Browse
Submit Data
Databases
API
Help

Dataset Information

19 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Rif1 promotes association of G-quadruplex (G4) by its specific G4 binding and oligomerization activities.

ABSTRACT: Rif1 is a conserved protein regulating replication timing and binds preferentially to the vicinity of late-firing/dormant origins in fission yeast. The Rif1 binding sites on the fission yeast genome have an intrinsic potential to generate G-quadruplex (G4) structures to which purified Rif1 preferentially binds. We previously proposed that Rif1 generates chromatin architecture that may determine replication timing by facilitating the chromatin loop formation. Here, we conducted detailed biochemical analyses on Rif1 and its G4 binding. Rif1 prefers sequences containing long stretches of guanines and binds preferentially to the multimeric G4 of parallel or hybrid/mix topology. Rif1 forms oligomers and binds simultaneously to multiple G4. We present a model on how Rif1 may facilitate the formation of chromatin architecture through its G4 binding and oligomerization properties.

SUBMITTER: Masai H

PROVIDER: S-EPMC6565636 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Rif1 promotes association of G-quadruplex (G4) by its specific G4 binding and oligomerization activities.

Masai Hisao H Fukatsu Rino R Kakusho Naoko N Kanoh Yutaka Y Moriyama Kenji K Ma Yue Y Iida Keisuke K Nagasawa Kazuo K

Scientific reports 20190613 1

Rif1 is a conserved protein regulating replication timing and binds preferentially to the vicinity of late-firing/dormant origins in fission yeast. The Rif1 binding sites on the fission yeast genome have an intrinsic potential to generate G-quadruplex (G4) structures to which purified Rif1 preferentially binds. We previously proposed that Rif1 generates chromatin architecture that may determine replication timing by facilitating the chromatin loop formation. Here, we conducted detailed biochemic ...[more]

PMID: 31197198

Similar Datasets

G4 Quadruplex Binding Proteins

Project description:In this study we discover proteins that bind to G4 quadruplex DNA structure. We use modified c-myc quadruplex as a bait, and compare it to the control bait - T15 oligo. We use spectral counts and G-test to determine significant binders. T15 and G4 datafiles are uploaded

2016-07-07 | PXD003501 | Pride

G4-quadruplex-binding proteins: review and insights into selectivity.

Project description:There are over 700,000 putative G4-quadruplexes (G4Qs) in the human genome, found largely in promoter regions, telomeres, and other regions of high regulation. Growing evidence links their presence to functionality in various cellular processes, where cellular proteins interact with them, either stabilizing and/or anchoring upon them, or unwinding them to allow a process to proceed. Interest in understanding and manipulating the plethora of processes regulated by these G4Qs has spawned a new area of small-molecule binder development, with attempts to mimic and block the associated G4-binding protein (G4BP). Despite the growing interest and focus on these G4Qs, there is limited data (in particular, high-resolution structural information), on the nature of these G4Q-G4BP interactions and what makes a G4BP selective to certain G4Qs, if in fact they are at all. This review summarizes the current literature on G4BPs with regards to their interactions with G4Qs, providing groupings for binding mode, drawing conclusions around commonalities and highlighting information on specific interactions where available.

| S-EPMC9250568 | biostudies-literature

Molecular architecture of G-quadruplex structures generated on duplex Rif1-binding sequences.

Project description:G-quadruplexes (G4s) are four-stranded DNA structures comprising stacks of four guanines, are prevalent in genomes, and have diverse biological functions in various chromosomal structures. A conserved protein, Rap1-interacting factor 1 (Rif1) from fission yeast (Schizosaccharomyces pombe), binds to Rif1-binding sequence (Rif1BS) and regulates DNA replication timing. Rif1BS is characterized by the presence of multiple G-tracts, often on both strands, and their unusual spacing. Although previous studies have suggested generation of G4-like structures on duplex Rif1BS, its precise molecular architecture remains unknown. Using gel-shift DNA binding assays and DNA footprinting with various nuclease probes, we show here that both of the Rif1BS strands adopt specific higher-order structures upon heat denaturation. We observed that the structure generated on the G-strand is consistent with a G4 having unusually long loop segments and that the structure on the complementary C-strand does not have an intercalated motif (i-motif). Instead, we found that the formation of the C-strand structure depends on the G4 formation on the G-strand. Thus, the higher-order structure generated at Rif1BS involved both DNA strands, and in some cases, G4s may form on both of these strands. The presence of multiple G-tracts permitted the formation of alternative structures when some G-tracts were mutated or disrupted by deazaguanine replacement, indicating the robust nature of DNA higher-order structures generated at Rif1BS. Our results provide general insights into DNA structures generated at G4-forming sequences on duplex DNA.

| S-EPMC6222110 | biostudies-literature

Membrane Binding Promotes Annexin A2 Oligomerization.

Project description:Annexin A2 (AnxA2) is a cytosolic Ca2+ regulated membrane binding protein that can induce lipid domain formation and plays a role in exocytosis and endocytosis. To better understand the mode of annexin-membrane interaction, we analyzed membrane-bound AnxA2 assemblies by employing a novel 3-armed chemical crosslinker and specific AnxA2 mutant proteins. Our data show that AnxA2 forms crosslinkable oligomers upon binding to membranes containing negatively charged phospholipids. AnxA2 mutants with amino acid substitutions in residues predicted to be involved in lateral protein-protein interaction show compromised oligomer formation, albeit still being capable of binding to negatively charged membranes in the presence of Ca2+. These results suggest that lateral protein-protein interactions are involved in the formation of AnxA2 clusters on a biological membrane.

| S-EPMC7291239 | biostudies-literature

Oligomer formation and G-quadruplex binding by purified murine Rif1 protein, a key organizer of higher-order chromatin architecture.

Project description:Rap1-interacting protein 1 (Rif1) regulates telomere length in budding yeast. We previously reported that, in metazoans and fission yeast, Rif1 also plays pivotal roles in controlling genome-wide DNA replication timing. We proposed that Rif1 may assemble chromatin compartments that contain specific replication-timing domains by promoting chromatin loop formation. Rif1 also is involved in DNA lesion repair, restart after replication fork collapse, anti-apoptosis activities, replicative senescence, and transcriptional regulation. Although multiple physiological functions of Rif1 have been characterized, biochemical and structural information on mammalian Rif1 is limited, mainly because of difficulties in purifying the full-length protein. Here, we expressed and purified the 2418-amino-acid-long, full-length murine Rif1 as well as its partially truncated variants in human 293T cells. Hydrodynamic analyses indicated that Rif1 forms elongated or extended homo-oligomers in solution, consistent with the presence of a HEAT-type helical repeat segment known to adopt an elongated shape. We also observed that the purified murine Rif1 bound G-quadruplex (G4) DNA with high specificity and affinity, as was previously shown for Rif1 from fission yeast. Both the N-terminal (HEAT-repeat) and C-terminal segments were involved in oligomer formation and specifically bound G4 DNA, and the central intrinsically disordered polypeptide segment increased the affinity for G4. Of note, pulldown assays revealed that Rif1 simultaneously binds multiple G4 molecules. Our findings support a model in which Rif1 modulates chromatin loop structures through binding to multiple G4 assemblies and by holding chromatin fibers together.

| S-EPMC5846147 | biostudies-literature

Lipid binding promotes oligomerization and focal adhesion activity of vinculin.

Project description:Adherens junctions (AJs) and focal adhesion (FA) complexes are necessary for cell migration and morphogenesis, and for the development, growth, and survival of all metazoans. Vinculin is an essential regulator of both AJs and FAs, where it provides links to the actin cytoskeleton. Phosphatidylinositol 4,5-bisphosphate (PIP2) affects the functions of many targets, including vinculin. Here we report the crystal structure of vinculin in complex with PIP2, which revealed that PIP2 binding alters vinculin structure to direct higher-order oligomerization and suggests that PIP2 and F-actin binding to vinculin are mutually permissive. Forced expression of PIP2-binding-deficient mutants of vinculin in vinculin-null mouse embryonic fibroblasts revealed that PIP2 binding is necessary for maintaining optimal FAs, for organization of actin stress fibers, and for cell migration and spreading. Finally, photobleaching experiments indicated that PIP2 binding is required for the control of vinculin dynamics and turnover in FAs. Thus, through oligomerization, PIP2 directs a transient vinculin sequestration at FAs that is necessary for proper FA function.

| S-EPMC4259812 | biostudies-literature

Yeast Sub1 and human PC4 are G-quadruplex binding proteins that suppress genome instability at co-transcriptionally formed G4 DNA.

Project description:G-quadruplex or G4 DNA is a non-B secondary DNA structure consisting of a stacked array of guanine-quartets that can disrupt critical cellular functions such as replication and transcription. When sequences that can adopt Non-B structures including G4 DNA are located within actively transcribed genes, the reshaping of DNA topology necessary for transcription process stimulates secondary structure-formation thereby amplifying the potential for genome instability. Using a reporter assay designed to study G4-induced recombination in the context of an actively transcribed locus in Saccharomyces cerevisiae, we tested whether co-transcriptional activator Sub1, recently identified as a G4-binding factor, contributes to genome maintenance at G4-forming sequences. Our data indicate that, upon Sub1-disruption, genome instability linked to co-transcriptionally formed G4 DNA in Top1-deficient cells is significantly augmented and that its highly conserved DNA binding domain or the human homolog PC4 is sufficient to suppress G4-associated genome instability. We also show that Sub1 interacts specifically with co-transcriptionally formed G4 DNA in vivo and that yeast cells become highly sensitivity to G4-stabilizing chemical ligands by the loss of Sub1. Finally, we demonstrate the physical and genetic interaction of Sub1 with the G4-resolving helicase Pif1, suggesting a possible mechanism by which Sub1 suppresses instability at G4 DNA.

| S-EPMC5449603 | biostudies-literature

Custom G4 Microarrays Reveal Selective G-Quadruplex Recognition of Small Molecule BMVC: A Large-Scale Assessment of Ligand Binding Selectivity.

Project description:G-quadruplexes (G4) are considered new drug targets for human diseases such as cancer. More than 10,000 G4s have been discovered in human chromatin, posing challenges for assessing the selectivity of a G4-interactive ligand. 3,6-bis(1-Methyl-4-vinylpyridinium) carbazole diiodide (BMVC) is the first fluorescent small molecule for G4 detection in vivo. Our previous structural study shows that BMVC binds to the MYC promoter G4 (MycG4) with high specificity. Here, we utilize high-throughput, large-scale custom DNA G4 microarrays to analyze the G4-binding selectivity of BMVC. BMVC preferentially binds to the parallel MycG4 and selectively recognizes flanking sequences of parallel G4s, especially the 3'-flanking thymine. Importantly, the microarray results are confirmed by orthogonal NMR and fluorescence binding analyses. Our study demonstrates the potential of custom G4 microarrays as a platform to broadly and unbiasedly assess the binding selectivity of G4-interactive ligands, and to help understand the properties that govern molecular recognition.

| S-EPMC7436161 | biostudies-literature

Characterization of G-quadruplex antibody reveals differential specificity for G4 DNA forms.

Project description:Accumulating evidence suggests that human genome can fold into non-B DNA structures, when appropriate sequence and favourable conditions are present. Among these, G-quadruplexes (G4-DNA) are associated with gene regulation, chromosome fragility and telomere maintenance. Although several techniques are used in detecting such structures in vitro, understanding their intracellular existence has been challenging. Recently, an antibody, BG4, was described to study G4 structures within cells. Here, we characterize BG4 for its affinity towards G4-DNA, using several biochemical and biophysical tools. BG4 bound to G-rich DNA derived from multiple genes that form G-quadruplexes, unlike complementary C-rich or random sequences. BLI studies revealed robust binding affinity (Kd = 17.4 nM). Gel shift assays show BG4 binds to inter- and intramolecular G4-DNA, when it is in parallel orientation. Mere presence of G4-motif in duplex DNA is insufficient for antibody recognition. Importantly, BG4 can bind to G4-DNA within telomere sequence in a supercoiled plasmid. Finally, we show that BG4 binds to form efficient foci in four cell lines, irrespective of their lineage, demonstrating presence of G4-DNA in genome. Importantly, number of BG4 foci within the cells can be modulated, upon knockdown of G4-resolvase, WRN. Thus, we establish specificity of BG4 towards G4-DNA and discuss its potential applications.

| S-EPMC7711166 | biostudies-literature

Expression data by G-quadruplex (G4) forming oligonucleotides transfection in 3D culture.

Project description:Telomere erosion causes cell mortality, suggesting that longer telomeres allow greater number of cell division. In telomerase-positive human cancer cells, however, telomeres are often kept shorter than the surrounding normal tissues. Recently, we have shown that telomere elongation in cancer cells represses innate immune genes and promotes their differentiation in vivo. This implies that short telomeres contribute to cancer malignancy, but it is unclear how such genetic repression is caused by long telomeres. Here we report that telomeric repeat-containing RNA (TERRA) induces genome-wide alteration of gene expression in telomere-elongated cancer cells in vivo. Using three different cell lines, we found that G4 forming oligonucleotide repressed innate immune genes in vivo 3D culture conditions. Most of the suppressed genes belonged to innate immune system categories and were upregulated in various cancers. We propose that TERRA G4 counteracts cancer malignancy through suppression of innate immune genes. Six samples are G4 oligo-transfected cells (PC-3/(uuaggg)^4, PC-3/AS1411, HBC4/(uuaggg)^4, HBC4/AS1411, MKN74/(uuaggg)^4 and MKN74/AS1411), and the other six samples are control oligo-transfected cells.

2015-02-05 | E-GEOD-56239 | biostudies-arrayexpress

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data