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T Igo-Kemenes

Publications and source records attributed to T Igo-Kemenes.

At least 19 recordsLinked to original sources
PubMed1998

Characterization of the chicken and rat DCoH gene domains using an improved ligation-mediated PCR method.

The bifunctional protein DCoH (dimerization cofactor of HNF-1) is a transcriptional coactivator of homeodomain proteins and a cytoplasmic enzyme. Here we report on the isolation, nucleotide sequence and genomic structure of the chicken and rat DCoH genes. They have an unusual first exon coding for a single amino acid. The promoters are located within CpG-islands and lack TATA and CAAT consensus sequences. Analysis of the number of CpG dinucleotides in the promoter indicates little changes since man and chicken were separated in evolution whereas a 67% reduction was observed in the rat lineage. The DNA sequences were obtained entirely without cloning. For walking into unknown genomic regions we describe a modification of the ligation-mediated polymerase chain reaction (LM-PCR) that can replace conventional screening and cloning methods and greatly expedite the isolation of genomic DNA.

Animals↗
PubMed1993

Hepatic nuclear factor 1 alpha (HNF-1 alpha) is expressed in the oviduct of hens and interacts with regulatory elements of the lysozyme gene.

HNF-1 alpha is a nuclear transcriptional regulatory protein required for the expression of a variety of liver-specific genes. This factor was previously considered liver-specific but later shown to be expressed also in a few other mammalian tissues. Here we report on the occurrence of HNF-1 alpha in the avian oviduct. This finding is of particular interest because HNF-1 alpha is not expressed in female reproductive organs of mammals. The avian oviduct is the site of assembly for the avian egg and the site of tissue-specific synthesis of the major egg white proteins, such as lysozyme. We also demonstrate that the chicken lysozyme gene contains HNF-1 recognition sites within two of its important upstream regulatory elements. The presence of HNF-1 recognition elements in functionally significant regulatory sites of the lysozyme gene and high levels of HNF-1 alpha in the oviduct is a strong indication for the involvement of HNF-1 alpha in the control of the lysozyme gene and possibly other egg white protein genes in the chicken oviduct. Apparently, HNF-1 alpha performs functions in the avian oviduct that were lost upon development from birds to mammals.

Amino Acid Sequence↗
PubMed1993

Genomic structure of the POU-related hepatic transcription factor HNF-1 alpha.

Hepatocyte Nuclear Factor-1 alpha (HNF-1 alpha) is a vertebrate transcriptional regulatory protein which plays a prominent role in the activation of a large family of hepatocyte-specific genes. It is the prototype of a new subfamily of POU-related proteins important in embryogenesis and cell-type specification. Here we report on the genomic structure and DNA sequence of HNF-1 alpha of the hen. Functional domains are non-uniformly conserved between hen and mammals consistent with the notion of functions in the hen oviduct that were lost in mammals after avian mammalian separation. The unusual homeo-related DNA-binding domain is well preserved between birds and mammals. The exon/intron pattern of the gene changed during evolution. The rat gene consists of 9 exons, the chicken gene of 10. In mammals one intron was eliminated that in hens subdivides the serine-rich transactivation domain. Comparison of the sequences at this junction indicates the involvement of elements of the V(D)J recombination system in the process of intron elimination.

Amino Acid Sequence↗
PubMed1987

Partial overlapping of binding sequences for steroid hormone receptors and DNaseI hypersensitive sites in the rabbit uteroglobin gene region.

Four DNaseI hypersensitive (HS) chromatin regions were found in the uteroglobin locus located at -3.7, -2.4, -0.1 and +4.1 kb with respect to the transcription start site of the gene. The three sites upstream of the gene are only detected in the hormonally stimulated endometrium and disappear after hormone withdrawal, whereas the site at +4.1 is also found in tissues that do not express uteroglobin. In the -2.4 HS region, which is strictly dependent on progesterone treatment, three DNaseI sites are clustered within a 240 bp DNA segment that contains 20 imperfect repeats of an octanucleotide motif. Upstream of the uteroglobin gene there are three regions containing binding sites for the glucocorticoid and the progesterone receptors, located at -3.7, -2.6/-2.7 and -2.4. The -2.4 region contains two binding sites for the hormone receptors flanking the central HS site. In footprinting experiments with naked DNA binding of the receptor also renders this site more susceptible towards digestion with DNaseI. The -2.6/-2.7 region contains three binding sites for the hormone receptors located 140 bp upstream of the HS -2.4. While the -3.7 HS is also located within a receptor binding fragment, there is no binding of the hormone receptors to the promoter region. Thus, interaction of the receptor with DNA sequences far upstream from the promoter alters the chromatin conformation of neighbouring sequences and results in transcriptional activation.

Animals↗
PubMed1987

DNase I-hypersensitive sites in the chromatin structure of the lysozyme gene in steroid hormone target and non-target cells.

The pattern of DNase I-hypersensitive sites in the chromatin domain of the lysozyme gene was investigated in several organs and cell-types of the chicken. In the cluster of hypersensitive chromatin sites framing the gene, different classes of sites could be discerned: A subset was common to essentially all cells examined except for erythrocytes. Thus several highly nuclease susceptible structures exist around the gene even in its repressed state. Beside the promoter region a second site 6.1 kb upstream of the transcriptional start site of the gene strictly correlates with the transcriptionally active or potentially active state of the gene in both, oviduct cells and macrophages. A final class of sites is specific for the particular lysozyme expressing tissue and the presence of its members distinguish whether the gene is steroid regulated or in a steroid independent active mode.

Animals↗
PubMed1986

The DNase I sensitive domain of the chicken lysozyme gene spans 24 kb.

We have determined the DNase I sensitive chromatin domain of the lysozyme gene in the hen oviduct. When nuclei were digested with DNase I, about 14 kb of upstream and 6 kb of downstream sequences in addition to the 4 kb long transcribed region were preferentially degraded. The transcription start site is located near the center of the approximately 24 kb long sensitive domain. At the 3' boundary there is a rather abrupt transition from the DNase I sensitive to the resistant chromatin configuration whereas at the 5' border this transition occurs in a gradual fashion over 6-7 kb of DNA. No obvious correlation between the boundaries of the domain and repetitive sequences could be established. DNase I-hypersensitive sites are clustered within the boundaries of the sensitive domain which seems to represent a functional unit of the gene.

Animals↗
PubMed1984

Positioning of nucleosomes in satellite I-containing chromatin of rat liver.

The location of nucleosomes on rat satellite I DNA has been investigated using a new approach. Nucleosome cores were prepared from rat liver nuclei with micrococcal nuclease, exonuclease III and nucleases S1. From the total population of core DNA fragments the satellite-containing fragments were isolated by molecular cloning and the complete sequence of 50 clones was determined. The location of nucleosomes along the satellite sequence was found to be non-random. Our results show that nucleosomes occupy a number of positions on satellite I DNA. About 35 to 50% of all nucleosomes are positioned in two corresponding major sites, the remainder in about 16 less preferred sites. The major nucleosome positions are apparently strictly defined with the precision of a single base-pair. These results were confirmed by other approaches, including restriction nuclease digestion experiments. There are good indications of a defined long-range organization of the satellite chromatin fiber in two or more oligonucleosomal arrays with distinct nucleosome configurations.

Animals↗
PubMed1983

Nuclease-hypersensitive sites in the chromatin domain of the chicken lysozyme gene.

We have examined the chromatin structure of a 22 kilobase-pair chromosomal region containing the lysozyme gene in laying hen. Nuclease-hypersensitive sites were probed with DNAase I by using an indirect end-labeling technique. Eight DNAase I-hypersensitive sites could be mapped in the flanking regions of the gene in oviduct cells, in which the gene is expressed. The same sensitive sites were detected by utilization of an endogenous nuclease activity present in oviduct nuclei. Only one hypersensitive site was detected in the chromatin from erythrocytes, in which the gene is not expressed. The 3'-terminus of the lysozyme gene is highly exposed in nuclei from both tissues. Of special interest is the hypersensitive site at the 5'-terminus of the actively transcribed gene since it maps at the region of multiple initiation sites of transcription and the putative control regions of steroid hormones. DNAase I-hypersensitive chromatin structures also at a greater distance from the gene may take part in the control of gene expression.

Animals↗
PubMed1982

Chromatin.

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Animals↗
PubMed1980

Different repeat lengths in rat satellite I DNA containing chromatin and bulk chromatin.

The nucleosome repeat structure of a rat liver chromatin component containing the satellite I DNA (repeat length 370 bp) was investigated. Digestion experiments with micrococcal nuclease, DNAase II, and the Ca2+/Mg2+-dependent endogenous nuclease of rat liver nuclei revealed a repeat unit of 185 nucleotide pairs which is shorter by approximately 10 bp than the repeat unit of the bulk chromatin of this cell type. The difference seems not to be related to the histone composition which was found to be similar in the two types of chromatin.

Animals↗
PubMed1980

Non-random arrangement of nucleosomes in satellite I containing chromatin of rat liver.

The location of nucleosomes on the nucleotide sequence of rat satellite I DNA was investigated using micrococcal nuclease, exonuclease III, and restriction nucleases as tools. Hae III cleaved the satellite DNA containing chromatin very preferentially in the linker region. Nucleosomes were found predominantly in three defined positions on the 370 bp satellite I monomer unit. This type of arrangement occurs on not more than half of the satellite DNA containing chromatin while the rest of this chromatin is arranged differently. The arrangement of nucleosomes with high probability in preferred frames and with low probability in less preferred frames may be a general phenomenon which can be discussed as a possible mechanism to modulate sequence recognition.

Animals↗
PubMed1979

Nucleotide sequence of a highly repetitive component of rat DNA.

A highly repetitive component of rat DNA which could not yet be enriched by density gradient centrifugation was isolated with the help of the restriction nuclease Sau3AI. This nuclease converted the bulk of the DNA to small fragments and left a repetitive DNA component as large fragments which were subsequently purified by gel filtration and electrophoresis. This DNA component which was termed rat satellite DNA I is composed of tandemly repeated 370 bp blocks. According to sequence analysis the 370 bp repeats consist of alternating 92 and 93 bp units with homologous but not identical sequences. Methylation of CpG residues was correlated to the rate of cleavage by restriction nucleases. Significant homologies exist between the sequences of rat satellite DNA I and satellite DNAs of several other organisms. The divergence of the sequence of rat satellite DNA I was discussed with respect to evolutionary considerations.

Animals↗
PubMed1979

The non-histone proteins of the rat liver nucleus and their distribution amongst chromatin fractions as produced by nuclease digestion.

The search for proteins involved in maintaining higher order chromatin structures has led to a systematic examination of the non-histone proteins (NHP) of rat liver nuclei in the context of nuclease digestion studies. 40-45% of the 3H-tryptophan labelled NHP originally present could be removed by extensive washing in a "physiological" buffer, incubation at 37 degrees C with or without nuclease and a further wash step. Nuclei at this stage had a remarkably constant NHP content (ca. 0.73 micrograms/micrograms DNA), independent of the degree of digestion with micrococcal nuclease or HaeIII. The solubilized chromatin produced by limited digestion with either nuclease contained 0.3-0.5 microgram NHP/microgram DNA, this value falling to ca. 0.16 after more extensive cleavage. Insoluble chromatin fractions were between 2-fold (very limited digestion) and 16-fold (extensive digestion) richer in NHP than the corresponding soluble fractions. Gel electrophoresis revealed about 12 NHP bands in soluble fractions, the most prominent of M.Wt. 41.400, while the insoluble material had at least 50 components. These properties were independent of whether lysis of nuclei occurred in 0.2 or 50 mM ionic strength. The large disparity in NHP content between complementary soluble and insoluble chromatin fractions is considered in terms of chromatin organization in vivo and the possible role of NHP migration.

Animals↗
PubMed1978

Characterization of restriction nuclease prepared chromatin by electron microscopy.

Chromatin was solubilized from rat liver nuclei by digestion with the restriction nuclease EcoRI or HaeIII in the presence or absence of EDTA and sodium chloride. The samples were investigated by electron microscopy after positive and negative staining with uranyl acetate under a number of conditions. Depending on the salt concentration during solubilization the chromatin appeared as beads on the string or in more compact form. Solenoid- and superbead-like structures were seen as had been reported for chromatin solubilized with micrococcal nuclease.

Animals↗
PubMed1977

Prepartation of soluble chromatin and specific chromatin fractions with restriction nucleases.

By digestion of rat liver nuclei with EndoR HaeIII, EndoR EcoRI, and EndoR Bam and subsequent lysis of the nuclei approx. 90%, 40%, and 45%, respectively, of the chromatin were solubilized. The plateau values of solubilization are in agreement with a model in which the chromatin strands are crosslinked and/or attached to a supporting structure. The distribution of DNA lengths in the soluble and insoluble chromatin fractions were determined. According to digestion experiments with restriction nucleases rat liver DNA contains highly repetitive sequences, some of which are arranged in tandem repeats of 95 and 380 nucleotide pairs, respectively. With EndoR EcoRI chromatin containing the repetitive RNA was preferentially solubilized and, by subsequent sucrose gradient centrifugation, purified to about 90%. The useful properties of chromatin prepared by the specific action of restriction nucleases are discussed.

Animals↗
PubMed1976

Nuclease digestion in between and within nucleosomes.

In the course of digestions of rat liver nuclei with micrococcal nuclease the size of the nucleosomal DNA is shortened by 50-60 nucleotide pairs from starting lengths of about 200, 400, 600, 800, etc. nucleotide pairs in the monomeric and oligomeric nucleosomes, respectively. Acid soluble DNA material is created relatively slowly as compared to the rate of formation of subnucleosomal material. More DNA with lengths in between the 200, 400, etc. nucleotide pairs of nucleosomal DNA is formed when digestions with micrococcal nuclease are carried out at 0 to 10 degrees C compared to 40 degrees C. With DNAase II, on the other hand, formation of a 200 nucleotide pair pattern is favoured at the low temperatures. Apparently, the accessibility of potential cleavage sites in between and within nucleosomes depends strongly on the conditions of digestion. Possible reasons for this dependence are discussed.

Animals↗