(1/142) The orphan nuclear receptor COUP-TFII is required for angiogenesis and heart development.
The embryonic expression of COUP-TFII, an orphan nuclear receptor, suggests that it may participate in mesenchymal-epithelial interactions required for organogenesis. Targeted deletion of the COUP-TFII gene results in embryonic lethality with defects in angiogenesis and heart development. COUP-TFII mutants are defective in remodeling the primitive capillary plexus into large and small microcapillaries. In the COUP-TFII mutant heart, the atria and sinus venosus fail to develop past the primitive tube stage. Reciprocal interactions between the endothelium and the mesenchyme in the vascular system and heart are essential for normal development of these systems. In fact, the expression of Angiopoietin-1, a proangiogenic soluble factor thought to mediate the mesenchymal-endothelial interactions during heart development and vascular remodeling, is down-regulated in COUP-TFII mutants. This down-regulation suggests that COUP-TFII may be required for bidirectional signaling between the endothelial and mesenchymal compartments essential for proper angiogenesis and heart development. (+info)
(2/142) Heterodimeric interactions between chicken ovalbumin upstream promoter-transcription factor family members ARP1 and ear2.
Members of the chicken ovalbumin upstream promoter-transcription factor (COUP-TF) subfamily of orphan nuclear receptors, which minimally includes COUP-TFI and ARP1, are highly expressed in brain and are generally considered to be constitutive repressors of transcription. We have used a yeast two-hybrid system to isolate proteins expressed in brain that interact with ARP1. One of the proteins isolated in this screen was Ear2, another orphan receptor that has been suggested to be a member of the COUP-TF subfamily. Here we demonstrate that ARP1 and Ear2 form heterodimers in solution and on directly repeated response elements with high efficiency and a specificity differing from that of homodimeric complexes composed of either receptor. ARP1 and Ear2 were observed to interact in mammalian cells, and the tissue distribution of Ear2 transcripts was found to overlap precisely with the expression pattern of ARP1 in several mouse tissues and embryonal carcinoma cell lines. Heterodimeric interactions between ARP1 and Ear2 may define a distinct pathway of orphan receptor signaling. (+info)
(3/142) Functional study of the E276Q mutant hepatocyte nuclear factor-4alpha found in type 1 maturity-onset diabetes of the young: impaired synergy with chicken ovalbumin upstream promoter transcription factor II on the hepatocyte nuclear factor-1 promoter.
Seven mutations in the hepatocyte nuclear factor (HNF)-4alpha gene have been shown to correlate with type 1 maturity-onset diabetes of the young (MODY 1), a monogenic form of type 2 diabetes. Up to now, only the functional properties of two MODY 1 HNF-4alpha mutants, Q268X and V393I, have been investigated to address how the mutations in the HNF-4alpha gene, found by genetic studies, can give rise to impaired activities of mutated HNF-4alpha proteins and can cause this disease. The E276Q mutation results in a nonconservative substitution occurring in the HNF-4alpha E domain, which is involved in dimerization and transactivation activities as well as in protein-protein interactions with other transcription factors or coactivators. Using the mutated human HNF-4alpha2, we have found that, in the absence of chicken ovalbumin upstream promoter transcription factor II (COUP TFII), the E276Q substitution does not significantly affect the dimerization and transactivating activities of HNF-4alpha, at least on the promoters studied herein. On the other hand, in the presence of COUP TFII, the substitution impairs the enhancement of HNF-4-mediated activation of HNF-1 promoter. The impaired synergy between COUP TFII and HNF-4 on the HNF-1 promoter results from an alteration of their interaction. HNF-1 expression plays a crucial role in transactivation of insulin promoter and of numerous genes coding for enzymes involved in glucose homeostasis. Therefore, its downregulation resulting from the E276Q mutation in HNF-4alpha gene most probably impairs the function of pancreatic beta-cells. (+info)
(4/142) Dorsal and ventral retinal territories defined by retinoic acid synthesis, break-down and nuclear receptor expression.
Determination of the dorso-ventral dimension of the vertebrate retina is known to involve retinoic acid (RA), in that high RA activates expression of a ventral retinaldehyde dehydrogenase and low RA of a dorsal dehydrogenase. Here we show that in the early eye vesicle of the mouse embryo, expression of the dorsal dehydrogenase is preceded by, and transiently overlaps with, the RA-degrading oxidase CYP26. Subsequently in the embryonic retina, CYP26 forms a narrow horizontal boundary between the dorsal and ventral dehydrogenases, creating a trough between very high ventral and moderately high dorsal RA levels. Most of the RA receptors are expressed uniformly throughout the retina except for the RA-sensitive RARbeta, which is down-regulated in the CYP26 stripe. The orphan receptor COUP-TFII, which modulates RA responses, colocalizes with the dorsal dehydrogenase. The organization of the embryonic vertebrate retina into dorsal and ventral territories divided by a horizontal boundary has parallels to the division of the Drosophila eye disc into dorsal, equatorial and ventral zones, indicating that the similarities in eye morphogenesis extend beyond single molecules to topographical patterns. (+info)
(5/142) Chicken ovalbumin upstream promoter-transcription factor II, a new partner of the glucose response element of the L-type pyruvate kinase gene, acts as an inhibitor of the glucose response.
Transcription of the L-type pyruvate kinase (L-PK) gene is induced by glucose in the presence of insulin and repressed by glucagon via cyclic AMP. The DNA regulatory sequence responsible for mediating glucose and cyclic AMP responses, called glucose response element (GlRE), consists of two degenerated E boxes spaced by 5 base pairs and is able to bind basic helix-loop-helix/leucine zipper proteins, in particular the upstream stimulatory factors (USFs). From ex vivo and in vivo experiments, it appears that USFs are required for correct response of the L-PK gene to glucose, but their expression and binding activity are not known to be regulated by glucose. A genetic screen in yeast has allowed us to identify a novel transcriptional factor binding to the GlRE, i.e. the chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII). Binding of COUP-TFII to the GlRE was confirmed by electrophoretic mobility shift assays, and COUP-TFII-containing complexes were detectable in liver nuclear extracts. Neither abundance nor binding activity of COUP-TFII appeared to be significantly regulated by diets. In footprinting experiments, two COUP-TFII-binding sites overlapping the E boxes were detected. Overexpression of COUP-TFII abrogated the USF-dependent transactivation of an artificial GlRE-dependent promoter in COS cells and the glucose responsiveness of the L-PK promoter in hepatocytes in primary culture. In addition, a mutated GlRE with increased affinity for USF and very low affinity for COUP-TFII conferred a dramatically decreased glucose responsiveness on the L-PK promoter in hepatocytes in primary culture by increasing activity of the reporter gene in low glucose condition. We propose that COUP-TFII could be a negative regulatory component of the glucose sensor complex assembled on the GlRE of the L-PK gene and most likely of other glucose-responsive genes as well. (+info)
(6/142) Identification of COUP-TF as a transcriptional repressor of the c-mos proto-oncogene.
The c-mos proto-oncogene is specifically expressed in the male and female germ cells of the mouse and other vertebrates. We previously identified a 15-base pair sequence element (B2) as the binding site of a candidate repressor of c-mos transcription in somatic cells. In the present study, we used the yeast one-hybrid system to isolate HeLa cell cDNAs encoding proteins that specifically bound to the c-mos B2 element. Nucleotide sequencing identified several of the clones isolated in this screen as the orphan nuclear receptors COUP-TFI and COUP-TFII. A COUP-TF-binding site was then identified within the B2 sequence. Complexes formed between purified COUP-TFs and the c-mos B2 probe comigrated in electrophoretic mobility shift assays with those formed using whole nuclear extracts of NIH 3T3 or HeLa cells. Moreover, the complexes formed with NIH 3T3 nuclear extracts and B2 probe were supershifted with antibody against COUP-TF, identifying COUP-TF as the candidate repressor previously detected in these somatic cell extracts. Substitution of a consensus COUP-TF-binding site for the c-mos negative regulatory element suppressed expression from the c-mos promoter in transfected somatic cells, demonstrating the functional activity of COUP-TF as a repressor of c-mos transcription. (+info)
(7/142) HNF4 and COUP-TFII interact to modulate transcription of the cholesterol 7alpha-hydroxylase gene (CYP7A1).
The gene for cholesterol 7alpha-hydroxylase (CYP7A1) contains a sequence at nt -149 to -118 that was found to play a large role in determining the overall transcriptional activity and regulation of the promoter. Hepatocyte nuclear factor 4 (HNF4) and chicken ovalbumin upstream promoter transcription factor II (COUP-TFII) synergistically activate transcription of the CYP7A1 promoter. Transactivation of CYP7A1 by HNF4 in the human hepatoma cell line, HepG2, was enhanced by cotransfection with COUP-TFII or the basal transcription element binding protein (BTEB). HNF4 prepared from rat liver nuclear extracts bound to oligomers homologous to the nt -146 to -134 sequences in electrophoretic mobility shift assays (EMSA), which corresponded to a conserved region containing a direct repeat of hormone response elements spaced by one nucleotide (DR1). The sequences surrounding this DR1 were found to be essential for the HNF4 transactivation. In vitro-translated COUP-TFII was found to bind the adjacent sequences from nt -139 to -128 (DR0), but COUP-TFII interacted with this region at a much lower affinity than to the COUP-TFII-site at nt -72 to -57 (DR4). Mutations at nt -139 to -128 or nt -72 to -57 reduced the COUP-TFII and HNF4 synergy; however, these COUP-TFII-binding sequences were not absolutely required for the cooperative effect of HNF4 and COUP-TFII on transactivation. These results indicated that the observed transactivation was the result of protein/protein interactions facilitated by the juxtaposition of the binding elements. (+info)
(8/142) ARP1 in Golgi organisation and attachment of manchette microtubules to the nucleus during mammalian spermatogenesis.
Actin related protein of vertebrate, Arp1, is a major component of the dynactin complex. To characterise and localise Arp1 during mammalian spermatogenesis, polyclonal antibodies were raised against a human recombinant Arp1. Anti-Arp1 antibodies were used for western-immunoblotting, indirect immunofluorescence and immunoelectron microscopy. In round spermatids, Arp1 was detected at the centrosome and at the Golgi apparatus. In elongated spermatids, Arp1 was predominantly found along microtubules of the manchette and at their site of attachment to the nuclear envelope. In maturing spermatids, Arp1 was still present in the pericentriolar material, but in testicular spermatozoa it was not detectable. These various localisations of Arp1 and their changes during spermatid differentiation suggest that the dynactin complex in association with dynein might contribute to several activities: the functional organisation of the centrosome and of the Golgi apparatus and the shaping of the nucleus by manchette microtubules. (+info)