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ホーム > Séminaires > Archives séminaires > Séminaires 2013 > Séminaire MSC Lundi 7 Octobre à 11h30. Maxime Dahan (Physico-Chimie Curie). "Probing the target search of individual DNA-binding proteins in mammalian cells".

Séminaire MSC Lundi 7 Octobre à 11h30. Maxime Dahan (Physico-Chimie Curie). "Probing the target search of individual DNA-binding proteins in mammalian cells"

Sauf mention contraire, les séminaires et les soutenances se déroulent à 11h30 en salle 454A du bâtiment Condorcet.

Maxime Dahan Physico Chimie Curie, Paris

Lundi 7 Octobre à 11h30.

Probing the target search of individual DNA-binding proteins in mammalian cells

D. Normanno1,2,3, L. Boudarene1,2, J. Chen3, C. Dugast-Darzacq2, X. Darzacq2,3, and M. Dahan1,3

1 Laboratoire Physico-Chimie, Institut Curie, CNRS UMR168, Paris, France. 2 Functional imaging of transcription, IBENS, Paris, France 3 Transcription Imaging Consortium, HHMI Janelia Farm, Ashburn, Virginia.

For many cellular functions, DNA-binding proteins (DBPs) need to find specific target sites in the genome. Facilitated diffusion (FD), namely the combination of one-dimensional motion along non-specific DNA and three-dimensional exploration, is the dominant model for the target search (TS) of DBPs. Yet, this model has hardly been tested in vivo, particularly in the complex environment of a mammalian nucleus, and it is still controversial whether it accelerates association to specific DNA binding sites. To address that question, we have implemented a TS assay using human cells with a unique target locus for an inducible exogenous searcher, the tetracycline repressor (TetR). Using single-molecule tracking and in situ biochemical measurements of association kinetics, we directly characterize the mobility of TetR, its transient interaction with non-cognate DNA and the kinetics of binding to the specific locus. Overall, we find that the searcher follows a FD strategy but it does not accelerate its search compared to a 3D diffusion-limited process, due to low association efficiency at the target site. I will also present on-going efforts to develop 3D imaging tools for single-molecule tracking and super-resolution. In particular, I will discuss the performances of a novel multifocus microscope that allows volumetric imaging and will be instrumental for future studies of the functional dynamics of nuclear factors.

Related publications : 1. Ibrahim Cisse, Ignacio Izeddin, Sebastien Causse, Lydia Boudarene, Adrien Senecal, Leila Muresan, Claire Dugast-Darzacq, Bassam Hajj, Maxime Dahan, Xavier Darzacq, « Real time dynamics of RNA Polymerase II clustering in live human cells », Science (2013) 341, 664-7. 2. M. El Beheiry and M. Dahan, “ViSP : representing single particle localization in three dimensions”, Nature Methods (2013) 10, 689–690. 3. S. Abrahamsson, J. Chen, B. Hajj, S. Stallinga, A. Katsov, J. Wisniewski, G. Mizuguchi, P. Soulle, F. Mueller, C. Dugast Darzacq, X. Darzacq, C.Wu, C. I. Bargmann, D. A. Agard, M. Dahan and M.G.L. Gustafsson, “Fast multi-color 3D imaging using aberration corrected multi-focus microscopy”, Nature Methods (2013) 10(1):60-3. 4. D. Normanno, M. Dahan, and X. Darzacq, “Intra-nuclear mobility and target search mechanisms of transcription factors : a single-molecule perspective on gene expression and regulation”, Biophysica and Bichemica Acta (2012), 1819, 482–493

Contact : Équipe séminaires / Seminar team - Published on / Publié le 23 septembre 2013

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