scholarly journals A Study of p53 Action on DNA at the Single Molecule Level

2021 ◽  
Author(s):  
Kiyoto Kamagata
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Dynamic Properties ◽  
Diffusion Mechanism ◽  
Target Search ◽  
Single Molecule Level ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Biophysical Study ◽  
Model Protein

The transcription factor p53 searches for and binds to target sequences within long genomic DNA, to regulate downstream gene expression. p53 possesses multiple disordered and DNA-binding domains, which are frequently observed in DNA-binding proteins. Owing to these properties, p53 is used as a model protein for target search studies. It counters cell stress by utilizing a facilitated diffusion mechanism that combines 3D diffusion in solution, 1D sliding along DNA, hopping/jumping along DNA, and intersegmental transfer between two DNAs. Single-molecule fluorescence microscopy has been used to characterize individual motions of p53 in detail. In addition, a biophysical study has revealed that p53 forms liquid-like droplets involving the functional switch. In this chapter, the target search and regulation of p53 are discussed in terms of dynamic properties.

scholarly journals Single-molecule analysis reveals two separate DNA-binding domains in the Escherichia coli UvrA dimer

2009 ◽  
Vol 37 (6) ◽  
pp. 1962-1972 ◽  
Author(s):  
Koen Wagner ◽  
Geri Moolenaar ◽  
John van Noort ◽  
Nora Goosen
Keyword(s):  
Escherichia Coli ◽  
Dna Binding ◽  
Single Molecule ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Single Molecule Analysis

scholarly journals Slow, heterogeneous NFκB single molecule conformational dynamics and implications for function

2021 ◽  
Author(s):  
Wei Chen ◽  
Wei Liu ◽  
Peter Wolynes ◽  
Elizabeth A. Komives
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Electrostatic Interactions ◽  
Bound States ◽  
Conformational Dynamics ◽  
Dna Binding Domains ◽  
Single Molecule Fret ◽  
Binding Domains ◽  
Lower Amplitude ◽  
Slow Motions

The transcription factor NFκB (RelA-p50) is a multidomain protein that binds DNA and its inhibitor, IκBα with apparently different conformations. We used single-molecule FRET to characterize the interdomain motions of the N-terminal DNA-binding domains in the free protein and also in various bound states. Several surprising results emerged from this study. First, the domains moved with respect to each other on several widely different timescales from hundreds of milliseconds to minutes. The free NFκB displayed stochastic motions leading to a broad distribution of states, ranging from very low-FRET states to high-FRET states. Varying the ionic strength altered the slow motions suggesting that they may be due to different weak electrostatic interactions between the domains creating a rugged energy landscape. Third, the DNA-binding domains continued to be mobile even when the protein was bound to its cognate DNA, but in this case the majority of the states were either high-FRET, a state expected from the available x-ray structures, or low-FRET, a state consistent with one of the DNA-binding domains dissociated. The fluctuations of the DNA-bound states were of lower amplitude and slightly faster frequency. Fourth, the inhibitor, IκBα freezes the domains into a low-FRET state, expected to be incapable of binding DNA. Neutralization of five acidic residues in the IκBα PEST sequence, which was previously shown to impair IκBαs ability to strip NFκB from the DNA, also impaired its ability to freeze the domains into a low-FRET state indicating that the freezing of motions of the DNA-binding domains is essential for efficient molecular stripping.

scholarly journals Pioneering, chromatin remodeling, and epigenetic constraint in early T-cell gene regulation by PU.1

2018 ◽  
Author(s):  
Jonas Ungerbäck ◽  
Hiroyuki Hosokawa ◽  
Xun Wang ◽  
Tobias Strid ◽  
Brian A. Williams ◽  
...  
Keyword(s):  
T Cell ◽  
Dna Binding ◽  
Target Genes ◽  
Dynamic Properties ◽  
Chromatin Accessibility ◽  
Site Quality ◽  
Open Chromatin ◽  
Loss Of Function ◽  
Dna Binding Domains ◽  
Binding Domains

AbstractPU.1 is a dominant but transient regulator in early T-cell precursors and a potent transcriptional controller of developmentally important pro-T cell genes. Before T-lineage commitment, open chromatin is frequently occupied by PU.1, and many PU.1 sites lose accessibility when PU.1 is later downregulated. Pioneering activity of PU.1 was tested in in this developmentally dynamic context, by quantitating the relationships between PU.1 occupancy and site quality and accessibility as PU.1 levels naturally declined in pro-T cell development, and by using stage-specific gain and loss of function perturbations to relate binding to effects on target genes. PU.1 could bind closed genomic sites, but rapidly opened many of them, despite the absence of its frequent collaborators, C/EBP factors. The dynamic properties of PU.1 engagements implied that PU.1 binding affinity and concentration determine its occupancy choices, but with quantitative tradeoffs for occupancy between site sequence quality and stage-dependent site accessibility in chromatin. At non-promoter sites PU.1 binding criteria were more stringent than at promoters, and PU.1 was also much more effective as a transcriptional regulator at non-promoter sites where local chromatin accessibility depended on the presence of PU.1. Runx motifs and Runx1 binding were often linked to PU.1 at open sites, but highly expressed PU.1 could bind its sites without Runx1. Notably, closed chromatin presented a qualitative barrier to occupancy by the PU.1 DNA binding domain alone. Thus, effective pioneering at closed chromatin sites also depends on requirements beyond site recognition served by non-DNA binding domains of PU.1.

scholarly journals Mechanistic Heterogeneity in Site Recognition by the Structurally Homologous DNA-binding Domains of the ETS Family Transcription Factors Ets-1 and PU.1

2014 ◽  
Vol 289 (31) ◽  
pp. 21605-21616 ◽  
Author(s):  
Shuo Wang ◽  
Miles H. Linde ◽  
Manoj Munde ◽  
Victor D. Carvalho ◽  
W. David Wilson ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Ets Family ◽  
Site Recognition

scholarly journals Protein Kinase A Signaling Pathway Regulates Transcriptional Activity of SAF-1 by Unmasking Its DNA-binding Domains

2003 ◽  
Vol 278 (25) ◽  
pp. 22586-22595 ◽  
Author(s):  
Alpana Ray ◽  
Papiya Ray ◽  
Nicole Guthrie ◽  
Arvind Shakya ◽  
Deepak Kumar ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Dna Binding ◽  
Protein Kinase A ◽  
Signaling Pathway ◽  
Transcriptional Activity ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Kinase A

scholarly journals Role of Papillomavirus E1 Initiator Dimerization in DNA Replication

2005 ◽  
Vol 79 (13) ◽  
pp. 8661-8664 ◽  
Author(s):  
Stephen Schuck ◽  
Arne Stenlund
Keyword(s):  
Dna Replication ◽  
Dna Binding ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Severe Effect ◽  
Origin Of Dna Replication ◽  
Initiation Of Dna Replication

ABSTRACT Viral initiator proteins are polypeptides that form oligomeric complexes on the origin of DNA replication (ori). These complexes carry out a multitude of functions related to initiation of DNA replication, and although many of these functions have been characterized biochemically, little is understood about how the complexes are assembled. Here we demonstrate that loss of one particular interaction, the dimerization between E1 DNA binding domains, has a severe effect on DNA replication in vivo but has surprisingly modest effects on most individual biochemical activities in vitro. We conclude that the dimer interaction is primarily required for initial recognition of ori.

scholarly journals Cdc13 N-Terminal Dimerization, DNA Binding, and Telomere Length Regulation

2010 ◽  
Vol 30 (22) ◽  
pp. 5325-5334 ◽  
Author(s):  
Meghan T. Mitchell ◽  
Jasmine S. Smith ◽  
Mark Mason ◽  
Sandy Harper ◽  
David W. Speicher ◽  
...  
Keyword(s):  
Dna Binding ◽  
Telomere Length ◽  
Functional Characterization ◽  
Point Mutations ◽  
Telomeric Dna ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Length Regulation ◽  
Telomere Length Regulation

ABSTRACT The essential yeast protein Cdc13 facilitates chromosome end replication by recruiting telomerase to telomeres, and together with its interacting partners Stn1 and Ten1, it protects chromosome ends from nucleolytic attack, thus contributing to genome integrity. Although Cdc13 has been studied extensively, the precise role of its N-terminal domain (Cdc13N) in telomere length regulation remains unclear. Here we present a structural, biochemical, and functional characterization of Cdc13N. The structure reveals that this domain comprises an oligonucleotide/oligosaccharide binding (OB) fold and is involved in Cdc13 dimerization. Biochemical data show that Cdc13N weakly binds long, single-stranded, telomeric DNA in a fashion that is directly dependent on domain oligomerization. When introduced into full-length Cdc13 in vivo, point mutations that prevented Cdc13N dimerization or DNA binding caused telomere shortening or lengthening, respectively. The multiple DNA binding domains and dimeric nature of Cdc13 offer unique insights into how it coordinates the recruitment and regulation of telomerase access to the telomeres.

Conformational changes in the DNA-binding domains of the ecdysteroid receptor during the formation of a complex with the hsp27 response element

2012 ◽  
Vol 30 (4) ◽  
pp. 379-393 ◽  
Author(s):  
Szymon Pakuła ◽  
Marek Orłowski ◽  
Grzegorz Rymarczyk ◽  
Tomasz Krusiński ◽  
Michał Jakób ◽  
...  
Keyword(s):  
Dna Binding ◽  
Conformational Changes ◽  
Response Element ◽  
Ecdysteroid Receptor ◽  
Dna Binding Domains ◽  
Binding Domains

scholarly journals Differential Gene Regulation by Selective Association of Transcriptional Coactivators and bZIP DNA-Binding Domains

2006 ◽  
Vol 26 (16) ◽  
pp. 5969-5982 ◽  
Author(s):  
Benoit Miotto ◽  
Kevin Struhl
Keyword(s):  
Dna Binding ◽  
Regulation Of Gene Expression ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Transcriptional Coactivators ◽  
Differential Gene Regulation ◽  
Arginine Residues ◽  
Contact Dna ◽  
Bzip Proteins ◽  
Basic Region

ABSTRACT bZIP DNA-binding domains are targets for viral and cellular proteins that function as transcriptional coactivators. Here, we show that MBF1 and the related Chameau and HBO1 histone acetylases interact with distinct subgroups of bZIP proteins, whereas pX does not discriminate. Selectivity of Chameau and MBF1 for bZIP proteins is mediated by residues in the basic region that lie on the opposite surface from residues that contact DNA. Chameau functions as a specific coactivator for the AP-1 class of bZIP proteins via two arginine residues. A conserved glutamic acid/glutamine in the linker region underlies MBF1 specificity for a subgroup of bZIP factors. Chameau and MBF1 cannot synergistically coactivate transcription due to competitive interactions with the basic region, but either protein can synergistically coactivate with pX. Analysis of Jun derivatives that selectively interact with these coactivators reveals that MBF1 is crucial for the response to oxidative stress, whereas Chameau is important for the response to chemical and osmotic stress. Thus, the bZIP domain mediates selective interactions with coactivators and hence differential regulation of gene expression.

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