Biomedical subjects

K Umesono

Publications and source records attributed to K Umesono.

At least 55 records · Page 3Linked to original sources

PubMed1993

Functional inhibition of retinoic acid response by dominant negative retinoic acid receptor mutants.

The diverse effects of retinoids on the development, growth, and homeostasis of vertebrate organisms are mediated in part by three distinct isoforms of retinoic acid receptors (RARs). These proteins, which are structurally and functionally closely related to thyroid hormone receptors and the oncogene product v-ErbA, regulate patterns of gene expression in target tissues. One approach to study the distinct effects of retinoic acid in cells is to subvert this activity of endogenous receptors by expression of dominant negative receptor derivatives. We demonstrate here that RAR alpha, RAR beta, and RAR gamma can be converted into potent negative transcriptional regulators that block wild-type RAR function. Furthermore, these mutant RARs, but not the wild-type receptors, actively repress the basal transcription level of target promoters. When expressed in transgenic mice, the most potent of these inhibitory receptor mutants is apparently able to disturb developmental processes by inducing a cleft palate in transgenic offspring.

Amino Acid Sequence↗

PubMed1993

Retinoid receptors.

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Amino Acid Sequence↗

PubMed1993

Determinants for selective RAR and TR recognition of direct repeat HREs.

Recently, we have shown that receptors for vitamin D3 (VDR), thyroid hormone (TR), and retinoic acid (RAR) activate preferentially through direct repeats (DRs) spaced by 3, 4, and 5 nucleotides, respectively. In addition, the RAR can activate weakly through DRs spaced by 2 nucleotides. A common feature of RAR, TR, and VDR is their ability to heterodimerize with the retinoid X receptor (RXR) through their ligand-binding domains (LBDs) to form high-affinity DNA-binding complexes that are specific for appropriately spaced repeats. In this paper we demonstrate that selective binding of RAR-RXR and TR-RXR heterodimers to their cognate DRs is a consequence of a novel cooperative dimer interaction within the DNA-binding domains (DBDs). Accordingly, a region in the first zinc finger of the TR and RAR DBDs interacts with the second zinc finger in the RXR DBD to promote selective DNA-binding to DRs spaced by 4 and 5 nucleotides, respectively. The resulting polarity established by this interaction places RXR in the 5' position of the direct repeats. These data provide a mechanism for selective receptor recognition of a restricted set of target sequences in DR DNA and explains the structural basis for physiological specificity.

Amino Acid Sequence↗

PubMed1992

Convergence of 9-cis retinoic acid and peroxisome proliferator signalling pathways through heterodimer formation of their receptors.

Peroxisomes are cytoplasmic organelles which are important in mammals in modulation of lipid homeostasis, including the metabolism of long-chain fatty acids and conversion of cholesterol to bile salts (reviewed in refs 1 and 2). Amphipathic carboxylates such as clofibric acid have been used in man as hypolipidaemic agents and in rodents they stimulate the proliferation of peroxisomes. These agents, termed peroxisome proliferators, and all-trans retinoic acid activate genes involved in peroxisomal-mediated beta-oxidation of fatty acids. Here we show that the receptor activated by peroxisome proliferators and the retinoid X receptor-alpha (ref. 6) form a heterodimer that activates acyl-CoA oxidase gene expression in response to either clofibric acid or the retinoid X receptor-alpha ligand, 9-cis retinoic acid, an all-trans retinoic acid metabolite; simultaneous exposure to both activators results in a synergistic induction of gene expression. These data demonstrate the coupling of the peroxisome proliferator and retinoid signalling pathways and provide evidence for a physiological role for 9-cis retinoic acid in modulating lipid metabolism.

Acyl-CoA Oxidase↗

PubMed1992

Retinoic acid receptors initiate induction of the cytomegalovirus enhancer in embryonal cells.

Reactivation of latent virus is believed to result from a signal transduction event that induces immediate-early (IE) gene transcription. Evidence is presented that the major IE promoter (MIEP) of human cytomegalovirus (hCMV) is activated by physiological levels of retinoic acid (RA) in human embryonal carcinoma cells. Mutagenesis experiments localized in the MIEP enhancer, a retinoic acid-responsive element composed of a direct repeat separated by five nucleotides. Protein-DNA binding experiments revealed that this element functions as a specific target site for the direct interaction of nuclear receptor proteins for RA. These findings implicate the biologically active derivative of vitamin A (RA) as a potential modulator of hCMV pathogenesis in infants and immunocompromised adults.

Animals↗

PubMed1992

Retinoid X receptor-COUP-TF interactions modulate retinoic acid signaling.

We have recently described the properties of direct repeats (DRs) of the half-site AGGTCA as hormone response elements (HREs). According to our results, spacing the half sites by 3, 4, or 5 nucleotides determines specificity of response for vitamin D3, thyroid hormone, and retinoic acid receptors, respectively. This so-called 3-4-5 rule led to the prediction that remaining spacing options of 0, 1, and 2 might serve as targets for other nuclear receptors. A concurrent prediction is that receptors recognizing common sites might display more complex or combinatorial interactions. In exploring these predictions, we discovered that both the retinoid X receptor (RXR) and COUP-TF bind preferentially to a DR-1 motif. In vivo, RXR and COUP-TF display antagonistic action such that RXR-mediated activation is fully repressed by COUP-TF. In vitro studies reveal that COUP-TF and RXR form heterodimers on DR-1. Thus, these results support a general proposal in which the half-site spacing preferences may be used as a means to decipher potentially complex and interactive regulatory circuits.

Base Sequence↗

PubMed1992

Retinoid X receptor interacts with nuclear receptors in retinoic acid, thyroid hormone and vitamin D3 signalling.

Cellular responsiveness to retinoic acid and its metabolites is conferred through two structurally and pharmacologically distinct families of receptors: the retinoic acid receptors (RAR) and the retinoid X receptors (RXR). Here we report that the transcriptional activity of RAR and RXR can be reciprocally modulated by direct interactions between the two proteins. RAR and RXR have a high degree of cooperativity in binding to target DNA, consistent with previous reports indicating that the binding of either RAR or RXR to their cognate response elements is enhanced by factors present in nuclear extracts. RXR also interacts directly with and enhances the binding of nuclear receptors conferring responsiveness to vitamin D3 and thyroid hormone T3; the DNA-binding activities of these receptors are also stimulated by the presence of nuclear extracts. Together these data indicate that RXR has a central role in multiple hormonal signalling pathways.

Animals↗

PubMed1992

Transcriptional regulation of the c-jun gene by retinoic acid and E1A during differentiation of F9 cells.

Differentiation of mouse F9 embryonal carcinoma (EC) cells can be induced by exposure to retinoic acid (RA) or by expression of adenovirus E1A. The transcription of the c-jun gene is stimulated by either RA or E1A. We report here that both RA and E1A strongly induce the expression of chloramphenicol acetyltransferase (CAT) from c-jun promoter/CAT reporter construct (c-jun/CAT), which is stably integrated into F9 cells, in a manner that is independent of both copy number and integration locus. The induction of c-jun/CAT expression is observed in undifferentiated F9 cells, but not in differentiated F9 cells, adenovirus-infected F9 cells or HeLa cells. Deletion analysis of the promoter region of the c-jun gene indicates that the sequence elements required for the RA- and E1A-mediated induction are identical and they have been defined as a region of 145 bp between -190 and -46 of the 5' flanking region of c-jun. This RA and E1A response element (RERE) contains five variants of the motif CGCGGTGACGNT. The upstream two motifs are adjacent and extend in opposite directions, creating an imperfect palindrome. The downstream four motifs are located at 35 or 36 bp intervals in the same orientation. Substitution and insertion analysis indicates that these motifs and their regular intervals are important for the activity of the RERE.

Adenoviruses, Human↗

PubMed1992

Modulation of glucocorticoid receptor function by protein kinase A.

Protein kinase A (PKA) has been shown to modulate the pattern of gene expression via transcription factors such as cAMP response element binding protein. However, in F9 embryonal carcinoma cells which lack endogenous functional cAMP response element binding protein, we have found that PKA is still able to control gene transcription through the glucocorticoid receptor (GR) by up-regulating its hormone-dependent trans-activation. Dose-response analysis indicates that PKA does not alter the ligand binding affinity of GR. PKA seems to act through the DNA binding domain of GR, since GR mutants which lack either the amino-terminal or the ligand binding domain are still able to be up-regulated by PKA. In support of this proposal, we demonstrate that PKA can enhance the DNA binding activity of GR. Our results suggest a novel mechanism by which PKA modulates the steroid sensitivity of a target cell by enhancing the DNA binding activity of GR for its cognate hormone response elements.

Animals↗

PubMed1991

Chromosomal translocation t(15;17) in human acute promyelocytic leukemia fuses RAR alpha with a novel putative transcription factor, PML.

A unique mRNA produced in leukemic cells from a t(15;17) acute promyelocytic leukemia (APL) patient encodes a fusion protein between the retinoic acid receptor alpha (RAR alpha) and a myeloid gene product called PML. PML contains a cysteine-rich region present in a new family of apparent DNA-binding proteins that includes a regulator of the interleukin-2 receptor gene (Rpt-1) and the recombination-activating gene product (RAG-1). Accordingly, PML may represent a novel transcription factor or recombinase. The aberrant PML-RAR fusion product, while typically retinoic acid responsive, displays both cell type- and promoter-specific differences from the wild-type RAR alpha. Because patients with APL can be induced into remission with high dose RA therapy, we propose that the nonliganded PML-RAR protein is a new class of dominant negative oncogene product. Treatment with RA would not only relieve this inhibition, but the activated PML-RAR protein may actually promote myelocyte differentiation.

Amino Acid Sequence↗

PubMed1991

A direct repeat in the cellular retinol-binding protein type II gene confers differential regulation by RXR and RAR.

The vitamin A derivative retinoic acid exerts its effects on transcription through two distinct classes of nuclear receptors, the retinoic acid receptor (RAR) and the retinoid X receptor (RXR). We provide evidence that expression of the gene for cellular retinol-binding protein type II (CRBPII), a key protein in the intestinal absorption of vitamin A, is dramatically up-regulated by retinoic acid in the presence of RXR but not RAR. This regulation is conferred through a specific cis element in the CRBPII promoter that contains five nearly perfect tandem repeats of the sequence AGGTCA spaced by a single nucleotide. The discovery of this new RX response element provides a means for distinguishing between the two retinoid receptor systems and suggests that an RXR-mediated pathway exists for modulating vitamin A metabolism.

Animals↗

PubMed1991

Direct repeats as selective response elements for the thyroid hormone, retinoic acid, and vitamin D3 receptors.

We report here the identification of thyroid hormone response elements (TREs) that consist of a direct repeat, not a palindrome, of the half-sites. Unlike palindromic TREs, direct repeat TREs do not confer a retinoic acid response. The tandem TRE can be converted into a retinoic acid response element by increasing the spacing between the half-sites by 1 nucleotide, and the resulting retinoic acid response element is no longer a TRE. Decreasing the half-site spacing by 1 nucleotide converts the TRE to a vitamin D3 response element, while eliminating response to T3. These results correlate well with DNA-binding affinities of the thyroid hormone, retinoic acid, and vitamin D3 receptors. This study points to the general importance of tandem repeat hormone response elements and suggests a simple physiologic code exists in which half-site spacing plays a critical role in achieving selective hormonal response.

Animals↗

PubMed1990

Jun-Fos and receptors for vitamins A and D recognize a common response element in the human osteocalcin gene.

We present evidence that the vitamin D response element in the human osteocalcin gene confers responsiveness to the vitamin A metabolite, retinoic acid. Retinoic acid receptor (RAR) expressed in E. coli binds to this sequence in vitro. Transfection of RAR expression vectors in cultured cells activates heterologous promoters containing this sequence in vivo. This response element contains a consensus AP-1 site TGACTCA and in vitro is bound by the Jun-Fos complex. Unexpectedly, cotransfection of Jun and Fos expression vectors suppresses basal level transcription of the osteocalcin gene and suppresses induction by both retinoic acid and vitamin D3. Additional studies delimit an 11 nucleotide segment as a minimal hormone response element containing the AP-1 site as its core. These results indicate that two distinct classes of transcription factors can recognize common regulatory sequences, a phenomenon we refer to as cross-coupling.

Animals↗

PubMed1990

A functional retinoic acid receptor encoded by the gene on human chromosome 12.

A cDNA clone derived from a human hepatocellular carcinoma has been isolated on the basis of homology to the alpha human retinoic acid receptor (RAR alpha) gene. Expression of this cDNA produces a high affinity nuclear binding protein for retinoic acid. The product of this clone when expressed in transfected cells is able to activate transcription of a reporter plasmid through specific DNA sequences in response to the addition of retinoic acid to the medium. Dose-dependent profiles upon trans-activation of the reporter indicate that apparent sensitivity to retinoic acid of this protein is approximately 10-fold higher than that of human RAR alpha and is comparable to that of the second human RAR, RAR beta. This gene has been mapped to human chromosome 12, which is distinct from those coding for either alpha or beta RAR, and thus encodes a third human RAR.

Amino Acid Sequence↗

PubMed1989

Comparative and functional anatomy of group II catalytic introns--a review.

The 70 published sequences of group II introns from fungal and plant mitochondria and plant chloroplasts are analyzed for conservation of primary sequence, secondary structure and three-dimensional base pairings. Emphasis is put on structural elements with known or suspected functional significance with respect to self-splicing: the exon-binding and intron-binding sites, the bulging A residue involved in lariat formation, structural domain V and two isolated base pairs, one of them involving the last intron nucleotide and the other one, the first nt of the 3' exon. Separate sections are devoted to the 29 group II-like introns from Euglena chloroplasts and to the possible relationship of catalytic group II introns to nuclear premessenger introns. Alignments of all available sequences of group II introns are provided in the APPENDIX.

Animals↗

PubMed1989

Determinants of target gene specificity for steroid/thyroid hormone receptors.

The molecular specificity of the receptors for steroid and thyroid hormones is achieved by their selective interaction with DNA binding sites referred to as hormone response elements (HREs). HREs can differ in primary nucleotide sequence as well as in the spacing of their dyadic half-sites. The target gene specificity of the glucocorticoid receptor can be converted to that of the estrogen receptor by changing three amino acids clustered in the first zinc finger. Remarkably, a single Gly to Glu change in this region produces a receptor that recognizes both glucocorticoid and estrogen response elements. Further replacement of five amino acids in the stem of the second zinc finger transforms the specificity to that of the thyroid hormone receptor. These findings localize structural determinants required for discrimination of HRE sequence and half-site spacing, respectively, and suggest a simple pathway for the coevolution of receptor DNA binding domains and hormone-responsive gene networks.

Amino Acid Sequence↗

PubMed1989

Steroid hormone receptor homologs in development.

The steroid/thyroid receptor superfamily are ligand-dependent transcription factors which consist of distinct functional domains required for transcriptional control of a network of genes. Members of this superfamily are beginning to be studied for their contribution to embryogenesis. Two human receptors for the vertebrate morphogen retinoic acid have been isolated and further characterized on model promoters. Moreover, the presence of homologs of these receptors in Drosophila reveals that members of this superfamily predate the divergence of the vertebrates and invertebrates. One locus is knirps-related (knrl), whose product is closely related to that of the gap segmentation gene knirps (kni). knrl is one of the most diverged steroid receptor-like molecules and displays a spatially restricted blastoderm pattern.

Animals↗

PubMed1988

Retinoic acid and thyroid hormone induce gene expression through a common responsive element.

Studies of steroid receptors have led to the identification of a superfamily of ligand-inducible regulatory proteins that includes receptors for thyroid hormones and retinoic acid. This family of receptors regulates gene expression through binding to short cis-acting sequences referred to as hormone-response elements. Identification of a functional retinoic acid responsive element is crucial to our understanding of the mechanisms by which retinoic acid receptors activate gene expression and regulate cell differentiation. One impediment to such a study is the absence of any identified gene whose transcription is directly dependent on the receptor-hormone complex. Because the DNA-binding domains of the retinoic acid and thyroid hormone receptors are highly related (62% identical in their amino acid sequences), we have investigated the possibility that the retinoic acid receptor could activate gene expression through a thyroid hormone response element. We now report that a human retinoic acid receptor expressed from cloned complementary DNA or the endogenous retinoic acid receptor present in F9 teratocarcinoma cells can activate gene expression from promoters fused to a natural or synthetic thyroid hormone response element. The product translated in vitro from the human retinoic acid receptor cDNA can bind to a thyroid hormone response element with high affinity. The unexpected implication of these findings is that retinoic acid and thyroid hormones, acting through their respective receptors, could control overlapping gene networks involved in the regulation of vertebrate morphogenesis and homeostasis.

Binding Sites↗