PubMed Health⌕ Search

Biomedical subjects

T de Lange

Publications and source records attributed to T de Lange.

51 records · Page 3Linked to original sources
PubMed1995

A human telomeric protein.

Telomeres are multifunctional elements that shield chromosome ends from degradation and end-to-end fusions, prevent activation of DNA damage checkpoints, and modulate the maintenance of telomeric DNA by telomerase. A major protein component of human telomeres has been identified and cloned. This factor, TRF, contains one Myb-type DNA-binding repeat and an amino-terminal acidic domain. Immunofluorescent labeling shows that TRF specifically colocalizes with telomeric DNA in human interphase cells and is located at chromosome ends during metaphase. The presence of TRF along the telomeric TTAGGG repeat array demonstrates that human telomeres form a specialized nucleoprotein complex.

Amino Acid Sequence↗
PubMed1995

Telomerase activity in normal and malignant hematopoietic cells.

Bone marrow and peripheral blood leukocytes from 19 leukemia patients were found to contain telomerase activity detectable by a PCR-based assay. Telomerase was also detectable in nonmalignant bone marrow and peripheral blood leukocytes from normal donors, including fractions enriched for granulocytes, T lymphocytes, and monocytes/B cells. Semiquantitative comparison revealed considerable overlap between telomerase activities in samples from normal subjects and leukemia patients, confounding evaluation of the role of telomerase in this disease. These data indicate that human telomerase is not restricted to immortal cells and suggest that the somatic expression of this enzyme may be more widespread than was previously inferred from the decline of human telomeres.

Acute Disease↗
PubMed1994

Stringent sequence requirements for the formation of human telomeres.

In human cells, transfection of telomeric T2AG3 repeats induces the formation of functional telomeres at previously interstitial sites. We report that telomere formation has stringent sequence requirements. While (T2AG3)n telomere seeds formed telomeres in approximately 70% of the transfected cells, five T2AG3-related heterologous telomeric DNAs seeded new telomeres in < 5% of the transfectants. Telomere formation did not correlate with the ability of human telomerase to elongate telomeric sequences in vitro. Homologous recombination is probably also not involved because a (T2AG3)n telomere seed with nontelomeric DNA at 160-bp intervals formed new telomeres frequently. Instead, the sequence dependence of telomere formation matched the in vitro binding requirements for the mammalian T2AG3 repeat binding factor (TRF). Human TRF failed to bind ineffective heterologous telomere seeds and had a 4-fold lower affinity for (T2AG5)2T2AG3 repeats, which seeded telomeres with reduced frequency. The results suggest that telomere seeds interact with TRF and predict that mammalian artificial chromosomes will require wild-type telomeric repeats at, or near, their termini.

Base Sequence↗
PubMed1994

Unusual chromatin in human telomeres.

We report that human telomeres have an unusual chromatin structure characterized by diffuse micrococcal nuclease patterns. The altered chromatin manifested itself only in human telomeres that are relatively short (2 to 7 kb). In contrast, human and mouse telomeres with telomeric repeat arrays of 14 to 150 kb displayed a more canonical chromatin structure with extensive arrays of tightly packed nucleosomes. All telomeric nucleosomes showed a shorter repeat size than bulk nucleosomes, and telomeric mononucleosomal particles were found to be hypersensitive to micrococcal nuclease. However, telomeric nucleosomes were similar to bulk nucleosomes in the rate at which they sedimented through sucrose gradients. We speculate that mammalian telomeres have a bipartite structure with unusual chromatin near the telomere terminus and a more canonical nucleosomal organization in the proximal part of the telomere.

Animals↗
PubMed1993

Telomeric structure in cells with chromosome end associations.

End-to-end associations of metaphase chromosomes have been observed in a variety of human tumors, ageing cells, and several chromosome instability syndromes. Since telomeres of tumor cells and ageing tissues are often reduced in length, it has been suggested that chromosome end associations may be due to loss of telomeric repeats. We report the molecular structure of telomeres of two human tumor cell lines with frequent end-to-end associations of metaphase chromosomes. These telomeres were shown to be severely reduced compared with most other human cells with functional telomeres. However, we also describe two cell lines with severely shortened telomeres that are not detectably compromised in their function. We suggest that telomeric length is not the only determinant of the fusigenic behavior of human telomeres in tumor cells.

Base Sequence↗
PubMed1993

A Xenopus egg factor with DNA-binding properties characteristic of terminus-specific telomeric proteins.

We have identified a Xenopus laevis protein factor that specifically recognizes vertebrate telomeric repeats at DNA ends. This factor, called Xenopus telomere end factor (XTEF), is detected predominantly in extracts of Xenopus eggs and ovaries, which are estimated to contain sufficient XTEF to bind approximately 3 x 10(7) DNA ends. In contrast, XTEF is much less abundant (approximately 90 per cell) in extracts of somatic cell nuclei. Mobility retardation analysis of the XTEF activity in egg extracts indicates that this factor binds the vertebrate telomeric repeat sequence (TTAGGG)2 when present in a single-stranded 3' overhang. Single-stranded 3' extensions of (TTTGGG)2, (AAAGGG)2, (TTACCC)2, or a nonrepetitive sequence fail to bind XTEF efficiently, whereas changes in the double-stranded sequence 5' to the TTAGGG repeat tail are tolerated. TTAGGG repeats are not recognized at internal position, at a 5' protruding end, or in double-stranded DNA. In addition, the factor does not bind RNA with single-stranded UUAGGG repeats at a 3' end. XTEF-DNA complexes form and are stable in high salt. The DNA-binding properties of XTEF resemble the characteristics of a class of terminus-specific telomere proteins identified previously in hypotrichous ciliates.

Acetates↗
PubMed1992

Human telomeres are attached to the nuclear matrix.

This report shows that human telomeres are tightly associated with the nuclear matrix. Telomere attachment is observed in several cell types and in all stages of interphase. Mapping experiments show that telomeres are anchored via their TTAGGG repeats; a subtelomeric repeat located immediately proximal to the telomeric TTAGGG repeats is quantitatively released from the nuclear matrix by restriction endonuclease cleavage. TTAGGG repeats introduced at chromosome-internal sites by DNA transfection do not behave as matrix attached loci, suggesting that the telomeric position of the repeats is required for their interaction with the nuclear matrix. These findings are consistent with the idea that telomeres function as a nucleoprotein complex.

Base Sequence↗
PubMed1992

A mammalian factor that binds telomeric TTAGGG repeats in vitro.

We have identified a DNA-binding activity with specificity for the TTAGGG repeat arrays found at mammalian telomeres. This factor, called TTAGGG repeat factor (TRF), is present in nuclear extracts of human, mouse, and monkey cells. TRF from HeLa cells was characterized in detail by electrophoretic mobility shift assays. It binds double-stranded TTAGGG repeats in linear and circular DNAs. Single-stranded repeats are not recognized. The optimal site for TRF appears to contain more than six contiguous TTAGGG repeats. Tandem arrays of TAGGG, TTTAGGG, TTTTAGGG, TTGGGG, and TTAGGC repeats do not bind TRF well, indicating that TRF preferentially recognizes the telomeric repeat sequence present at mammalian chromosome ends. The apparent molecular mass of this factor, based on recovery of TRF from sodium dodecyl sulfate-polyacrylamide gels, is approximately 50 kDa. We suggest that TRF binds along the length of mammalian telomeres.

Animals↗
PubMed1991

A map of the distal region of the long arm of human chromosome 21 constructed by radiation hybrid mapping and pulsed-field gel electrophoresis.

We have used radiation hybrid (RH) mapping and pulsed-field gel electrophoresis (PFGE) to determine the order and positions of 28 DNA markers from the distal region of the long arm of human chromosome 21. The maps generated by these two methods are in good agreement. This study, combined with that of D. R. Cox et al. (1990, Science 250:245-250), results in an RH map that covers the long arm of chromosome 21 (21q). We have used a subtelomeric probe to show that our map includes the telomere and have identified single-copy genes and markers within 200 kbp of the telomere. Comparison of the physical and RH maps with genetic linkage maps shows "hot spots" of meiotic recombination in the distal region, one of which is close to the telomere, in agreement with previous cytogenetic observations of increased recombination frequency near telomeres.

Cell Line↗
PubMed1990

Structure and variability of human chromosome ends.

Mammalian telomeres are thought to be composed of a tandem array of TTAGGG repeats. To further define the type and arrangement of sequences at the ends of human chromosomes, we developed a direct cloning strategy for telomere-associated DNA. The method involves a telomere enrichment procedure based on the relative lack of restriction endonuclease cutting sites near the ends of human chromosomes. Nineteen (TTAGGG)n-bearing plasmids were isolated, two of which contain additional human sequences proximal to the telomeric repeats. These telomere-flanking sequences detect BAL 31-sensitive loci and thus are located close to chromosome ends. One of the flanking regions is part of a subtelomeric repeat that is present at 10 to 25% of the chromosome ends in the human genome. This sequence is not conserved in rodent DNA and therefore should be a helpful tool for physical characterization of human chromosomes in human-rodent hybrid cell lines; some of the chromosomes that may be analyzed in this manner have been identified, i.e., 7, 16, 17, and 21. The minimal size of the subtelomeric repeat is 4 kilobases (kb); it shows a high frequency of restriction fragment length polymorphisms and undergoes extensive de novo methylation in somatic cells. Distal to the subtelomeric repeat, the chromosomes terminate in a long region (up to 14 kb) that may be entirely composed of TTAGGG repeats. This terminal segment is unusually variable. Although sperm telomeres are 10 to 14 kb long, telomeres in somatic cells are several kilobase pairs shorter and very heterogeneous in length. Additional telomere reduction occurs in primary tumors, indicating that somatic telomeres are unstable and may continuously lose sequences from their termini.

Animals↗
PubMed1987

Coincident multiple activations of the same surface antigen gene in Trypanosoma brucei.

Trypanosomes with a coat of variant surface glycoprotein (VSG) 118, consistently appear around day 20 when a rabbit is infected with Trypanosoma brucei strain 427. There is a single chromosome-internal gene for VSG 118 and this is activated by duplicative transposition to a telomeric expression site. We show here that the expression-linked extra copy of VSG gene 118 in a day 18 population of a chronic infection is heterogeneous, and we infer that the population is not monoclonal but is the result of multiple independent activations of the 118 gene. We show that the heterogeneity of expression-linked extra copies is also present in other trypanosome populations expressing chromosome-internal VSG genes. We present a model for the timing of VSG gene activation during chronic infection that emphasizes two features: the relative activation and inactivation frequencies of different expression sites, and the degree of homology of the sequences flanking VSG genes with expression sites.

Antigens, Protozoan↗
PubMed1987

Definition of regions in human c-myc that are involved in transformation and nuclear localization.

To study the relationship between the primary structure of the c-myc protein and some of its functional properties, we made in-frame insertion and deletion mutants of the normal human c-myc coding domain that was expressed from a retroviral promoter-enhancer. We assessed the effects of these mutations on the ability of c-myc protein to cotransform normal rat embryo cells with a mutant ras gene, induce foci in a Rat-1-derived cell line (Rat-1a), and localize in nuclei. Using the cotransformation assay, we found two regions of the protein (amino acids 105 to 143 and 321 to 439) where integrity was critical: one region (amino acids 1 to 104) that tolerated insertion and small deletion mutations, but not large deletions, and another region (amino acids 144) to 320) that was largely dispensable. Comparison with regions that were important for transformation of Rat-1a cells revealed that some are essential for both activities, but others are important for only one or the other, suggesting that the two assays require different properties of the c-myc protein. Deletion of each of three regions of the c-myc protein (amino acids 106 to 143, 320 to 368, and 370 to 412) resulted in partial cytoplasmic localization, as determined by immunofluorescence or immunoprecipitation following subcellular fractionation. Some abnormally located proteins retained transforming activity; most proteins lacking transforming activity appeared to be normally located.

Animals↗
PubMed1979

Energy requirement for maintenance of the transmembrane potassium gradient in Klebsiella aerogenes NCTC 418: a continuous culture study.

With a glucose-limited culture of Klebsiella aerogenes, growing at a fixed dilution rate (0.4 h-1), the specific respiration rate varied progressively as a function of the transmembrane K+ gradient. The latter was varied by changing the input K+ concentration and, under these conditions, the specific respiration rate was linearly related to the electrochemical potential of the K+ gradient. Increasing or decreasing the transmembrane K+ gradient in putatively potassium-limited cultures elicited marked changes in respiration rate consistent with the conclusion that the exceptionally high respiration rates expressed by fully glucose-sufficient potassium-limited cultures (i.e., values in excess of 25 mmol O2/g dry weight organisms . h, at D = 0.4 h-1) are necessary to scavenge traces of K+ from the environment and hence maintain an exceptionally high transmembrane K+ gradient.

Cell Membrane Permeability↗