SEARCH · PubMed Health

Results for “Locus Control Region”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources

PubMed1997

Molecular analysis of the Turkish form of deletion-inversion (delta beta)(0) thalassaemia.

Two unrelated (delta beta)(0)-thalassaemia patients from Southern Turkey are presented. DNA studies indicated that both of them are homozygous for the Turkish type of (delta beta)(0)-thalassaemia characterized by one large deletion of 11.5 kb including the delta and beta globin genes at the 5' end and one small deletion of 1.6 kb at the 3' end, which are separated by an inverted 7.6 kb long DNA segment that includes L1 repetitive sequence. In the present study a PCR-based method was performed to produce a unique deletion-specific product and subjected to sequence analysis for the determination of the breakpoint. DNA polymorphisms in the beta-globin gene cluster of deletion-inversion type of (delta beta)(0)-thalassaemia, IVS-I-6 and beta-39 globin genes were examined. Analysis of sequence variations in regulatory regions including the 5' hypersensitive site-2 of the locus control region (LCR), the delta, (G)gamma and (A)gamma 5' flanking regions and the second intervening sequence (IVS-II) of (A)gamma and (G)gamma genes indicated the presence of close similarities between the chromosome carrying the Turkish form of deletion-inversion (delta beta)(0)-thalassaemia and the chromosome associated with beta-39 nonsense mutation in haplotype II. These two chromosomes are characterized by the presence of a 4 base pair deletion in the (A)gamma(T) globin gene promoter. A C --> T alteration at position -199 5' to the delta gene was also found to be associated with the Turkish type of (delta beta)(0)-thalassaemia and beta-39 chromosome.

Adult↗

PubMed1993

Far upstream regions of class II MHC Ea are necessary for position-independent, copy-dependent expression of Ea transgene.

The chromatin upstream of the class II MHC Ea gene contains specific, DNase I hypersensitive (DH) sites (groups I-V), overlapping and extending the promoter proximal and distal control regions. To determine whether the Ea DH groups I-V define a functionally important chromatin domain or locus control region (LCR), we have used wild type Ead gene constructs to generate transgenic mouse lines from strains that do not express an endogenous Ea gene product. Constructs contained either DH groups I-V 'Longs' or DH groups I-II 'Shorts', of the hypersensitive sites defined within 20 kb 5' of Ea. We show that position-independent, copy number-dependent expression of the Ead gene occurs only with the Long construct (8/8 transgenic mouse lines, over a range of copy numbers, 1-30 copies); in contrast, the Short constructs are subject to position-dependent effects. This suggests that the region delineated by Ea DH groups I-II is necessary but not sufficient as an LCR, which requires the presence of the upstream regions containing DH III-V for complete position-independent, copy number-dependent expression. These results introduce an immunologically-important, putative LCR which can be used to target genes to cells of the B cell lineage, as well as to other class II MHC expressing cells, and highlight the importance of chromatin structure analysis as a means to locate DNA regions of regulatory interest which are dispersed over a large distance.

Animals↗

PubMed1993

Use of long sequence alignments to study the evolution and regulation of mammalian globin gene clusters.

The determination of long segments of DNA sequences encompassing the beta- and alpha-globin gene clusters has provided an unprecedented data base for analysis of genome evolution and regulation of gene clusters. A newly developed computer tool kit generates local alignments between such long sequences in a space-efficient manner, helps the user analyze the alignments effectively, and finds consistently aligning blocks of sequences in multiple pairwise comparisons. Such sequence analyses among the beta-like globin gene clusters of human, galago, rabbit, and mouse have revealed the general patterns of evolution of this gene cluster. Alignments in the flanking regions are very useful in assigning orthologous relationships. Investigation of such matches between the mouse and human beta-like globin gene clusters has led to a reassessment of some orthologous assignments in mouse and to a revision of the proposed pathway for evolution of this gene cluster. In general, the interspersed repetitive elements have inserted independently, presumably via a retrotransposition mechanism, in the different mammalian lineages. However, some examples of ancient L1 repeats are found, including one between the epsilon- and gamma-globin genes that appears to have been in the ancestral eutherian gene cluster. Prominent matching sequences are found in a long region 5' to the epsilon-globin gene, the locus control region (LCR) that is a positive regulator of the entire gene cluster. Three-way alignments among the human, goat, and rabbit sequences can extend for > or = 3 kb in part of the LCR (DNase hypersensitive site 3), indicating that the cis-acting components of this complex regulatory region cover a long segment of DNA. In contrast to the beta-like globin gene clusters, the alpha-like globin gene clusters of many mammals occur in very G+C-rich isochores and contain prominent CpG islands. The regions between the alpha-like globin genes are evolving faster than the intergenic regions of the beta-like globin gene clusters. The contrasts between the two gene clusters can be attributed to differences in DNA metabolism in the isochore. The proximal control elements of the rabbit alpha-globin gene are located both 5' to and within the gene. All of this region is part of a prominent CpG island that may be acting as an extended, enhancer-independent promoter. One can hypothesize that the analogue to the LCR in the alpha-globin gene cluster may interface with the distinctive alpha-globin promoter in ways different from the interaction between the beta LCR and the promoters of beta-like globin genes.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1993

A 5' control region of the human epsilon-globin gene is sufficient for embryonic specificity in transgenic mice.

When introduced as part of DNA constructions containing the human beta-globin locus control region (LCR), the human embryonic beta-globin gene, epsilon, is expressed in primitive but not definitive erythroid cells of recipient transgenic mice. In contrast to this pattern, the human fetal beta-globin gene, gamma, has been shown to be expressed in both primitive and definitive erythroid cells of transgenic mice when introduced in similar LCR-containing constructions. To begin to identify the minimal sequence(s) necessary for the epsilon expression pattern, we have fused a DNA fragment that contains the human epsilon-globin gene promoter region, and 13.7-kilobase (kb) of contiguous upstream flanking sequence containing super-hypersensitive (HS) sites 5'HS-2 and 5'HS-1 of the globin LCR, to the structural portion and near 3'-flanking region of the human gamma-globin gene. This construction, and one containing an intact human gamma-globin gene with the same 3'-flanking sequence and 383 base pairs of 5'-flanking sequence linked to LCR DNA from -0.86 to -13.7 kb upstream of epsilon, were each microinjected to produce transgenic mice. While the construction containing the intact gamma-globin gene is transcriptionally active in primitive and definitive erythroid cells of the transgenic mice, the fusion construction, in which the gamma-globin gene promoter and promoter proximal region is essentially replaced by that of epsilon, is not active in definitive erythroid cells and expresses with the same pattern as an intact epsilon gene. These results indicate that the promoter and near 5'-flanking region of epsilon, when linked to the LCR, is sufficient for embryonic-specific expression in transgenic mice. The level of expression of the fusion construction in primitive erythroid cells of transgenic mice is similar to that previously observed for the intact epsilon gene when identically cloned. This suggests that the epsilon 5'-region of the fusion construction also contains all the sequence necessary for the LCR-dependent activation of epsilon in transgenic mice.

Animals↗

PubMed1997

Dyad symmetry within the mouse 3' IgH regulatory region includes two virtually identical enhancers (C alpha3'E and hs3).

The transcription of the murine Ig heavy chain locus is regulated not only by the intronic enhancer, E mu, but also by a 3' regulatory region located downstream of the C alpha membrane exon. Several DNase I-hypersensitive sites (hs1-4) and enhancer elements (e.g., C alpha3'E) have been identified in this 3' regulatory region, and some of these were suggested to comprise a locus control region. However, little is known about the coordinate regulation or function of these individual elements. Here we provide evidence that C alpha3'E and hs3 are virtually mirror images of each other and demarcate the edges of an approximately 25-kb region of quasi-dyad symmetry with 3'alphaE(hs1,2) at its center. Flanking 3'alphaE(hs1,2) are inverted repeats and families of repetitive sequences uniquely located in this region. We have observed that, like 3'alphaE(hs1,2) and hs3, C alpha3'E is DNase I hypersensitive in plasma cell lines, but not in a pre-B cell line. Additionally, we found that C alpha3'E and hs3 show significant transcriptional synergy in transfection assays only in a plasma cell line. The DNA topology of the 3' regulatory region coupled with new and existing data on the activity of its individual enhancers during B cell differentiation lead us to propose a biphasic model for the activity of this region. According to our model, one unit, consisting of the 3'-most enhancer, hs4, is active early and throughout B cell development. The second unit, which comprises C alpha3'E, 3'alphaE(hs1,2), and hs3, becomes active later in development, when it contributes to such processes as class switching and increased levels of Ig heavy chain gene transcription in plasma cells.

Animals↗

PubMed1994

HLA class I chromosomal region, genes, and products: facts and questions.

Among the various areas of recent investigation in the field of human MHC class I antigens, the following have been selected for discussion in this review: (1) classical HLA class I genes: are they ubiquitously expressed?, what are the special features of their polymorphism?, are HLA-C molecules functional?, (2) non-classical HLA class I gene products: how restricted is their tissue distribution?, do they exhibit a little polymorphism?, what is their function, if any? (3) non-HLA genes recently detected in the HLA class I chromosomal region: are some of them involved in immunological function and development?, (4) other novel coding sequences present, or possibly present, in the region: the hemochromatosis gene, grc region and associated tumor suppressor genes, housekeeping genes, human equivalent of the murine H-2M region and Ped gene; (5) transcriptional regulation: are there cis-regulatory elements, including locus control region(s), located elsewhere than in the promoters? are CpG methylation, gene imprinting, chromatin structure, DNA rearrangement also implicated? what are the transcription factors involved and how do they interact with each other? is there HLA class I locus-, allele-, or isoform-specific regulation? is class I gene expression dysregulated in human tumors? The answers to these questions are crucial for the development of the future directions for research.

Base Sequence↗

PubMed1995

Evidence for distinct DNA binding forms of the erythroid-specific transcription factor NF-E2.

The transcriptional activity of the beta-globin genes is regulated by a complex genetic element, the locus control region (LCR), at the 5'-end of the beta-globin locus. Tandem binding sites for the erythroid-specific transcription factor NF-E2 are important for the transcriptional activation function of the LCR. We discovered that vanadate strongly stimulates the DNA binding activity of NF-E2 in crude and fractionated nuclear extracts. The other oxyanions, molybdate and tungstate, do not affect NF-E2 DNA binding. Quantitative DNA binding experiments indicated that vanadate stimulates NF-E2 DNA binding by increasing the number of NF-E2 molecules that are competent to bind to DNA, rather than influencing the affinity of binding. Gel filtration analysis revealed a similar Stokes' radius for NF-E2, in the absence or presence of vanadate, inconsistent with a role for vanadate in stabilizing the heteromeric NF-E2 complex. Distinct NF-E2 forms, which were either weakly or strongly induced by vanadate, were resolved by cation and anion exchange chromatography. A model is proposed in which two conformers of NF-E2 share an identical subunit composition, but differ in DNA binding activity. Vanadate may interact directly with one of the conformers to generate the high-affinity DNA binding state. The presence of a non-DNA binding pool of NF-E2 suggests that the formation of an active NF-E2 heteromer may be a regulated step in the cell.

Base Sequence↗

PubMed1992

Human gamma- to beta-globin gene switching using a mini construct in transgenic mice.

The developmental regulation of the human globin genes involves a key switch from fetal (gamma-) to adult (beta-) globin gene expression. It is possible to study the mechanism of this switch by expressing the human globin genes in transgenic mice. Previous work has shown that high-level expression of the human globin genes in transgenic mice requires the presence of the locus control region (LCR) upstream of the genes in the beta-globin locus. High-level, correct developmental regulation of beta-globin gene expression in transgenic mice has previously been accomplished only in 30- to 40-kb genomic constructs containing the LCR and multiple genes from the locus. This suggests that either competition for LCR sequences by other globin genes or the presence of intergenic sequences from the beta-globin locus is required to silence the beta-globin gene in embryonic life. The results presented here clearly show that the presence of the gamma-globin gene (3.3 kb) alone is sufficient to down-regulate the beta-globin gene in embryonic transgenic mice made with an LCR-gamma-beta-globin mini construct. The results also show that the gamma-globin gene is down-regulated in adult mice from most transgenic lines made with LCR-gamma-globin constructs not including the beta-globin gene, i.e., that the gamma-globin gene can be autonomously regulated. Evidence presented here suggests that a region 3' of the gamma-globin gene may be important for down-regulation in the adult. The 5'HS2 gamma en beta construct described is a suitable model for further study of the mechanism of human gamma- to beta-globin gene switching in transgenic mice.

Aging↗

PubMedDate not supplied

The intricacies of beta-globin gene expression.

Gene expression is an extremely complicated process in which several mechanisms are involved. Owing to its developmental and tissue-specific expression, the beta-globin gene is an excellent model for studying gene expression. beta-Globin gene expression involves an interplay between several different mechanisms. Chromatin structure is thought to be altered by the locus control region (LCR) located far upstream of the beta-globin gene locus. As well, multiple transcription factors come into play both in the LCR and in the individual promoters and enhancers of the beta-globin genes. The interaction between these then allows for delicate regulation of beta-globin gene expression. In the following review the elaborate system of beta-globin gene expression will briefly be examined.

Beta-Globulins↗

PubMed1993

Distal regulatory elements from the mouse metallothionein locus stimulate gene expression in transgenic mice.

DNA regions of 10 and 7 kb that flank the mouse metallothionein II (MT-II) and MT-I genes, respectively, were combined with a minimally marked MT-I (MT-I*) gene and tested in transgenic mice. This construct resulted in (i) position-independent expression of MT-I* mRNA and copy number-dependent expression, (ii) levels of hepatic MT-I mRNA per cell per transgene that were about half that derived from endogenous MT-I genes, (iii) appropriate regulation by metals and hormones, and (iv) tissue distribution of transgene mRNA that resembled that of endogenous MT-I mRNA. These features were not observed when MT-I* was tested without the flanking regions. These MT-I flanking sequences also improved the expression of rat growth hormone reporter genes, with or without introns, that were under the control of the MT-I promoter. Moreover, they enhanced expression from two of four heterologous promoters/enhancers that were tested. Deletion analysis indicated that regions known to have DNase I-hypersensitive sites were necessary but not sufficient for high-level expression. These data suggest that the DNA regions flanking the mouse MT-I and MT-II genes have functions like the locus control regions described for other genes.

Animals↗

PubMed1991

Characterization of the major regulatory element upstream of the human alpha-globin gene cluster.

The major positive regulatory activity of the human alpha-globin gene complex has been localized to an element associated with a strong erythroid-specific DNase I hypersensitive site (HS -40) located 40 kb upstream of the zeta 2-globin mRNA cap site. Footprint and gel shift analyses of the element have demonstrated the presence of four binding sites for the nuclear factor GATA-1 and two sites corresponding to the AP-1 consensus binding sequence. This region resembles one of the major elements of the beta-globin locus control region in its constitution and characteristics; this together with evidence from expression studies suggests that HS -40 is a primary element controlling alpha-globin gene expression.

Base Sequence↗

PubMed1997

Concomitant downregulation of IgH 3' enhancer activity and c-myc expression in a plasmacytoma x fibroblast environment: implications for dysregulation of translocated c-myc.

Regulation of immunoglobulin heavy chain (IgH) gene expression is controlled by a B cell-specific promoter, intronic enhancer and additional B cell-specific enhancer elements identified recently in the 3' end of the IgH locus. One of the latter elements, the IgH 3' enhancer, is of particular interest: (1) it is B cell-specific and active only in late B cell development; (2) in rodent plasmacytomas and in some human Burkitt's lymphomas it is part of a locus control region (LCR) that is involved in deregulation of the c-myc oncogene as a result of translocation into the IgH locus; and (3) it has been implicated in the mechanisms that control Ig gene class switch recombination. We have used a somatic cell hybridization approach to genetically analyse regulation of the activity of the IgH 3' enhancer. When mouse MPC11 plasmacytoma cells, in which the IgH 3' enhancer is active, are fused with fibroblasts, Ig expression is extinguished at the level of transcription. Here we show that in a MPC11 plasmacytoma x fibroblast environment, the IgH 3' enhancer is transcriptionally inactive. Furthermore, we demonstrate that binding of several B cell-specific transcription factors, essential for IgH 3' enhancer activity, is lacking, which may explain 3' enhancer inactivity, although the binding of repressors cannot be excluded. Moreover, the high expression level of c-myc, characteristic of the parental MPC11 cells carrying the t(12;15) translocation, is down-regulated in the hybrids to that in unfused fibroblasts. Therefore, inactivation of the IgH 3' enhancer is a multifactorial process affecting several transcription factors that control the cell-specific and developmental activity of the enhancer.

Animals↗

PubMed1994

Possible factors influencing the haemoglobin and fetal haemoglobin levels in patients with beta-thalassaemia due to a homozygosity for the IVS-I-6 (T-->C) mutation.

We have collected haematological, haemoglobin (Hb) and DNA sequence data for 29 patients with a homozygosity for the IVS-I-6 (T-->C) mutation with the intention of identifying factors contributing to the observed variability in the severity of the disease. None of the patients had received blood transfusion therapy for at least 6 months prior to the study. Hb levels varied from 5.0 to 9.9 g/dl. Patients with high Hb F (more than 1.5 g/dl or > 20%) had high total Hb levels (7.5-9.7 g/dl) but some with low Hb F also had high total Hb levels; two had a concomitant alpha-thalassaemia-2 (alpha-thal-2) heterozygosity. An inverse correlation between the Hb F and Hb A2 levels was observed. The majority of the patients were homozygous for haplotype VI (49/58 chromosomes) but haplotypes IV (2/58) and VII (7/58) were also present. The only haplotype IV homozygote had high Hb F levels with high G gamma values and the C-->T mutation at position -158 in the G gamma promoter, while both high and low Hb F levels were observed among patients with haplotypes VI and VII. Analysis of sequence variations in regulatory regions included the 5' hypersensitive sites (HS) 4. 3 and 2 of the locus control region (LCR), the G gamma and A gamma 5' flanking regions, the second intervening sequence (IVS-II), and the 5' beta-globin gene region in two patients with high Hb F (one homozygote each for haplotypes VI and IV), and in two patients with low Hb F levels (one homozygote each for haplotypes VI and VII). Haplotype specific differences were observed in the LCR 5' HS-2 and in the G gamma and A gamma flanking and IVS-II regions; however, no differences were present between the low and high Hb F-producing haplotype VI chromosomes, suggesting a major role for factors which are not linked to the beta-globin gene cluster in mediating gamma-globin gene expression in patients with this type of beta-thal.

Adolescent↗

PubMed1993

Germ-line transmission and developmental regulation of a 150-kb yeast artificial chromosome containing the human beta-globin locus in transgenic mice.

Sequential expression of the genes of the human beta-globin locus requires the formation of an erythroid-specific chromatin domain spanning > 200 kb. Regulation of this gene family involves both local interactions with proximal cis-acting sequences and long-range interactions with control elements upstream of the locus. To make it possible to analyze the interactions of cis-acting sequences of the human beta-globin locus in their normal spatial and sequence context, we characterized two yeast artificial chromosomes (YACs) 150 and 230 kb in size, containing the entire beta-globin locus. We have now successfully integrated the 150-kb YAC into the germ line of transgenic mice as a single unrearranged fragment that includes the locus control region, structural genes, and 30 kb of 3' flanking sequences present in the native locus. Expression of the transgenic human beta-globin locus is tissue- and developmental stage-specific and closely follows the pattern of expression of the endogenous mouse beta-globin locus. By using homology-directed recombination in yeast and methods for the purification and transfer of YACs into transgenic mice, it will now be feasible to study the physiological role of cis-acting sequences in specifying an erythroid-specific chromatin domain and directing expression of beta-globin genes during ontogeny.

Age Factors↗

PubMed1992

The developmental regulation of the human zeta-globin gene in transgenic mice employing beta-galactosidase as a reporter gene.

We have investigated the developmental and tissue specific expression of the human embryonic zeta-globin gene in transgenic mice. A construct containing 550 bp of zeta-globin 5' flanking region, fused to a beta-galactosidase (lacZ) reporter gene and linked to the locus control region (LCR)-like alpha positive regulatory element (alpha PRE) was employed for the production of transgenic mice. Firstly, we compared the number of live born transgenic mice containing this construct to the number of live born transgenic mice containing the entire zeta-globin gene linked to the alpha PRE or the beta LCR. Data showed that 12% of mice generated from eggs injected with zeta-promoter/lacZ/alpha PRE DNA were transgenic compared to only 2% of mice generated from eggs injected with the entire zeta-globin gene linked to the alpha PRE or the beta LCR. The reduced number of live born transgenic mice containing the latter constructs suggests that death of transgenic embryos, possibly due to thalassaemia, may be occurring. X-gal staining of whole embryos containing the lacZ gene revealed that zeta-globin promoter activity was most pronounced at 8.5-9.5 days of development and was restricted to erythroid cells. By 15 days of development, no zeta-globin promoter activity was detected. These results suggest that the alpha PRE can direct high level expression from the zeta-globin promoter and that sequences required for the correct tissue and developmental specific expression of the human zeta-globin gene are present within 550 bp's of 5' flanking region. Sequences within the body of the zeta-globin gene or 3' of the cap site do not appear to be necessary for correct zeta-globin developmental regulation.

Animals↗

PubMed1997

Properties of the mouse alpha-globin HS-26: relationship to HS-40, the major enhancer of human alpha-globin gene expression.

HS-26, the mouse homologue of HS-40, is the major regulatory element of the mouse alpha-globin gene locus. Like HS-40, HS-26 is located within an intron of a house-keeping gene; comparison of the nucleotide sequences of HS-26 and HS-40 reveals conservation of the sequences and positions of several DNA binding motifs in the 5' regions of both elements (3 GATA, 2 NFE-2, and 1 CACCC sites) and the absence in HS-26 of three CACCC sites and one GATA site that are present in the 3' region of HS-40, suggesting that the two elements might not be identical. We report here that when HS-26 is linked to a 1.5 kb Pstl human alpha-globin gene fragment, it has a weak enhancer activity in induced MEL cells and in transgenic embryos, and it does not have any detectable activity in adult transgenic mice. This suggests that HS-26 does not have Locus Control Region (LCR) activity but can act as an enhancer during the embryonic life when integrated at a permissive locus. To further test the importance of HS-26 at its natural locus, we have generated embryonic stem cells and chimeric animals in which 350 bp containing HS-26 have been replaced by a neomycin resistance gene by homologous recombination. The sizes of the chimeras' red cells were then estimated by measuring forward scattering on a FacsScan apparatus in hypotonic conditions. This revealed that a fraction of the chimeric animals' red cells were smaller than normal mouse red cells and were very similar to cells from mice heterozygous for alpha-thalassemia. Density gradient analysis also suggested the presence of thalassemic cells. These results indicated that despite its lack of LCR activity, HS-26 is important for the regulation of the mouse alpha-globin gene locus.

Animals↗

PubMed1996

Heterochromatin effects on the frequency and duration of LCR-mediated gene transcription.

Locus control regions (LCRs) are responsible for initiating and maintaining a stable tissue-specific open chromatin structure of a locus. In transgenic mice, LCRs confer high level expression on linked genes independent of position in the mouse genome. Here we show that an incomplete LCR loses this property when integrated into heterochromatic regions. Two disruption mechanisms were observed. One is classical position-effect variegation, resulting in continuous transcription in a clonal subpopulation of cells. The other is a novel mechanism resulting in intermittent gene transcription in all cells. We conclude that only a complete LCR fully overcomes heterochromatin silencing and that it controls the level of transcription by ensuring activity in all cells at all times rather than directly controlling the rate of transcription.

Animals↗

PubMed1995

Isolation of a differentially regulated splicing isoform of human NF-E2.

The transcription factor NF-E2 (nuclear factor erythroid 2), interacting via DNA motifs within regulatory regions of several hematopoietic genes, is thought to mediate the enhancer activity of the globin locus control regions. By screening a human fetal liver cDNA library with probes derived from mouse NF-E2, we have isolated a splicing variant of the NF-E2 gene (fNF-E2) that differs in the 5' untranslated region from the previously reported cDNA (aNF-E2). The fNF-E2 isoform is transcribed from an alternative promoter located in the 3' end of the first intron and joined by alternative splicing to the second and third exons, which are shared by both RNA isoforms. Although the two forms produce the same protein, they are expressed in different ratios during development. fNF-E2 is more abundant in the fetal liver and less abundant in the adult bone marrow compared to the previously described form. Their distribution apparently follows the differential expression of fetal and adult hemoglobins.

Alternative Splicing↗