Protein:DNA interactions at chromosomal loop attachment sites

Genome ◽  
1989 ◽  
Vol 31 (2) ◽  
pp. 503-509 ◽  
Author(s):  
Veronica C. Blasquez ◽  
Ann O. Sperry ◽  
Peter N. Cockerill ◽  
William T. Garrard
Keyword(s):  
Nuclear Matrix ◽  
Binding Sites ◽  
Topoisomerase Ii ◽  
Regulatory Sequences ◽  
Immunoglobulin Gene ◽  
Sequence Motifs ◽  
Cis Acting ◽  
Consensus Sequences ◽  
Dna And Rna ◽  
Multiple Binding Sites

We have recently identified an evolutionarily conserved class of sequences that organize chromosomal loops in the interphase nucleus, which we have termed "matrix association regions" (MARs). MARs are about 200 bp long, AT-rich, contain topoisomerase II consensus sequences and other AT-rich sequence motifs, often reside near cis-acting regulatory sequences, and their binding sites are abundant (> 10 000 per mammalian nucleus). Here we demonstrate that the interactions between the mouse κ immunoglobulin gene MAR and topoisomerase II or the "nuclear matrix" occur between multiple and sometimes overlapping binding sites. Interestingly, the sites most susceptible to topoisomerase II cleavage are localized near the breakpoints of a previously described illegitimate recombination event. The presence of multiple binding sites within single MARs may allow DNA and RNA polymerase passage without disrupting primary loop organization.Key words: MARs, chromatin loops, topoisomerase II, nuclear matrix.

scholarly journals Identification of regulatory sequences and protein-binding sites in the liver-type promoter of a gene encoding 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

1991 ◽  
Vol 11 (2) ◽  
pp. 1099-1106 ◽  
Author(s):  
F P Lemaigre ◽  
S M Durviaux ◽  
G G Rousseau
Keyword(s):  
Protein Binding ◽  
Binding Sites ◽  
Specific Protein ◽  
Regulatory Sequences ◽  
Alternative Promoters ◽  
Gene Encoding ◽  
Protein Binding Sites ◽  
Cis Acting ◽  
Dnase I Footprinting ◽  
Nuclear Extracts

The liver-type and muscle-type isozymes of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase are encoded by one gene that uses two alternative promoters. We have identified cis-acting sequences and protein-binding sites on the liver-type promoter. Transfection assays with deleted promoters showed that maximal promoter activity is contained within 360 bp upstream of the cap site. DNase I footprinting experiments with liver and spleen nuclear extracts and with purified proteins revealed several protein-binding sites in this region. These included four binding sites for nuclear factor I, one site that contains an octamer consensus but showed a liver-specific footprint pattern, two liver-specific protein-binding sites, and one poly(dG)-containing binding site. Transfection of cells of hepatic origin suggested that all these sites except one are involved in transcriptional regulation. The region between -360 and -2663 contained an element that functioned as a silencer in a nonhepatic cell line. We conclude that in liver transcription from the liver-type promoter of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene is controlled by ubiquitous and tissue-specific factors and involves activating and derepressing mechanisms.

Role of AP1/NFE2 binding sites in endogenous α-globin gene transcription

Blood ◽  
2003 ◽  
Vol 102 (12) ◽  
pp. 4223-4228 ◽  
Author(s):  
Melanie R. Loyd ◽  
Yasuhiro Okamoto ◽  
Mindy S. Randall ◽  
Paul A. Ney
Keyword(s):  
Gene Transcription ◽  
Binding Sites ◽  
Regulatory Element ◽  
Globin Gene ◽  
Gene Activation ◽  
Loxp Site ◽  
Regulatory Sequences ◽  
Cis Acting ◽  
High Level

Abstract High-level α-globin expression depends on cis-acting regulatory sequences located far upstream of the α-globin cluster. Sequences that contain the α-globin positive regulatory element (PRE) activate α-globin expression in transgenic mice. The α-globin PRE contains a pair of composite binding sites for the transcription factors activating protein 1 and nuclear factor erythroid 2 (AP1/NFE2). To determine the role of these binding sites in α-globin gene transcription, we mutated the AP1/NFE2 sites in the α-globin PRE in mice. We replaced the AP1/NFE2 sites with a neomycin resistance gene (neo) that is flanked by LoxP sites (floxed). Mice with this mutation exhibited increased embryonic death and α-thalassemia intermedia. Next, we removed the neo gene by Cre-mediated recombination, leaving a single LoxP site in place of the AP1/NFE2 sites. These mice were phenotypically normal. However, α-globin expression, measured by allele-specific RNA polymerase chain reaction (PCR), was decreased 25%. We examined the role of the hematopoietic-restricted transcription factor p45Nfe2 in activating expression through these sites and found that it is not required. Thus, we have demonstrated that AP1/NFE2 binding sites in the murine α-globin PRE contribute to long-range α-globin gene activation. The proteins that mediate this effect remain to be determined. (Blood. 2003;102:4223-4228)

scholarly journals Cloning and characterization of the human osteopontin gene and its promoter

1994 ◽  
Vol 303 (1) ◽  
pp. 255-262 ◽  
Author(s):  
N Hijiya ◽  
M Setoguchi ◽  
K Matsuura ◽  
Y Higuchi ◽  
S Akizuki ◽  
...  
Keyword(s):  
Human Monocyte ◽  
Mouse Gene ◽  
Upstream Region ◽  
Regulatory Sequences ◽  
Major Band ◽  
Cis Acting ◽  
Consensus Sequences ◽  
Shift Analysis ◽  
Intron Structure ◽  
The Difference

We isolated the human osteopontin (hOP) gene and the 5′ upstream region, and analysed its exon-intron structure and potential regulatory sequences of the promoter region in comparison with those of the mouse and porcine gene. The coding sequence is split into 7 exons which are similar to those of the mouse gene, although the hOP gene is longer than the mouse gene. The difference in length is mainly due to variations in intron 3, which is approximately 2.7-fold longer than that of the mouse OP gene. The 5′ upstream region of the hOP, which is highly conserved up to nucleotide -250, contains a number of potential cis regulatory consensus sequences. A series of sequentially 5′-deleted chimeric clones was tested for the ability to stimulate chloramphenicol acetyltransferase (CAT). Initial CAT analysis demonstrated that nucleotides at positions -474 to -270, -124 to -80, and -55 to -39 contained cis-acting enhancing sequences in a human monocyte cell line, SCC-3, although the -124 to -80 region was much more active than other regions. Deletion of the sequences between -474 and -270 localized this cis region to the sequence at positions -439 to -410, whereas the deletion between -124 to -80 localized the regions to -124 to -115, and -94 to -80. Gel-shift analysis using as probes synthesized double-stranded DNA corresponding to the 10 and 15 bp region at positions -124 to -115 and -94 to -80 respectively revealed that each probe formed a major band complexed with nuclear proteins prepared from SCC-3 cells.

Identification of regulatory sequences and protein-binding sites in the liver-type promoter of a gene encoding 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase

1991 ◽  
Vol 11 (2) ◽  
pp. 1099-1106
Author(s):  
F P Lemaigre ◽  
S M Durviaux ◽  
G G Rousseau
Keyword(s):  
Protein Binding ◽  
Binding Sites ◽  
Specific Protein ◽  
Regulatory Sequences ◽  
Alternative Promoters ◽  
Gene Encoding ◽  
Protein Binding Sites ◽  
Cis Acting ◽  
Dnase I Footprinting ◽  
Nuclear Extracts

The liver-type and muscle-type isozymes of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase are encoded by one gene that uses two alternative promoters. We have identified cis-acting sequences and protein-binding sites on the liver-type promoter. Transfection assays with deleted promoters showed that maximal promoter activity is contained within 360 bp upstream of the cap site. DNase I footprinting experiments with liver and spleen nuclear extracts and with purified proteins revealed several protein-binding sites in this region. These included four binding sites for nuclear factor I, one site that contains an octamer consensus but showed a liver-specific footprint pattern, two liver-specific protein-binding sites, and one poly(dG)-containing binding site. Transfection of cells of hepatic origin suggested that all these sites except one are involved in transcriptional regulation. The region between -360 and -2663 contained an element that functioned as a silencer in a nonhepatic cell line. We conclude that in liver transcription from the liver-type promoter of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene is controlled by ubiquitous and tissue-specific factors and involves activating and derepressing mechanisms.

NMDAR PAMs: Multiple Chemotypes for Multiple Binding Sites

2019 ◽  
Vol 19 (24) ◽  
pp. 2239-2253 ◽  
Author(s):  
Paul J. Goldsmith
Keyword(s):  
Binding Sites ◽  
Receptor Activation ◽  
Research Area ◽  
Allosteric Modulators ◽  
Nonselective Cation Channels ◽  
Multiple Binding Sites ◽  
Multiple Binding ◽  
Excitatory Neurotransmission ◽  
Psychiatric Conditions ◽  
High Level

The N-methyl-D-aspartate receptor (NMDAR) is a member of the ionotropic glutamate receptor (iGluR) family that plays a crucial role in brain signalling and development. NMDARs are nonselective cation channels that are involved with the propagation of excitatory neurotransmission signals with important effects on synaptic plasticity. NMDARs are functionally and structurally complex receptors, they exist as a family of subtypes each with its own unique pharmacological properties. Their implication in a variety of neurological and psychiatric conditions means they have been a focus of research for many decades. Disruption of NMDAR-related signalling is known to adversely affect higherorder cognitive functions (e.g. learning and memory) and the search for molecules that can recover (or even enhance) receptor output is a current strategy for CNS drug discovery. A number of positive allosteric modulators (PAMs) that specifically attempt to overcome NMDAR hypofunction have been discovered. They include various chemotypes that have been found to bind to several different binding sites within the receptor. The heterogeneity of chemotype, binding site and NMDAR subtype provide a broad landscape of ongoing opportunities to uncover new features of NMDAR pharmacology. Research on NMDARs continues to provide novel mechanistic insights into receptor activation and this review will provide a high-level overview of the research area and discuss the various chemical classes of PAMs discovered so far.

Effects of Multiple Binding Sites on Studies of Hydrogen Bonding between Nitroxide Radicals and Solvent Molecules

1993 ◽  
Vol 58 (1) ◽  
pp. 47-52 ◽  
Author(s):  
Imad Al-Bala'a ◽  
Richard D. Bates
Keyword(s):  
Hydrogen Bonding ◽  
Magnetic Resonance ◽  
Binding Site ◽  
Binding Sites ◽  
Hyperfine Coupling Constant ◽  
Hydrogen Donor ◽  
Complex Formation Constants ◽  
Multiple Binding Sites ◽  
Multiple Binding ◽  
Solvent Molecules

The role of more than one binding site on a nitroxide free radical in magnetic resonance determinations of the properties of the complex formed with a hydrogen donor is examined. The expression that relates observed hyperfine couplings in EPR spectra to complex formation constants and concentrations of each species in solution becomes much more complex when multiple binding sites are present, but reduces to a simpler form when binding at the two sites occurs independently and the binding at the non-nitroxide site does not produce significant differences in the hyperfine coupling constant in the complexed radical. Effects on studies of hydrogen bonding between multiple binding site nitroxides and hydrogen donor solvent molecules by other magnetic resonance methods are potentially more extreme.

Sign in / Sign up

Export Citation Format

Share Document