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J R Tata

Publications and source records attributed to J R Tata.

At least 55 records · Page 3Linked to original sources
PubMed1991

Developmental and regional expression of thyroid hormone receptor genes during Xenopus metamorphosis.

A characteristic feature of the obligatory control of amphibian metamorphosis by thyroid hormones is the early acquisition of response of tadpole tissues to these hormones well before the latter are secreted, with 'exponentially' increasing hormonal sensitivity upon the onset of metamorphosis. We have therefore analyzed the expression of the two thyroid hormone receptor genes (TR alpha and beta) before, during and after metamorphosis in Xenopus tadpoles and froglets. Using non-cross-hybridizing cRNA probes for 5' and 3' sequences of Xenopus TR alpha and beta transcripts for RNAase protection assays, the two mRNAs can be detected in tadpoles as early as stage 39. Their concentration increases abruptly at stage 44 and continues to increase differentially at the onset of metamorphosis (stage 55) and through metamorphic climax at stages 58-62, after which they decline upon completion of metamorphosis at stage 66. Quantitative densitometric scanning of autoradiograms showed that, although the concentration of TR beta transcripts is about 1/30th of that of TR alpha mRNA at stages 44-48, depending on the region, it accumulates 3-10 times more rapidly than does the alpha isoform during further development. A substantial proportion of the increase in TR beta mRNA is localized to the head region of tadpoles. Using the hormone-binding domain (HBD) and 3' end of Xenopus TR alpha cRNA as probe for in situ hybridization, the highest concentration of TR transcripts in stage 44 tadpoles is seen in the brain and spinal cord. High concentrations of mRNA are also present in the intestinal epithelium and tail tip, tissues programmed for regression. At later stages (55 onwards), strong hybridization signals are also exhibited by hindlimb buds. This pattern persists through metamorphic climax, after which TR mRNAs decline in all tissues to low levels in froglets at stage 66. In developing froglets, TR transcripts were detected in large amounts in the cytoplasm of stage 1 and 2 oocytes but the rate of their accumulation did not increase with further oocyte growth. This observation raises the possibility that the response to thyroid hormones at early stages of tadpoles (42-44) may be due to TR synthesized on maternally derived mRNA. Exposure of tadpoles at premetamorphic stages (48-52) to exogenous thyroid hormone (T3) substantially enhanced the accumulation of TR mRNA, especially that of TR beta message, which could explain the accelerated increase in sensitivity of tadpoles to thyroid hormones at the onset of natural metamorphosis.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1990

Autoinduction of estrogen receptor is associated with FOSP-1 mRNA induction by estrogen in primary cultures of Xenopus oviduct cells.

The number of nuclear and cytosolic estrogen receptors (ER) per cell and the steady-state levels of the mRNA encoding a tissue-specific, estrogen-inducible protein (FOSP-1) were measured as a function of time following the addition of estradiol-17 beta (E2) to primary cultures of Xenopus oviduct cells. After a lag period of about 12 h, 10(-9) to 10(-7) M E2 caused a 10 to 15-fold increase in FOSP-1 mRNA by 60 h, whereas it was only 2-fold with 10(-7) M progesterone. Under the same conditions, E2 doubled its own total receptor content within the first 12 h, reaching a 4-fold increase in nuclear ER by 48 h. Cycloheximide treatment in the presence of 10(-7) M estradiol reduced the functional ER content by 75.90%. Treatment with the anti-estrogen ICI 164,384 of oviduct cells in which FOSP-1 mRNA was pre-induced to high levels with the hormones caused a drastic reduction in nuclear ER and a total loss of FOSP-1 mRNA in 72 h. The close correlation between the kinetics of autoinduction of ER and the induction of FOSP-1 mRNA, as was shown earlier for vitellogenin mRNA in hepatocytes (Perlman et al. (1984) Mol. Cell. Endocrinol. 38, 151-161), strongly suggests that Xenopus egg protein gene activation by estrogen requires the up-regulation of its own receptor by the hormone.

Animals↗
PubMed1990

Accumulation of proto-oncogene c-erb-A related transcripts during Xenopus development: association with early acquisition of response to thyroid hormone and estrogen.

The expression of genes encoding receptors for estrogen and thyroid hormones, as well as total c-erb-A related transcripts was determined in unfertilized eggs, all stages of embryonic and larval development and in adult tissues of Xenopus, by quantitative Northern and slot-blot hybridization. DNA and antisense RNA probes complementary to Xenopus c-myc, cytoskeletal actin and albumin mRNAs served as controls or developmental markers. Hybridization to full-length chicken c-erb-A cDNA at moderate stringency revealed a complex biphasic ontogenic pattern for several c-erb-A related mRNAs in all tissues and at all developmental stages, an increase of 4-fold in the accumulation of these transcripts occurring before metamorphosis (stages 30 to 40-42) and followed by a gradual build-up after mid-metamorphosis (stage 56). Using full-length or ligand-binding domain fragments of thyroid hormone (TR) and estrogen (ER) cDNAs under stringent hybridization conditions, transcripts of TR and ER were detected from stages 44 and 54 onwards, respectively. The alpha and beta forms of TR mRNAs exhibited different patterns of accumulation during development, the former transcript being present in substantially higher amounts at all developmental stages. The distinct patterns of accumulation of TR and ER mRNAs could be correlated with the differential pattern of early developmental acquisition of sensitivity of Xenopus larval tissues to thyroid hormone and estrogen.

Animals↗
PubMed1990

Rosalind Pitt-Rivers and the discovery of T3.

The discovery of T3 as a major thyroid hormone brought international fame to Rosalind Pitt-Rivers in the 1950s. Here we reflect on the life and work of this distinguished biochemist who died in January of this year.

Endocrinology↗
PubMed1988

Selective degradation of mRNA: the role of short-lived proteins in differential destabilization of insulin-induced creatine phosphokinase and myosin heavy chain mRNAs during rat skeletal muscle L6 cell differentiation.

This investigation concerns the combined effects of removal and readdition of insulin and inhibition of protein and RNA synthesis on the stability of insulin-induced mRNAs during and after differentiation of rat L6A1 myoblast cells in culture. Addition of insulin accompanying the withdrawal of the mitogenic stimulus of serum to myoblasts caused an 80-fold increase in creatine phosphokinase (CK) activity which was largely accounted for by a similar increase in the amount of CK mRNA. The latter was co-ordinately induced with myosin heavy chain (MHC) mRNA but not malic enzyme (ME) mRNA. Measurements of steady-state levels of mRNA showed that removal of insulin caused CK mRNA, but not MHC mRNA, to be rapidly degraded, the effect being reversed upon readdition of the hormone. Direct measurement of 3H-labeled CK, MHC and beta-actin mRNAs confirmed the selective stabilization and destabilization of CK mRNA by the hormone. Conditions were established for a time-window during which cycloheximide (Cx) produced a virtually total arrest of protein synthesis in myotubes that was reversible upon removal of the inhibitor. Under these conditions, Cx selectively prevented the degradation of CK mRNA in a reversible manner. Actinomycin D (Act D) also arrested the loss of this mRNA. Under the same conditions of mRNA stabilization during de-induction, a superinduction of CK mRNA, but not MHC mRNA, was observed if the two inhibitors were added during induction in the continuous presence of insulin. We conclude that a short-lived protein(s), encoded by a short-lived mRNA(s), selectively regulates the stability of reversibly inducible mRNA.

Animals↗
PubMed1988

FOSP-1 (frog oviduct-specific protein-1) gene: cloning of cDNA and induction by estrogen in primary cultures of Xenopus oviduct cells.

We describe the isolation of a cloned cDNA from a cDNA library of oviduct of estrogen-treated adult Xenopus. Although the protein encoded by this cDNA is not known, it is designated as FOSP-1 (frog oviduct-specific protein-1). A partial restriction map of FOSP-1 cDNA, which is 1.5 kb in size, is presented. Northern hybridization analysis showed that FOSP-1 cDNA codes for a single species of mRNA of 2.6 kb which is exclusively expressed in Xenopus oviduct. Southern blot analysis showed that the gene was present in only one or two copies. Sequencing of partial FOSP-1 cDNA did not reveal homology with any protein in the sequence data bank. Measurement of steady-state levels of FOSP-1 mRNA in primary cultures of Xenopus oviduct cells by a technique of quantitative slot-blot analysis showed that both 17 beta and 17 alpha stereoisomers of estradiol caused a rapid 5-fold enhancement of accumulation of the mRNA with maximum values obtained at 5 X 10(-8) M estrogen. Progesterone caused only a small increase in FOSP-1 mRNA concentration. This hormone-specific induction of mRNA makes FOSP-1 a valuable candidate for exploring tissue specificity of regulation by estrogen of gene expression.

Amino Acid Sequence↗
PubMed1987

Establishment and characterization of Xenopus oviduct cells in primary culture.

Based on a previously established procedure for Xenopus hepatocytes, we describe tubular oviduct cells in primary culture which continue to secrete substantial quantities of egg jelly for several days, as can be visualized microscopically. Freshly isolated cells exhibited a culture shock response [A. P. Wolffe, J. F. Glover, and J. R. Tata (1984) Exp. Cell Res. 154, 581], from which they recovered by the third day in culture. This recovery was characterized by (a) the diminished synthesis of heat shock proteins hsp 70 and hsp 85, (b) the cessation of the drop in number of estrogen receptor, and (c) the enhanced rate of synthesis of cellular and secreted proteins. The oviduct estrogen receptor had the same characteristics as those in other estrogen target tissues and was present in the same amount as in adult female Xenopus hepatocytes [A. J. Perlman, A. P. Wolffe, J. Champion, and J. R. Tata (1984) Mol. Cell. Endocrinol. 38, 51]. Unlike the latter in primary culture [M. P. R. Tenniswood, P. F. Searle, A. P. Wolffe, and J. R. Tata (1983) Mol. Cell. Endocrinol. 30, 329], oviduct cell cultures did not actively metabolize either estradiol or progesterone (t1/2 approximately equal to 55 and 7 h, respectively). The successful establishment and characterization of primary cultures of both liver and oviduct cells now fulfil the conditions required for investigating the basis for tissue specificity of regulation by estrogen of Xenopus egg protein gene expression in primary cell culture.

Animals↗
PubMed1987

Hormonal regulation of RNA synthesis and specific gene expression in Xenopus oviduct cells in primary culture.

The studies described in this report were carried out as a first step towards the elucidation of mechanisms underlying the tissue specificity of regulation of gene expression by estrogen. Using a procedure established earlier in our laboratory for primary culture of Xenopus hepatocytes, we have characterized how estradiol-17 beta and progesterone affect the rate of synthesis of total RNA and that of accumulation of two oviduct-specific mRNAs in Xenopus oviduct cells in primary culture. In cells that had recovered from 'culture shock' 3 days after they were plated out, both hormones had only a slight or no effect on the overall rate of labelling of newly synthesized RNA over 24 h. Cloned cDNA probes for two mRNAs, termed 7F and 6G and specifying as yet unknown proteins expressed in the oviduct and not in the liver, were used to quantify the two mRNAs. The levels of both mRNAs declined for the first 2 days in culture after which they were stabilized. When added to the oviduct cell cultures 3 days after they were plated out, estradiol increased the steady-state concentration (relative to total RNA) of 7F and 6G mRNAs by 3- to 7-fold after 60-80 h, but with different time-course and dose-response kinetics for the two messages. The antiestrogen tamoxifen also exerted different degrees of antagonist effect on the estrogen-induced accumulation of 7F and 6G mRNAs. Although the protein products of these two oviduct-specific mRNAs have not yet been characterized, these studies set the stage for comparing the regulation by estrogen of their genes with that of vitellogenin genes in primary cultures of Xenopus oviduct cells and hepatocytes.

Animals↗
PubMed1985

Specific switching on of silent egg protein genes in vitro by an S-100 fraction in isolated nuclei from male Xenopus.

A soluble post-nuclear extract (S-100) specifically switches on the permanently silent vitellogenin genes in male Xenopus liver nuclei in vitro. Using cloned cDNAs to albumin, globin and vitellogenin mRNAs as hybridization probes, and heparin as an inhibitor of initiation of transcription, run-on assays with nuclei faithfully reproduced the transcriptional status of these genes in vivo. In the absence of heparin, a liver S-100 extract from female or male Xenopus treated with estradiol, but not from hormonally untreated males, induced the de novo transcription of vitellogenin genes in nuclei from hormonally naive male hepatocytes. The activation of the dormant genes was dependent on pre-incubation of nuclei with the competent extract for a minimum of 45 min at 24 degrees C before transcription was initiated. The same extract did not alter significantly the transcription of albumin and globin genes. The S-100 fraction from hormonally stimulated male hepatocytes exhibited a high degree of tissue specificity; it did not activate the transcription of vitellogenin genes in Xenopus erythrocyte and oviduct nuclei, and only slightly in nuclei from Xenopus XTC-2 cells, while a soluble extract from oviduct only slightly induced their transcription in male hepatocyte nuclei. This is the first demonstration of a hormone- and tissue-specific de novo activation of dormant genes by a soluble cell extract in isolated nuclei and offers a new approach to analyzing steroid hormone action and tissue-specific transcription factors in the regulation of gene expression.

Animals↗
PubMed1985

Galactoside-binding serum lectin of Xenopus laevis. Estrogen-dependent hepatocyte synthesis and relationship to oocyte lectin.

Xenopus laevis serum contains a lectin which binds alpha- and beta-galactosides. It was purified to homogeneity by affinity chromatography and consists of a single subunit with Mr approximately 69,000, associated in a multimer. The lectin is synthesized and secreted by hepatic parenchymal cells, and its synthesis is increased about 2-fold by estrogen treatment, both in vivo and in primary cell cultures. The serum lectin has the same carbohydrate binding properties as an oocyte lectin from X. laevis described previously, is immunologically cross-reactive, and shows similarities in its peptide map. However, marked differences in amino acid composition preclude the possibility that the serum lectin is a precursor of the oocyte lectin.

Amino Acids↗
PubMed1985

Deinduction of transcription of Xenopus 74-kDa albumin genes and destabilization of mRNA by estrogen in vivo and in hepatocyte cultures.

The goal of this study is to explain the molecular basis of the marked deinduction of Xenopus albumin synthesis and secretion accompanying the activation of vitellogenin genes by estrogen. We have characterized by restriction analysis, DNA sequencing and hybrid-selected translation of mRNA, a cloned cDNA specifying the two 74-kDa albumins which constitute the predominant circulating form of albumin in Xenopus laevis. Using this recombinant DNA plasmid as a hybridization probe, we have determined the steady-state levels of albumin mRNA, the rate of transcription of the two 74-kDa albumin genes and the stability of the mRNA in male and female Xenopus hepatocytes in vivo and in primary cell cultures following estrogen treatment. In both whole liver and cultured hepatocytes estradiol caused a rapid drop in the steady-state levels of 74-kDa albumin mRNAs, which was reversed spontaneously in the continued presence of the hormone. The concentration of albumin mRNA was substantially higher in male than in female hepatocytes, the hormonal effect being more marked in male than in female hepatocytes. The decrease in steady-state levels of mRNA was anticipated in male hepatocytes by a 70% inhibition of rate of transcription of albumin genes within 2 h of exposure to estradiol, as measured by run-off transcription in liver nuclei isolated from animals treated in vivo or by determining the absolute transcription rate in cell cultures. In the latter the diminished transcription rate returned to normal within 12 h in the continued presence of the hormone. Estradiol caused a threefold destabilization of albumin mRNA in both male and female hepatocyte cultures to t 1/2 = 3 h and 2 h respectively. The combined effects on rate of or transcription and mRNA stability largely explain the changes in the steady-state levels of mRNA caused by hormone administration. Comparison of the kinetics of transcription rates of vitellogenin and albumin genes in vivo and in vitro reveals a striking reciprocity in the selective activation of the inducible genes and deinduction of the constitutively expressed genes at the early stages of response of Xenopus hepatocytes to estrogen.

Albumins↗
PubMed1985

Inhibition by estradiol of binding and mitogenic effect of epidermal growth factor in primary cultures of Xenopus hepatocytes.

We demonstrate here the presence of two classes of EGF (epidermal growth factor) binding sites in primary cultures of male Xenopus liver parenchymal cells. One of these corresponds to the high-affinity receptor described in other tissues and species, and which exhibits the property of autophosphorylation. The number of EGF receptors decreased sharply in freshly prepared cultures but recovered to maximum levels within 24 h thereafter. Addition of EGF and insulin to the hepatocyte cultures enhanced the rate of DNA synthesis as measured by the incorporation of [3H]thymidine. Estrogen abolished this increase, reducing the incorporation to that seen with hydroxyurea. At the same time, the addition of estradiol reduced the number or activity of EGF receptors in a dose-dependent manner. The latter paralleled the activation of transcription of vitellogenin genes in Xenopus hepatocytes so that a high rate of DNA synthesis is unnecessary for or incompatible with the activation of the steroid hormone-induced vitellogenin genes.

Animals↗
PubMed1985

Vitellogenin genes and their products in closely and distantly related species of Xenopus.

Plasma vitellogenins from two closely related species of Xenopus, X. laevis and X. borealis, and a more ancient species, X. tropicalis, exhibited the same size on gel electrophoresis and were immunologically related. Partial peptide maps of 125I-labelled plasma vitellogenins, however, revealed marked differences in th structure and organisation of vitellogenin in the three Xenopus species. Northern blot hybridisation of liver RNA from oestrogen-treated males and females, probed with cloned vitellogenin cDNA, revealed the presence of mRNA of the same size in the three species of Xenopus, which was absent in untreated male liver. Cell-free translation of total liver RNA showed the presence of functional mRNA coding for vitellogenin subunit of the same size (Mr congruent to 210,000). Restriction endonuclease digestion patterns of genomic DNA from the three Xenopus species, using cloned X. laevis vitellogenin cDNA as the hybridisation probe, revealed significant differences in the organisation of these genes, which occur at a higher multiplicity in X. laevis and X. borealis than in X. tropicalis. Thus, despite a high degree of conservation of size, overall sequence and immunological identity of vitellogenin genes and their products in the three species of Xenopus, there is a substantial structural rearrangement during evolution of Xenopus within this multigene family.

Animals↗