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Multiple-binding-site mechanism explains concentration-dependent unbinding rates of DNA-binding proteins.

ABSTRACT: Recent work has demonstrated concentration-dependent unbinding rates of proteins from DNA, using fluorescence visualization of the bacterial nucleoid protein Fis [Graham et al. (2011) (Concentration-dependent exchange accelerates turnover of proteins bound to double-stranded DNA. Nucleic Acids Res., 39:2249)]. The physical origin of this concentration-dependence is unexplained. We use a combination of coarse-grained simulation and theory to demonstrate that this behavior can be explained by taking into account the dimeric nature of the protein, which permits partial dissociation and exchange with other proteins in solution. Concentration-dependent unbinding is generated by this simple model, quantitatively explaining experimental data. This effect is likely to play a major role in determining binding lifetimes of proteins in vivo where there are very high concentrations of solvated molecules.

SUBMITTER: Sing CE

PROVIDER: S-EPMC3973338 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature

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Multiple-binding-site mechanism explains concentration-dependent unbinding rates of DNA-binding proteins.

Sing Charles E CE Olvera de la Cruz Monica M Marko John F JF

Nucleic acids research 20140106 6

Recent work has demonstrated concentration-dependent unbinding rates of proteins from DNA, using fluorescence visualization of the bacterial nucleoid protein Fis [Graham et al. (2011) (Concentration-dependent exchange accelerates turnover of proteins bound to double-stranded DNA. Nucleic Acids Res., 39:2249)]. The physical origin of this concentration-dependence is unexplained. We use a combination of coarse-grained simulation and theory to demonstrate that this behavior can be explained by taki ...[more]

PMID: 24393773

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Project description:Runx1 and Runx3 function redundantly in early T-development, and together drive T-lineage developmental progression by regulating distinct sets of genes in different stages. Context-specific Runx target genes are particularly enriched near Runx binding sites that dynamically shift from pre-T-commitment stages (Phase 1) to post-T-commitment stages (Phase 2). As total Runx activities (Runx1+Runx3) are maintained stably throughout early T-development stages, yet Runx factors physically interact with multiple collaborators, we hypothesized that different Runx-collaborating transcription factors compete to recruit a limited pool of Runx transcription factors. To test whether increasing Runx availability can alter Runx DNA-binding site choices, Runx1 overexpression vectors were introduced to two different systems. First, we increased Runx1 expression in a DN3-like cell line (representing a post-commitment, Phase 2 stage) in the presence or absence of exogenous Phase 1 co-factor, PU.1. Second, we increased Runx1 expression in Phase 1 primary cells which naturally express PU.1 and other Phase 1 collaborators. Then, Runx1 binding behaviors were measured using ChIP-seq or CUT&RUN (C&R). A modest increase of Runx1 levels (about 2-3 fold increase) substantially increased the number of Runx binding sites seen and the intensity of occupancy in both systems. When Runx expression was at physiological levels, PU.1 dominated Runx1 site choice before T commitment by recruiting Runx1 to PU.1 sites. The Phase 2 cell line system showed that it did this while depleting Runx1 from alternative high quality Runx motif sites. However, when Runx1 was overexpressed, Runx1 was still recruited to PU.1 sites, but this recruitment did not evacuate Runx1 occupancy from default preferred sites. Notably, Runx1 overexpression in primary Phase 1 cells caused precocious occupancy of post-commitment, Phase 2-specific sites. We found that these are often co-occupied with TCF1, E2A, and HEB, but have minimal co-binding with PU.1. In addition, Runx1 overexpression resulted in new binding sites that were not normally observed in pro-T cells, which are mostly sequestered by closed chromatin normally although they harbor more numerous Runx motifs. Thus, these data suggest that Runx DNA binding site choices are sensitive to Runx concentration and co-factors during early T-development.

Dataset Information

Multiple-binding-site mechanism explains concentration-dependent unbinding rates of DNA-binding proteins.

Publications

Multiple-binding-site mechanism explains concentration-dependent unbinding rates of DNA-binding proteins.

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