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T D Sargent

Publications and source records attributed to T D Sargent.

At least 37 records · Page 2Linked to original sources
PubMed1991

Developmental regulation of transcription factor AP-2 during Xenopus laevis embryogenesis.

We have isolated a cDNA clone encoding the Xenopus homologue of the transcription factor AP-2 (XAP-2). The predicted amino acid sequence derived from the Xenopus cDNA shows very strong conservation with the amino acid sequence of human AP-2, suggesting that this protein is evolutionarily conserved, at least among vertebrates. This is further substantiated by the demonstration that an in vitro translation product of XAP-2 cDNA bound specifically to an AP-2 binding site from the human MT-IIA gene. Northern blot analysis of Xenopus embryo RNA revealed the existence of three major XAP-2 mRNA species that were only detectable after the midblastula transition (when embryonic transcription is activated), with peak accumulation of the transcripts occurring during gastrulation. Therefore, in contrast to other Xenopus transcription factors, XAP-2 is not maternally derived but arises exclusively from zygotic transcription. Unlike the situation in cultured human teratocarcinoma (NT2) cells, retinoic acid treatment did not induce XAP-2 mRNA in Xenopus embryos, even though the treatment had a pronounced morphogenetic effect on the embryos. Our results suggest that XAP-2 may play a distinctive role during Xenopus embryogenesis.

Amino Acid Sequence↗
PubMed1991

Localized and inducible expression of Xenopus-posterior (Xpo), a novel gene active in early frog embryos, encoding a protein with a 'CCHC' finger domain.

Xenopus-posterior (Xpo) is a gene that is activated at or shortly after the midblastula transition (MBT). The RNA accumulates to a relatively low level, which remains constant until gastrulation, then rapidly and transiently increases in posterior ectoderm and mesoderm. A single copy of a putative finger motif, of the 'CCHC' type, is located near the carboxyl terminus. One or two copies of similar sequence motifs are found in the nucleocapsid protein of retroviruses where they are involved in protein-RNA interactions, and in cellular nucleic acid binding protein (CNBP), a protein that binds to the sterol regulatory element. Xpo expression is induced in ectodermal explants by treatment with basic fibroblast growth factor (bFGF) and with polypeptide growth factors found in medium conditioned by the Xenopus XTC cell line (XTC-CM). Taken together, these properties suggest a possible role for Xpo in the organization of the anteroposterior axis during development.

Animals↗
PubMed1991

Transcription factor AP-2 is tissue-specific in Xenopus and is closely related or identical to keratin transcription factor 1 (KTF-1).

This paper identifies a new, developmental role for transcription factor AP-2 in the activation of amphibian embryonic epidermal keratin gene expression. Keratin transcription factor KTF-1 is shown by several criteria to be identical or closely related to AP-2. KTF-1/AP-2 is shown to be tissue-specific from its first transcription in Xenopus embryos, and restricted to a small number of adult tissues, including skin. Epidermis-specific keratin transcription closely follows specification of the embryonic ectoderm in Xenopus, and is subject to regulation by growth factors and embryonic induction. We further show that in mouse basal keratinocytes, a KTF-1/AP-2-like factor is present and binds to a DNA sequence previously shown to be important in the regulation of the keratin K14 gene, which is actively expressed in these cells. Thus, the study of AP-2 and its role in the regulation of keratin gene transcription should enhance our understanding of both amphibian embryonic development and mammalian skin differentiation.

Animals↗
PubMed1990

Molecular approach to dorsoanterior development in Xenopus laevis.

RNA from ventralized Xenopus laevis embryos has been used to generate a subtracted library consisting of genes whose expression is dependent upon dorsoanterior development. Clones were selected on the basis of two criteria: (1) at least 10-fold reduction in ventralized versus control embryo RNA, and (2) absence of hybridization to RNA from adult muscle. One of these clones, designated UVS.2, was characterized further. UVS.2, was found by in situ hybridization to be expressed exclusively in the anterior neural fold of neurula stage embryos. By the tailbud stage, the UVS.2 protein and transcript were localized in specialized cephalic ectoderm, in a region probably corresponding to the hatching gland. UVS.2 expression could be induced in ectodermal explants by contact with dorsal mesodermal tissue. Furthermore, in lithium-treated embryos, which have exaggerated dorsoanterior structures, UVS.2 expression was increased four- to fivefold. A molecular marker such as UVS.2 will be useful in defining and studying the induction and organization of cephalic tissues.

Animals↗
PubMed1990

KTF-1, a transcriptional activator of Xenopus embryonic keratin expression.

Nuclear extracts from embryos of Xenopus laevis were shown to contain a protein activity, KTF-1, which binds in vitro to the promoter of the embryonic, epidermis-specific keratin gene, XK81A1. Mobility shift assays, methylation interference and footprinting analysis were used to show that the KTF-1 binding site contains an imperfect, palindromic sequence, ACCCTGAGGCT. This sequence occurs once in the XK81A1 promoter, 152-162 base pairs upstream of the transcription start site. A construct of the keratin gene in which this sequence was altered so that it no longer binds KTF-1 in vitro showed severely reduced transcription levels upon injection into Xenopus embryos, but retained epidermal specificity. Addition of KTF-1 binding sites also enhanced epidermal and non-epidermal activity of a heterologous promoter, Xenopus beta-globin, in embryos. These results suggest that KTF-1 is a general activator of embryonic keratin transcription, which acts in concert with other factors to produce high levels of epidermis-specific expression.

Animals↗
PubMed1989

Development of neural inducing capacity in dissociated Xenopus embryos.

During gastrulation in amphibians, dorsal ectoderm is converted by induction from an epidermal pathway to central nervous system (CNS). We show in this report that the ability of the embryo to manufacture this neural inducing signal is not dependent upon intercellular communication between early cleavage and early gastrula stages. This result is consistent with the interpretation that the cell lineage that induces CNS formation arises itself by a cell-autonomous mechanism, perhaps specified by materials inherited from the fertilized egg.

Actins↗
PubMed1989

Xenopus laevis serum albumin: sequence of the complementary deoxyribonucleic acids encoding the 68- and 74-kilodalton peptides and the regulation of albumin gene expression by thyroid hormone during development.

In adult Xenopus serum, albumin gene expression is regulated by estrogen through the selective destabilization of its mRNA during the vitellogenic response. The present study reports the cDNA sequence of both the 68K and 74K Xenopus albumin mRNAs, their derived amino acid sequence, and the regulation of albumin gene expression during embryogenesis. Albumin mRNA has a 39 nucleotide 5' untranslated region terminating in a consensus translation initiation site. The derived amino acid sequence yields a 24-amino acid hydrophobic leader sequence (terminating in Lys-Arg) that shares significant homology with the leader peptide of rat albumin. Overall there is 37% sequence identity between rat and frog albumin, with exact conservation of all but one Cys residue and the Pro residues responsible for the three domain structure of the mature protein. The 74K albumin (unlike the 68K albumin) is glycosylated; a point mutation converting Lys256 to Asn introduces an N-linked glycosylation site that is similar to one found in the sequence of mammalian alpha-fetoproteins. A larval albumin-like protein was not detectable by silver staining in serum of tadpoles before the beginning of metamorphosis at stage 48. Albumin mRNA is absent from early tadpoles (stages 22-47); however, it is rapidly induced at stage 48 as one of the earliest manifestations of metamorphosis. Exposure of embryos to 10(-8) M T3, which regulates amphibian metamorphosis, resulted in the premature induction of albumin mRNA, such that it is evident by stage 43.

Amino Acid Sequence↗
PubMed1989

Transcriptional regulation of a Xenopus embryonic epidermal keratin gene.

XK81A1 is a type I epidermal keratin gene expressed in early developmental stages of Xenopus (Jonas et al. 1985). Fusion of the keratin promoter (-5900 to +26) to a human beta globin gene led to fully epidermis-specific accumulation of human globin mRNA and protein when this DNA was injected into fertilized eggs. Further localization of regulatory sequences was accomplished by injecting marked, 5'-deleted keratin gene DNA into fertilized eggs and evaluating tissue specificity of expression. All 5' flanking DNA upstream from -487 could be removed without interfering with keratin gene expression or regulation. These results suggest that the primary mode of regulation of epidermis-specific keratin gene expression is at the level of transcription, and that sequence elements in the 5' flanking region of the keratin gene, between -487 and +26, are responsible for this regulation.

Animals↗
PubMed1988

Xenopus laevis in developmental and molecular biology.

Xenopus laevis is a prime system for the study of embryogenesis in vertebrates. Both prelocalized information in the egg and inductive interactions between cells contribute to the ordered increase in complexity during development. Embryonic induction, discovered in amphibians, is being studied intensely in Xenopus; recent work suggests a role for growth factors in this process. Contributions of the Xenopus system to the analysis of ribosomal and 5S RNA genes, and the diverse and highly productive applications of the oocyte injection technology, are also summarized.

Animals↗
PubMed1988

Accumulation and decay of DG42 gene products follow a gradient pattern during Xenopus embryogenesis.

The DG42 gene is expressed during a short window during embryogenesis of Xenopus laevis. The mRNA for this gene can be first detected just after midblastula, peaks at late gastrula, and decays by the end of neurulation. The sequence of the DG42 cDNA and genomic DNA predicts a 70,000-Da protein that is not related to any other known protein. Antibodies prepared against portions of the DG42 open reading frame that had been expressed in bacteria detected a 70,000-Da protein in the embryo with a temporal course of appearance and decay that follows that of the RNA by several hours. Localization of the mRNA in dissected embryos and immunohistochemical detection of the protein showed that DG42 expression moves as a wave or gradient through the embryo. The RNA is first detected in the animal region of the blastula, and by early gastrula is found everywhere except in the outer layer of the dorsal blastopore lip. By midgastrula DG42 protein is present in the inner ectodermal layer and the endoderm; it disappears from dorsal ectoderm as the neural plate is induced and later decays in a dorsoventral direction. The last remnants of DG42 protein are seen in ventral regions of the gut at the tailbud stage.

Animals↗
PubMed1988

Gene expression in amphibian embryogenesis.

The study of molecular events during the embryogenesis of Xenopus laevis has advanced as a result of the availability of molecular markers, i.e., nucleic acid and antibody probes for genes that are expressed in a temporally and spatially regulated fashion during development. In this article we summarize results on the localized expression of keratin genes and on the reconstruction of regulated transcription of the gastrula/neurula-specific DG42 gene. Furthermore, we discuss experiments that investigate molecular events during mesoderm induction and provide information on the nature of the inducing principle.

Animals↗
PubMed1988

Xenopus endo B is a keratin preferentially expressed in the embryonic notochord.

Screening of a cDNA library from neurula stage Xenopus laevis for notochord-specific sequences led to the isolation of a cDNA clone, XK endo B (Xenopus keratin endo B), which encodes a nonepidermal type I keratin. In situ and Northern blot hybridizations indicate that expression of XK endo B RNA is concentrated in the notochord, whereas expression in the endoderm is 5-10 times lower. XK endo B mRNA is present in the oocyte and increases from late gastrula. Accumulation peaks by late neurula and is greatly reduced by the tadpole stage; in the adult, a low level of XK endo B RNA is present in the liver. XK endo B shows sequence homology to mouse endo B; genomic Southern blots show that XK endo B is the most similar sequence to mouse endo B in the Xenopus genome, and vice versa, indicating that XK endo B and mouse endo B are homologs. The use of endo B as a marker and the germ layer derivation of the notochord are discussed in light of these results.

Amino Acid Sequence↗
PubMed1987

Cell-type-specific expression of epidermal cytokeratin genes during gastrulation of Xenopus laevis.

Analysis of the spatial pattern of expression of embryo-specific epidermal cytokeratin genes in Xenopus laevis shows earliest activity in the animal pole cells of stage-9 blastulae. These genes are transcribed predominantly in the epithelial or outer ectoderm, to a lesser extent in the sensorial or inner ectoderm, and at low levels if at all in other regions of the embryo. In the early gastrula the entire ectoderm, including preneural and preepidermal regions, expresses cytokeratin mRNAs; accumulation of these mRNAs in preneural cells is terminated after contact is made with involuting chordamesoderm. On the basis of this and earlier work (Sargent et al. 1986) we suggest that the pattern of expression of cytokeratin genes in frog embryogenesis is based on prelocalized components modulated by the inductive influence of involuting chordamesoderm. The cytokeratin proteins are deposited in the form of filamentous networks in both layers of the epidermis. In the epithelial layer, a much denser mesh of filaments is facing the outside of the embryo. This polarity is established at the onset of the polymerization of these filaments. Thus, intraembryonic and intracellular localization of keratin gene expression and protein deposition is established at the onset of activation of these genes.

Animals↗
PubMed1986

Cell interactions and the control of gene activity during early development of Xenopus laevis.

During embryonic development, regulation of the zygotic genome may be mediated by inductive interactions and by cell-autonomous inheritance of informational material from the egg; we have studied certain aspects of such regulatory events in Xenopus laevis. Embryos cultured in Ca2+/Mg2+-free medium can be dissociated and dispersed, eliminating cell-cell contact and thus precluding inductive interactions. Such manipulations revealed that activation of the muscle-specific alpha-actin genes is absolutely dependent upon cell contacts. Conversely, the endoderm-specific DG42 gene and the ectoderm-specific DG81 cytokeratin gene are activated in embryo cells dispersed throughout blastula stages and therefore appear to be controlled by inherited factors. Nevertheless, abnormal cell arrangements may prevent expression of the cytokeratin gene, suggesting that animal pole cells can be diverted from their normal ectodermal fate by inductive contact with vegetally derived cells. The interactions required for alpha-actin induction and inhibition of cytokeratin expression are independent of strong adhesion between embryonic cells mediated by high concentrations of divalent cations.

Actins↗