@article {48343,
	title = {Quantifying transcription factor binding dynamics at the single-molecule level in live cells.},
	journal = {Methods (San Diego, Calif.)},
	year = {2017},
	month = {2017 Mar 15},
	abstract = {<p>Progressive, technological achievements in the quantitative fluorescence microscopy field are allowing researches from many different areas to start unraveling the dynamic intricacies of biological processes inside living cells. From super-resolution microscopy techniques to tracking of individual proteins, fluorescence microscopy is changing our perspective on how the cell works. Fortunately, a growing number of research groups are exploring single-molecule studies in living cells. However, no clear consensus exists on several key aspects of the technique such as image acquisition conditions, or analysis of the obtained data. Here, we describe a detailed approach to perform single-molecule tracking (SMT) of transcription factors in living cells to obtain key binding characteristics, namely their residence time and bound fractions. We discuss different types of fluorophores, labeling density, microscope, cameras, data acquisition, and data analysis. Using the glucocorticoid receptor as a model transcription factor, we compared alternate tags (GFP, mEOS, HaloTag, SNAP-tag, CLIP-tag) for potential multicolor applications. We also examine different methods to extract the dissociation rates and compare them with simulated data. Finally, we discuss several challenges that this exciting technique still faces.</p>
},
	issn = {1095-9130},
	doi = {10.1016/j.ymeth.2017.03.014},
	author = {Presman, Diego M and Ball, David A and Paakinaho, Ville and Grimm, Jonathan B and Lavis, Luke D and Karpova, Tatiana S and Hager, Gordon L}
}
@article {47420,
	title = {Steroid receptors reprogram FoxA1 occupancy through dynamic chromatin transitions.},
	journal = {Cell},
	year = {2016},
	month = {2016 Apr 6},
	abstract = {<p>The estrogen receptor (ER), glucocorticoid receptor (GR), and forkhead box protein 1 (FoxA1) are significant factors in breast cancer progression. FoxA1 has been implicated in establishing ER-binding patterns though its unique ability to serve as a pioneer factor. However, the molecular interplay between ER, GR, and FoxA1 requires further investigation. Here we show that ER and GR both have the ability to alter the genomic distribution of the FoxA1 pioneer factor. Single-molecule tracking experiments in live cells reveal a highly dynamic interaction of FoxA1 with chromatin in\&nbsp;vivo. Furthermore, the FoxA1 factor is not associated with detectable footprints at its binding sites throughout the genome. These findings support a model wherein interactions between transcription factors and pioneer factors are highly dynamic. Moreover, at a subset of genomic sites, the role of pioneer can be reversed, with the steroid receptors serving to enhance binding of FoxA1.</p>
},
	issn = {1097-4172},
	doi = {10.1016/j.cell.2016.02.067},
	author = {Swinstead, Erin E and Miranda, Tina B and Paakinaho, Ville and Baek, Songjoon and Goldstein, Ido and Hawkins, Mary and Karpova, Tatiana S and Ball, David and Mazza, Davide and Lavis, Luke D and Grimm, Jonathan B and Morisaki, Tatsuya and Gr{\o}ntved, Lars and Presman, Diego M and Hager, Gordon L}
}
