article · 01/07/2015
Probing the target search of DNA-binding proteins in mammalian cells using TetR as model searcher
Résumé
Many cellular functions rely on DNA-binding proteins finding and associating to specific sites in the genome. Yet the mechanisms underlying the target search remain poorly understood, especially in the case of the highly organized mammalian cell nucleus. Using as a model Tet repressors (TetRs) searching for a multi-array locus, we quantitatively analyse the search process in human cells with single-molecule tracking and single-cell protein-DNA association measurements. We find that TetRs explore the nucleus and reach their target by 3D diffusion interspersed with transient interactions with non-cognate sites, consistent with the facilitated diffusion model. Remarkably, nonspecific binding times are broadly distributed, underlining a lack of clear delimitation between specific and nonspecific interactions. However, the search kinetics is not determined by diffusive transport but by the low association rate to nonspecific sites. Altogether, our results provide a comprehensive view of the recruitment dynamics of proteins at specific loci in mammalian cells.
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Normanno, D., Boudarene, L., Dugast-Darzacq, C., Chen, J., Richter, C., Proux, F., Benichou, O., Voituriez, R., Darzacq, X., & Dahan, M. (2015). Probing the target search of DNA-binding proteins in mammalian cells using TetR as model searcher. NATURE COMMUNICATIONS, 6(7357). https://doi.org/10.1038/ncomms8357@article{Normanno2015_196,
author = {Normanno, Davide and Boudarene, Lydia and Dugast-Darzacq, Claire and Chen, Jiji and Richter, Christian and Proux, Florence and Benichou, Olivier and Voituriez, Raphael and Darzacq, Xavier and Dahan, Maxime},
year = {2015},
month = {7},
title = {Probing the target search of DNA-binding proteins in mammalian cells using TetR as model searcher},
journal = {NATURE COMMUNICATIONS},
volume = {6},
number = {7357},
abstract = {Many cellular functions rely on DNA-binding proteins finding and associating to specific sites in the genome. Yet the mechanisms underlying the target search remain poorly understood, especially in the case of the highly organized mammalian cell nucleus. Using as a model Tet repressors (TetRs) searching for a multi-array locus, we quantitatively analyse the search process in human cells with single-molecule tracking and single-cell protein-DNA association measurements. We find that TetRs explore the nucleus and reach their target by 3D diffusion interspersed with transient interactions with non-cognate sites, consistent with the facilitated diffusion model. Remarkably, nonspecific binding times are broadly distributed, underlining a lack of clear delimitation between specific and nonspecific interactions. However, the search kinetics is not determined by diffusive transport but by the low association rate to nonspecific sites. Altogether, our results provide a comprehensive view of the recruitment dynamics of proteins at specific loci in mammalian cells.},
url = {http://www.dx.doi.org/10.1038/ncomms8357},
doi = {10.1038/ncomms8357},
issn = {2041-1723},
}TY - JOUR
AU - Normanno, Davide
AU - Boudarene, Lydia
AU - Dugast-Darzacq, Claire
AU - Chen, Jiji
AU - Richter, Christian
AU - Proux, Florence
AU - Benichou, Olivier
AU - Voituriez, Raphael
AU - Darzacq, Xavier
AU - Dahan, Maxime
PY - 2015
DA - 2015/07/01
TI - Probing the target search of DNA-binding proteins in mammalian cells using TetR as model searcher
JO - NATURE COMMUNICATIONS
VL - 6
IS - 7357
SN - 2041-1723
AB - Many cellular functions rely on DNA-binding proteins finding and associating to specific sites in the genome. Yet the mechanisms underlying the target search remain poorly understood, especially in the case of the highly organized mammalian cell nucleus. Using as a model Tet repressors (TetRs) searching for a multi-array locus, we quantitatively analyse the search process in human cells with single-molecule tracking and single-cell protein-DNA association measurements. We find that TetRs explore the nucleus and reach their target by 3D diffusion interspersed with transient interactions with non-cognate sites, consistent with the facilitated diffusion model. Remarkably, nonspecific binding times are broadly distributed, underlining a lack of clear delimitation between specific and nonspecific interactions. However, the search kinetics is not determined by diffusive transport but by the low association rate to nonspecific sites. Altogether, our results provide a comprehensive view of the recruitment dynamics of proteins at specific loci in mammalian cells.
DO - 10.1038/ncomms8357
UR - http://www.dx.doi.org/10.1038/ncomms8357
ER -