to the full-length protein overcomes the inhibition of nuclear import, suggesting that masking of the NLS function rather than cytoplasmic anchoring is responsible for the negative regulation of Xenopus B-Myb nuclear transfer. During Xenopus embryogenesis, when inhibition of nuclear import is relieved, Xenopus B-myb is preferentially expressed in the developing nervous system and neural crest cells. Within the developing neural tube, Xenopus B-myb gene transcription occurs preferentially in proliferating, non-differentiated cells.

Amino Acid Sequence↗

PubMed1999

Characterization of the Ets-type protein ER81 in Xenopus embryos.

A function for FGF-type peptide growth factors has been implied for early mesodermal patterning events in Xenopus laevis. FGF signalling operates via the MAP kinase cascade that can directly activate the transcription of organizer-expressed genes, such as Xbra and Xegr-1. We have recently provided evidence for a critical role of Ets-type transcription factors in FGF mediated Xegr-1 transcription activation. Here, we report on the identification of the Xenopus Ets-type protein ER81 that is expressed in a pattern overlapping with the ones of Xegr-1 and Xbra during gastrulation. Microinjection in XER81 encoding mRNA into ventral blastomeres of Xenopus embryos results in the induction of ectopic, tail-like protrusions, whereas dorsal overexpression results in disturbed eye development. In the animal cap assay, ectopic expression of XER81 is found to interfere with activin mediated induction of Xegr-1 and gsc, but not with the Xbra response to activin.

Activins↗

PubMed1999

Xpitx-1: a homeobox gene expressed during pituitary and cement gland formation of Xenopus embryos.

Pitx-1 is a member of the family of bicoid-related vertebrate homeobox genes; it was originally identified as a tissue-specific transcriptional regulator of the proopiomelacortin gene. Here we report on the embryonic expression of Xpitx-1, which is expressed in the anterior neural ridge and in the cement gland Anlage during late gastrulation/early neurulation. In tadpole stage embryos Xpitx-1 transcripts are primarily detected in the cement gland, stomodeal-hypophyseal Anlage, oral epithelia and lens placode. Therefore, Xpitx-1 may be part of the genetic network that controls the early development of these structures.

Amino Acid Sequence↗

PubMed1999

Characterization of a subfamily of related winged helix genes, XFD-12/12'/12" (XFLIP), during Xenopus embryogenesis.

The fork head domain family of genes defines a growing group of proteins that serve important regulatory functions in pattern-forming events of both invertebrates and vertebrates. Here we add three closely related, novel members to this family in Xenopus laevis, termed XFD-12, XFD-12' and XFD-12". All three genes reveal indistinguishable expression patterns during Xenopus embryogenesis. During gastrulation, XFD-12 type transcripts are detected exclusively in the superficial layer of cells within the Spemann organizer territory. In the open neural plate, XFD-12 type expression defines a row of cells located along the dorsal midline and destined to become the floor plate of the neural tube. After closure of the neural tube, XFD-12 type encoding mRNAs are only detected in the tailtip and a small area located at the midbrain/hindbrain boundary. Within the Spemann organizer and in the floor plate area, expression of XFD-12 type genes is only partially overlapping with XFD-1 expression.

Amino Acid Sequence↗

PubMed1998

Regionalized metabolic activity establishes boundaries of retinoic acid signalling.

The competence of a cell to respond to the signalling molecule retinoic acid (RA) is thought to depend largely on its repertoire of cognate zinc finger nuclear receptors. XCYP26 is an RA hydroxylase that is expressed differentially during early Xenopus development. In Xenopus embryos, XCYP26 can rescue developmental defects induced by application of exogenous RA, suggesting that the enzymatic modifications introduced inhibit RA signalling activities in vivo. Alterations in the expression pattern of a number of different molecular markers for neural development induced upon ectopic expression of XCYP26 reflect a primary function of RA signalling in hindbrain development. Progressive inactivation of RA signalling results in a stepwise anteriorization of the molecular identity of individual rhombomeres. The expression pattern of XCYP26 during gastrulation appears to define areas within the prospective neural plate that develop in response to different concentrations of RA. Taken together, these observations appear to reflect an important regulatory function of XCYP26 for RA signalling; XCYP26-mediated modification of RA modulates its signalling activity and helps to establish boundaries of differentially responsive and non-responsive territories.

Amino Acid Sequence↗

PubMed1998

Structural determinants in 5S RNA and TFIIIA for 7S RNP formation.

C2H2-type zinc-finger modules define a unique structural motif, which is capable of forming specific complexes with both DNA and RNA. While the principles governing DNA binding have been defined in great detail, the mode of RNA recognition remains only poorly understood. In the absence of information from three-dimensional structural analysis of a zinc-finger/RNA complex, we have performed a number of biochemical studies to gain further insight into the molecular details of the interaction of 5S ribosomal RNA with the zinc-finger protein TFIIIA. Previous work had indicated that zinc finger 6 of TFIIIA contacts 5S RNA in close proximity or directly in the loop-A region (nucleotides 10-13). Permutation analysis of this sequence reveals that three of the four nucleotides are of vital importance for RNA recognition. Exchange of unusual and therefore characteristic aromatic residues in finger 6 against aliphatic or other aromatic amino acids reveals that the aromatic character of tryptophan 177 is essential for RNA recognition. Association with helix V in 5S RNA appears to involve specific contacts with the phosphate backbone, as evidenced by ethylation-interference assays. Introduction of multiple internal and 3'-terminal as well as 5'-terminal deletions accompanied by stabilizing sequence substitutions defines a minimal RNA fragment that is sufficient for TFIIIA binding. This RNA molecule includes a truncated/mutated helix I, helix II and helix V, as well as structurally intact loops A and E. Permutation analysis of the loop-E region emphasizes its importance for TFIIIA recognition.

Amino Acid Sequence↗

PubMed1998

The Spemann organizer-expressed zinc finger gene Xegr-1 responds to the MAP kinase/Ets-SRF signal transduction pathway.

The transcriptional activity of a set of genes, which are all expressed in overlapping spatial and temporal patterns within the Spemann organizer of Xenopus embryos, can be modulated by peptide growth factors. We identify Xegr-1, a zinc finger protein-encoding gene, as a novel member of this group of genes. The spatial expression characteristics of Xegr-1 during gastrulation are most similar to those of Xbra. Making use of animal cap explants, analysis of the regulatory events that govern induction of Xegr-1 gene activity reveals that, in sharp contrast to transcriptional regulation of Xbra, activation of Ets-serum response factor (SRF) transcription factor complexes is required and sufficient for Xegr-1 gene expression. This finding provides the first indication for Ets-SRF complexes bound to serum response elements to be activated during gastrulation. MAP kinase signalling cascades can induce and sustain expression of both Xegr-1 and Xbra. Ectopic Xbra can induce Xegr-1 transcription by an indirect mechanism that appears to operate via primary activation of fibroblast growth factor secretion. These findings define a cascade of events that links Xbra activity to the signal-regulated control of Xegr-1 transcription in the context of early mesoderm induction in Xenopus laevis.

Amino Acid Sequence↗