scholarly journals Power-law behavior of transcription factor dynamics at the single-molecule level implies a continuum affinity model

2021 ◽  
Author(s):  
David A Garcia ◽  
Gregory Fettweis ◽  
Diego M Presman ◽  
Ville Paakinaho ◽  
Christopher Jarzynski ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Molecule ◽  
Power Law ◽  
Dwell Time ◽  
Specific Binding ◽  
Power Law Distribution ◽  
Single Molecule Level ◽  
Binding Behavior ◽  
Chromatin Template ◽  
Nuclear Domains

Abstract Single-molecule tracking (SMT) allows the study of transcription factor (TF) dynamics in the nucleus, giving important information regarding the diffusion and binding behavior of these proteins in the nuclear environment. Dwell time distributions obtained by SMT for most TFs appear to follow bi-exponential behavior. This has been ascribed to two discrete populations of TFs—one non-specifically bound to chromatin and another specifically bound to target sites, as implied by decades of biochemical studies. However, emerging studies suggest alternate models for dwell-time distributions, indicating the existence of more than two populations of TFs (multi-exponential distribution), or even the absence of discrete states altogether (power-law distribution). Here, we present an analytical pipeline to evaluate which model best explains SMT data. We find that a broad spectrum of TFs (including glucocorticoid receptor, oestrogen receptor, FOXA1, CTCF) follow a power-law distribution of dwell-times, blurring the temporal line between non-specific and specific binding, suggesting that productive binding may involve longer binding events than previously believed. From these observations, we propose a continuum of affinities model to explain TF dynamics, that is consistent with complex interactions of TFs with multiple nuclear domains as well as binding and searching on the chromatin template.

scholarly journals Power-law behaviour of transcription factor dynamics at the single-molecule level implies a continuum affinity model

2019 ◽  
Author(s):  
David A. Garcia ◽  
Gregory Fettweis ◽  
Diego M. Presman ◽  
Ville Paakinaho ◽  
Christopher Jarzynski ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Molecule ◽  
Power Law ◽  
Specific Binding ◽  
Power Law Distribution ◽  
Single Molecule Level ◽  
Complex Interactions ◽  
Chromatin Template ◽  
Nuclear Domains

ABSTRACTSingle-molecule tracking (SMT) allows the study of transcription factor (TF) dynamics in the nucleus, giving important information regarding the search and binding behaviour of these proteins in the nuclear environment. Dwell time distributions for most TFs have been described by SMT to follow bi-exponential behaviour. This is consistent with the existence of two discrete populations bound to chromatin in vivo, one non-specifically bound to chromatin (i.e. searching mode) and another specifically bound to target sites, as originally defined by decades of biochemical studies. However, alternative models have started to emerge, from multiple exponential components to power-law distributions. Here, we present an analytical pipeline with an unbiased model selection approach based on different statistical metrics to determine the model that best explains SMT data. We found that a broad spectrum of TFs (including glucocorticoid receptor, oestrogen receptor, FOXA1, CTCF) follow a power-law distribution, blurring the temporal line between non-specific and specific binding, and suggesting that productive binding may involve longer binding events than previously thought. We propose a continuum of affinities model to explain the experimental data, consistent with the movement of TFs through complex interactions with multiple nuclear domains as well as binding and searching on the chromatin template.

scholarly journals Transcription Factor Target Search Strategy at the Single Molecule Level in Eukaryotic Cells

2012 ◽  
Vol 102 (3) ◽  
pp. 288a
Author(s):  
Lydia Boudarene ◽  
Vincent Récamier ◽  
Davide Normanno ◽  
Ignacio Izzedine ◽  
Ibrahim Cissé ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Molecule ◽  
Search Strategy ◽  
Eukaryotic Cells ◽  
Target Search ◽  
Single Molecule Level ◽  
Transcription Factor Target

scholarly journals Tissue-specific transcription factor target identification in the Caenorhabditis elegans epidermis using targeted DamID

2020 ◽  
Author(s):  
Dimitris Katsanos ◽  
Michalis Barkoulas
Keyword(s):  
Transcription Factor ◽  
Cell Differentiation ◽  
Single Molecule ◽  
Regulatory Networks ◽  
Specific Binding ◽  
Wnt Signalling ◽  
Specific Gene ◽  
List Type ◽  
Tissue Specific ◽  
Transcription Factor Target

SummaryTranscription factors are key orchestrators of development in multicellular animals and display complex patterns of expression, as well as tissue-specific binding to targets. However, our ability to map transcription factor-target interactions in specific tissues of intact animals remains limited. We introduce here targeted DamID (TaDa) as a method to identify transcription factor targets with tissue-specific resolution in C. elegans. We focus on the epidermis as a paradigm and demonstrate that TaDa circumvents problems with Dam-associated toxicity and allows reproducible identification of putative targets. Using a combination of TaDa and single-molecule FISH (smFISH), we refine the positions of LIN-22 and NHR-25 within the epidermal gene network. We reveal direct links between these two factors and the cell differentiation programme, as well as the Wnt signalling pathway. Our results illustrate how TaDa and smFISH can be used to dissect the architecture of tissue-specific gene regulatory networks.HighlightsTaDa circumvents Dam-associated toxicity by keeping levels of Dam expression low.TaDa allows the recovery of tissue-specific methylation profiles representing TF binding.Methylation signal is enriched in regulatory regions of the genome.LIN-22 and NHR-25 targets reveal a link to cell differentiation and Wnt signalling.

scholarly journals Quantifying transcription factor binding dynamics at the single-molecule level in live cells

Methods ◽  
2017 ◽  
Vol 123 ◽  
pp. 76-88 ◽  
Author(s):  
Diego M. Presman ◽  
David A. Ball ◽  
Ville Paakinaho ◽  
Jonathan B. Grimm ◽  
Luke D. Lavis ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Molecule ◽  
Transcription Factor Binding ◽  
Live Cells ◽  
Single Molecule Level ◽  
Factor Binding

scholarly journals Correlative single-molecule fluorescence barcoding of gene regulation in Saccharomyces cerevisiae

2020 ◽  
Author(s):  
Sviatlana Shashkova ◽  
Thomas Nyström ◽  
Mark C Leake ◽  
Adam JM Wollman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Cell Growth ◽  
Single Molecule ◽  
Specific Binding ◽  
Carbon Sources ◽  
Growth Characteristics ◽  
List Type ◽  
Regulatory State ◽  
Single Molecule Fluorescence

AbstractMost cells adapt to their environment by switching combinations of genes on and off through a complex interplay of transcription factor proteins (TFs). The mechanisms by which TFs respond to signals, move into the nucleus and find specific binding sites in target genes is still largely unknown. Single-molecule fluorescence microscopes, which can image single TFs in live cells, have begun to elucidate the problem. Here, we show that different environmental signals, in this case carbon sources, yield a unique single-molecule fluorescence pattern of foci of a key metabolic regulating transcription factor, Mig1, in the nucleus of the budding yeast, Saccharomyces cerevisiae. This pattern serves as a ‘barcode’ of the gene regulatory state of the cells which can be correlated with cell growth characteristics and other biological function.HighlightsSingle-molecule microscopy of transcription factors in live yeastBarcoding single-molecule nuclear fluorescenceCorrelation with cell growth characteristicsGrowth in different carbon sources

scholarly journals Single molecule optical-probes measured power law distribution of polymer dynamics

2016 ◽  
Vol 65 (21) ◽  
pp. 218201
Author(s):  
Li Bin ◽  
Zhang Guo-Feng ◽  
Jing Ming-Yong ◽  
Chen Rui-Yun ◽  
Qin Cheng-Bing ◽  
...  
Keyword(s):  
Single Molecule ◽  
Power Law ◽  
Polymer Dynamics ◽  
Power Law Distribution ◽  
Optical Probes ◽  
Measured Power

Single molecule study of non-specific binding kinetics of LacI in mammalian cells

2015 ◽  
Vol 184 ◽  
pp. 393-400 ◽  
Author(s):  
Laura Caccianini ◽  
Davide Normanno ◽  
Ignacio Izeddin ◽  
Maxime Dahan
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Power Law ◽  
Mammalian Cells ◽  
Specific Binding ◽  
Lac Repressor ◽  
Striking Difference ◽  
Large Variability ◽  
U2os Cells ◽  
Kinetics Of

Many key cellular processes are controlled by the association of DNA-binding proteins (DBPs) to specific sites. The kinetics of the search process leading to the binding of DBPs to their target locus are largely determined by transient interactions with non-cognate DNA. Using single-molecule microscopy, we studied the dynamics and non-specific binding to DNA of the Lac repressor (LacI) in the environment of mammalian nuclei. We measured the distribution of the LacI–DNA binding times at non-cognate sites and determined the mean residence time to be τ1D = 182 ms. This non-specific interaction time, measured in the context of an exogenous system such as that of human U2OS cells, is remarkably different compared to that reported for the LacI in its native environment in E. coli (<5 ms). Such a striking difference (more than 30 fold) suggests that the genome, its organization, and the nuclear environment of mammalian cells play important roles on the dynamics of DBPs and their non-specific DNA interactions. Furthermore, we found that the distribution of off-target binding times follows a power law, similar to what was reported for TetR in U2OS cells. We argue that a possible molecular origin of such a power law distribution of residence times is the large variability of non-cognate sequences found in the mammalian nucleus by the diffusing DBPs.

scholarly journals Toward Sub-Diffraction Imaging of Single-DNA Molecule Sensors Based on Stochastic Switching Localization Microscopy

Sensors ◽  
2020 ◽  
Vol 20 (22) ◽  
pp. 6667
Author(s):  
Seungah Lee ◽  
Indra Batjikh ◽  
Seong Ho Kang
Keyword(s):  
Single Molecule ◽  
Specific Binding ◽  
Super Resolution ◽  
Localization Microscopy ◽  
Single Molecule Level ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Wide Range ◽  
Nanoscale Topography ◽  
Diffraction Imaging ◽  
Optical Reconstruction

The natural characteristics of deoxyribonucleic acid (DNA) enable its advanced applications in nanotechnology as a special tool that can be detected by high-resolution imaging with precise localization. Super-resolution (SR) microscopy enables the examination of nanoscale molecules beyond the diffraction limit. With the development of SR microscopy methods, DNA nanostructures can now be optically assessed. Using the specific binding of fluorophores with their target molecules, advanced single-molecule localization microscopy (SMLM) has been expanded into different fields, allowing wide-range detection at the single-molecule level. This review discusses the recent progress in the SR imaging of DNA nano-objects using SMLM techniques, such as direct stochastic optical reconstruction microscopy, binding-activated localization microscopy, and point accumulation for imaging nanoscale topography. Furthermore, we discuss their advantages and limitations, present applications, and future perspectives.

scholarly journals Probing Transcription Factor Dynamics at the Single-Molecule Level in a Living Cell

Science ◽  
2007 ◽  
Vol 316 (5828) ◽  
pp. 1191-1194 ◽  
Author(s):  
J. Elf ◽  
G.-W. Li ◽  
X. S. Xie
Keyword(s):  
Transcription Factor ◽  
Single Molecule ◽  
Living Cell ◽  
Single Molecule Level
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