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A Imhof

Publications and source records attributed to A Imhof.

At least 19 recordsLinked to original sources
PubMed2000

dMi-2 and ISWI chromatin remodelling factors have distinct nucleosome binding and mobilization properties.

Mi-2 and ISWI, two members of the Snf2 superfamily of ATPases, reside in separate ATP-dependent chromatin remodelling complexes. These complexes differ in their biochemical properties and are believed to perform distinct functions in the cell. We have compared the remodelling activity of recombinant Drosophila Mi-2 (dMi-2) with that of recombinant ISWI. Both proteins are nucleosome-stimulated ATPases and promote nucleosome mobilization. However, dMi-2 and ISWI differ in their interaction with nucleosome core particles, in their substrate requirements and in the direction of nucleosome mobilization. We have used antibodies to immobilize a complex containing dMi-2 and the dRPD3 histone deacetylase from Drosophila embryo extracts. This complex shares the nucleosome-stimulated ATPase and nucleosome mobilization properties of recombinant dMi-2, demonstrating that these activities are maintained in a physiological context. Its functional properties distinguish dMi-2 from both SWI2/SNF2 and ISWI, defining a new family of ATP-dependent remodelling machines.

Adenosine Triphosphatases↗
PubMed1999

p300 stimulates transcription instigated by ligand-bound thyroid hormone receptor at a step subsequent to chromatin disruption.

We investigate the role of the transcriptional coactivator p300 in gene activation by thyroid hormone receptor (TR) on addition of ligand. The ligand-bound TR targets chromatin disruption independently of gene activation. Exogenous p300 facilitates transcription from a disrupted chromatin template, but does not itself disrupt chromatin in the presence or absence of ligand-bound receptor. Nevertheless, the acetyltransferase activity of p300 is required to facilitate transcription from a disrupted chromatin template. Expression of E1A prevents aspects of chromatin remodeling and transcriptional activation dependent on TR and p300. E1A selectively inhibits the acetylation of non-histone substrates. E1A does not prevent the assembly of a DNase I-hypersensitive site induced by TR, but does inhibit topological alterations and the loss of canonical nucleosome arrays dependent on the addition of ligand. Mutants of E1A incompetent for interaction with p300 partially inhibit chromatin disruption but still allow nuclear receptors to activate transcription. We conclude that p300 has no essential role in chromatin disruption, but makes use of acetyltransferase activity to stimulate transcription at a subsequent step.

Acetyltransferases↗
PubMed1999

Purification and properties of the Xenopus Hat1 acetyltransferase: association with the 14-3-3 proteins in the oocyte nucleus.

We have purified the Xenopus histone acetyltransferase Hat1 holoenzyme from oocytes. The holoenzyme contains the catalytic subunit Hat1, the retinoblastoma associated protein RbAp48, and members of the phosphoserine binding family of 14-3-3 proteins. We have determined that the Hat1 holoenzyme specifically acetylates free histone H4 but not nucleosomal histones. RbAp48 is a phosphoprotein that contains a consensus recognition motif for the 14-3-3 proteins. The 14-3-3 proteins provide a regulatory function for the activity of many phosphoproteins. We find that the hugely abundant Hat1 holoenzyme is present in 10 000-fold excess over somatic cell levels. The holoenzyme is localized in the oocyte nucleus where acetylated histones are stored. The oocyte form of the Xenopus Hat1 holoenzyme may represent a specialized storage form of histone acetyltransferase. Following oocyte maturation and subsequent embryogenesis, the Hat1 enzyme is redistributed to the cytoplasm, where new histones are synthesized.

14-3-3 Proteins↗
PubMed1999

[A female patient with splenomegaly, interstitial pneumopathy and giant foam cells in bone marrow].

We describe a case of Niemann-Pick disease type B. A 13-year-old female adolescent of Turkish origin suffered from abdominal pain for several months, finally leading to hospitalisation. The investigations revealed splenomegaly and interstitial pneumopathy. The bone marrow contained giant foam cells typical of Niemann-Pick disease. Enzymatic analysis of a fibroblast culture confirmed the diagnosis of Niemann-Pick disease type B, with reduced activity of acid sphingomyelinase. Niemann-Pick disease is an inherited autosomal recessive lysosomal storage disorder of sphingolipids, resulting in an accumulation of sphingomyelin in the cells of the reticulo-histiocytic system due to an enzymatic defect. In Niemann-Pick disease type B the spleen and lung are the main organs affected. At present no treatment exists for this disorder.

Adolescent↗
PubMed1999

Risk factors for type I diabetes mellitus in children in Austria.

UNLABELLED: The aim of this study was to investigate environmental risk factors in the development of type 1 diabetes mellitus in a population-based case-control study. Parents of all patients with manifestation of type 1 diabetes between 1989 and 1994 in Vienna were asked to complete a questionnaire (n = 114). Control children (n = 495), matched for age and sex, were randomly recruited from all schools in Vienna. Fathers of diabetic children were significantly older at the time their children were born than fathers of control children (P = 0.015). Children with diabetes were more likely to be second- or third-born children (P<0.05) and fewer went to kindergarten than the control group children (P = 0.007). No significant difference in duration of gestation, percentage of delivery by caesarean section, birth weight or length was found. Neonatal jaundice was more often observed in the patient group (P = 0.038). Breast feeding was reported by 82.7% of mothers of diabetic children and by 81% of mothers of control children, and the duration of breast feeding was longer in patients than in controls (n.s.). CONCLUSION: In our study, the development of type 1 diabetes mellitus was associated with higher paternal age and neonatal jaundice. No correlation could be found with dietary intake of cow's milk products in early infancy, vaccination and other environmental factors.

Adolescent↗
PubMed1999

Transcriptional regulation of the AP-2alpha promoter by BTEB-1 and AP-2rep, a novel wt-1/egr-related zinc finger repressor.

AP-2 transcription factors have been suggested to exert key regulatory functions in vertebrate embryonic development, in tumorigenicity of various cancer cell types, and in controlling cell cycle and apoptotic effector genes. In this study, we investigated transcriptional regulation of the AP-2alpha gene promoter mediated by an autoregulatory element (referred to as A32) with a core consensus AP-2 binding site at position -336 relative to the mRNA initiation site. AP-2 and multiple different nuclear proteins in HeLa and Neuro2A cell extracts form specific bandshifts with the A32 element. By screening a mouse brain cDNA expression library, we isolated two different cDNAs encoding the transcription factor BTEB-1 and a novel zinc finger protein, AP-2rep. AP-2rep reveals a modular structure with homology to transcription factors of the wt-1/egr-1-family. AP-2rep, BTEB-1, and AP-2 interact in a mutually exclusive manner with overlapping binding sites in the A32 element. Transfection studies revealed that BTEB-1 is a strong activator of AP-2alpha promoter activity, whereas cotransfected AP-2alpha resulted in moderate autoactivation of promoter activity. In contrast, AP-2rep confers strong transcriptional repression to the AP-2alpha gene, and we observed an excellent correlation between induction of AP-2rep mRNA expression and downregulation of AP-2alpha mRNA during development of the kidney. In summary, we have identified multiple transcription factors and cloned from an expression library a novel zinc finger silencing factor, AP-2rep, mediating positive and negative regulation of AP-2alpha expression through a set of overlapping cis-regulatory promoter elements.

Amino Acid Sequence↗
PubMed1998

Transcription: gene control by targeted histone acetylation.

A transcriptional regulator in yeast, Gcn5p, activates transcription by targeted acetylation of specific lysine residues in the amino-terminal tails of histones. This targeted modification is restricted to nucleosomes assembled on the promoters of Gcn5p-responsive genes.

Acetylation↗
PubMed1998

Distinct requirements for chromatin assembly in transcriptional repression by thyroid hormone receptor and histone deacetylase.

Histone deacetylase and chromatin assembly contribute to the control of transcription of the Xenopus TRbetaA gene promoter by the heterodimer of Xenopus thyroid hormone receptor and 9-cis retinoic acid receptor (TR-RXR). Addition of the histone deacetylase inhibitor Trichostatin A (TSA) relieves repression of transcription due to chromatin assembly following microinjection of templates into Xenopus oocyte nuclei, and eliminates regulation of transcription by TR-RXR. Expression of Xenopus RPD3p, the catalytic subunit of histone deacetylase, represses the TRbetaA promoter, but only after efficient assembly of the template into nucleosomes. In contrast, the unliganded TR-RXR represses templates only partially assembled into nucleosomes; addition of TSA also relieves this transcriptional repression. This result indicates the distinct requirements for chromatin assembly in mediating transcriptional repression by the deacetylase alone, compared with those needed in the presence of unliganded TR-RXR. In addition, whereas hormone-bound TR-RXR targets chromatin disruption as assayed through changes in minichromosome topology and loss of a regular nucleosomal ladder on micrococcal nuclease digestion, addition of TSA relieves transcriptional repression but does not disrupt chromatin. Thus, TR-RXR can facilitate transcriptional repression in the absence of hormone through mechanisms in addition to recruitment of deacetylase, and disrupts chromatin structure through mechanisms in addition to the inhibition or release of deacetylase.

Amino Acid Sequence↗
PubMed1997

Acetylation of general transcription factors by histone acetyltransferases.

The acetylation of histones increases the accessibility of nucleosomal DNA to transcription factors [1,2], relieving transcriptional repression [3] and correlating with the potential for transcriptional activity in vivo [4 - 7]. The characterization of several novel histone acetyltransferases - including the human GCN5 homolog PCAF (p300/CBP-associated factor) [8], the transcription coactivator p300/CBP [9], and TAFII250 [10] - has provided a potential explanation for the relationship between histone acetylation and transcriptional activation. In addition to histones, however, other components of the basal transcription machinery might be acetylated by these enzymes and directly affect transcription. Here, we examine the acetylation of the basal transcriptional machinery for RNA polymerase II by PCAF, p300 and TAFII250. We find that all three acetyltransferases can direct the acetylation of TFIIEbetaand TFIIF, and we identify a preferred site of acetylation in TFIIEbeta. Human TFIIE consists of two subunits, alpha(p56) and beta(p34), which form a heterotetramer (alpha2 beta2) in solution ([11], reviewed in [12]). TFIIE enters the preinitiation complex after RNA polymerase II and TFIIF, suggesting that TFIIE may interact directly with RNA polymerase II and/or TFIIF [13,14]. In addition, TFIIE can facilitate promoter melting either in the presence or absence of TFIIH and can stimulate TFIIH-dependent phosphorylation of the carboxy-terminal domain of RNA polymerase II [15-18]. TFIIF has an essential role in both transcription initiation and elongation ([19,20], for review see [21]). We discuss the implications of the acetylation of TFIIEbetaand TFIIF for transcriptional control by PCAF, p300 and TAFII250.

Acetyl Coenzyme A↗
PubMed1997

Stability of Nonaqueous Emulsions

We examined the stability of emulsions of oil in several nonaqueous polar liquids using commercially available nonionic surfactants. Stable nonaqueous emulsions were only obtained with formamide and dimethylsulfoxide. Hydrogen bonding, and not polarity, appears to be the important factor determining the emulsifying power of a solvent. Ostwald ripening plays a much more important role in the stability of these nonaqueous emulsions than in the corresponding aqueous systems. This destabilizing process can be prevented, however, by addition to the oil phase of a small amount (1%) of an oil that has a very low solubility in the continuous phase. Furthermore, a larger size of the surfactant molecule protects emulsions against droplet coalescence. Thus, emulsions in formamide and dimethylsulfoxide did not show any breakdown when stabilized with a triblock copolymer of polyoxyethylene-polyoxypropylene-polyoxyethylene. Copyright 1997Academic Press

Journal Article↗
PubMed1996

Continuous monitoring of oxygen concentrations in several systems for cultivation of anaerobic bacteria.

Anaerobic chambers and jars are the two conventional methods used in clinical microbiology laboratories to produce an anaerobic atmosphere. The evacuation-replacement method, the Oxoid AnaeroGen, the Merck Anaerocult A, the BBL GasPak, the BBL GasPakPlus, the Adams Scientific GasGendicator, the Difco Anaerobic, and the bioMérieux Generbox anaer systems were compared for the timed decrease in the oxygen concentration in an anaerobic jar. The experiment was repeated 10 times for each system. The oxygen concentration was measured with an oxygen analyzer series 3600 instrument (Orbisphere Laboratories, Neuchâtel-Geneva Switzerland). The BBL GasPak, the BBL GasPakPlus, the bioMérieux Generbox, the Adams Scientific GasGendicator, and the Difco Anaerobic systems contain sodium borohydride, which liberates hydrogen. The Anaerocult A system contains iron powder which binds the oxygen. The active ingredient of the AnaeroGen system is ascorbic acid. The times to reach an O2 concentration of 0.5% were 8 to 15 min for the evacuation-replacement method, 26 to 41 min for the AnaeroGen system, 60 to 93 min for the Anaerocult A system, and 22 to 419 min for the sodium borohydride systems. The AnaeroGen system, the Anaerocult A system, and the evacuation-replacement method never failed to produce an anaerobic atmosphere. The sodium borohydride systems failed in 10 of 70 runs. These results suggest that the evacuation-replacement method or the Oxoid AnaeroGen system should be used to produce an anaerobic atmosphere. The Anaerocult A system showed a good reproducibility, but the length of time required to reach an appropriately low level of oxygen was too long. Because of the high failure rate, the borohydride systems cannot be recommended.

Anaerobiosis↗
PubMed1995

Transcriptional activation by Myc is under negative control by the transcription factor AP-2.

The Myc protein binds to and transactivates the expression of genes via E-box elements containing a central CAC(G/A)TG sequence. The transcriptional activation function of Myc is required for its ability to induce cell cycle progression, cellular transformation and apoptosis. Here we show that transactivation by Myc is under negative control by the transcription factor AP-2. AP-2 inhibits transactivation by Myc via two distinct mechanisms. First, high affinity binding sites for AP-2 overlap Myc-response elements in two bona fide target genes of Myc, prothymosin-alpha and ornithine decarboxylase. On these sites, AP-2 competes for binding of either Myc/Max heterodimers or Max/Max homodimers. The second mechanism involves a specific interaction between C-terminal domains of AP-2 and the BR/HLH/LZ domain of Myc, but not Max or Mad. Binding of AP-2 to Myc does not preclude association of Myc with Max, but impairs DNA binding of the Myc/Max complex and inhibits transactivation by Myc even in the absence of an overlapping AP-2 binding site. Taken together, our data suggest that AP-2 acts as a negative regulator of transactivation by Myc.

Animals↗
PubMed1995

Cloning and characterization of a second AP-2 transcription factor: AP-2 beta.

AP-2 has been characterized previously as a unique 52 x 10(3) M(r) transcription activator encoded by a single gene that is expressed in a restricted pattern during embryonic morphogenesis of the peripheral nervous system, face, skin and nephric tissues. Here we report the isolation of genomic and cDNA clones encoding for a second AP-2 related transcription factor, designated AP-2 beta. AP-2 beta binds specifically to a series of well-characterized AP-2 binding sites, consensus to the sequence G/CCCN3GGC, and transactivates transcription from a reporter plasmid under the control of an AP-2-dependent promoter. A C-terminal domain known to mediate homodimerization of the previously cloned AP-2 alpha transcription activator is highly conserved and sufficient to mediate interaction between the two proteins. Northern blot and in situ hybridizations revealed that the two genes are expressed in murine embryos between days 9.5 and 19.5 p.c. Coexpression of both mRNAs was detected in many tissues at day 13.5 and 15.5 of embryogenesis but some regions of the developing brain and face including the primordium of midbrain and the facial mesenchyme differed in their expression pattern of AP-2 genes. AP-2 alpha and AP-2 beta signals in the central and peripheral nervous system overlapped with regions of developing sensory neurons. In adult tissues AP-2 alpha expression was found mainly in the skin, eye and prostate and AP-2 beta expression in the kidney. In summary, our analyses of embryonic and adult mice demonstrate that two different AP-2 transcription factors are specifically expressed during differentiation of many neural, epidermal and urogenital tissues.

Amino Acid Sequence↗
PubMed1994

The genomic structure of the human AP-2 transcription factor.

The transcription factor AP-2 is encoded by a gene located on chromosome 6 near the HLA locus. Here we describe the genomic organization of the AP-2 gene including an initial characterization of the promoter. We have mapped two mRNA initiation sites, the entire exon-intron structure and located two polyadenylation sites. The mature AP-2 mRNA is spliced from 7 exons distributed over a region of 18 kb genomic DNA. A recently cloned inhibitory AP-2 protein is generated by alternative usage of a C-terminal exon. The proline-rich transactivation motif is encoded by a single exon within the N-terminal region in contrast to the complex DNA binding and dimerization motif which involves amino acid residues located on four different exons. The sites of mRNA initiation are located 220 and 271 bases upstream from the ATG translation start site. Although the promoter contains no canonical sequence motifs for basal transcription factors, such as TATA-, CCAAT- or SP-1 boxes, it mediates cell-type-specific expression of a CAT reporter gene in PA-1 human teratocarcinoma cells and is inactive in murine F9 teratocarcinoma cells. We demonstrate that the promoter of the AP-2 gene is subject to positive autoregulation by its own gene product. A consensus AP-2 binding site is located at position -622 with respect to the ATG. This site binds specifically to bacterially expressed AP-2 as well as to multiple proteins, including AP-2, present in PA-1 and HeLa cell nuclear extracts. A partial AP-2 promoter fragment including the AP-2 consensus binding site is approximately 5-fold transactivated by cotransfection of an AP-2 expression plasmid.

Animals↗