Single molecule tracking of Ace1p in Saccharomyces cerevisiae defines a characteristic residence time for non-specific interactions of transcription factors with chromatin

David Ball; Gunjan D. Mehta; Ronit Salomon-Kent; Davide Mazza; Tatsuya Morisaki; Florian Mueller; James G. Mcnally; Tatiana S. Karpova

Single molecule tracking of Ace1p in Saccharomyces cerevisiae defines a characteristic residence time for non-specific interactions of transcription factors with chromatin

David Ball ¹ Gunjan D. Mehta ¹ Ronit Salomon-Kent ¹ Davide Mazza ² Tatsuya Morisaki ³ Florian Mueller ⁴ James G. Mcnally ⁵ Tatiana S. Karpova ^{1, *}

* Corresponding author

1 NIH - National Cancer Institute

2 San Raffaele Scientific Institute

3 CSU - Colorado State University [Fort Collins]

4 Imagerie et Modélisation

5 Institute for Soft Matter and Functional Materials [Berlin]

Abstract : In vivo single molecule tracking has recently developed into a powerful technique for measuring and understanding the transient interactions of transcription factors (TF) with their chromatin response elements. However, this method still lacks a solid foundation for distinguishing between specific and non-specific interactions. To address this issue, we took advantage of the power of molecular genetics of yeast. Yeast TF Ace1p has only five specific sites in the genome and thus serves as a benchmark to distinguish specific from non-specific binding. Here, we show that the estimated residence time of the short-residence molecules is essentially the same for Hht1p, Ace1p and Hsf1p, equaling 0.12–0.32 s. These three DNA-binding proteins are very different in their structure, function and intracellular concentration. This suggests that (i) short-residence molecules are bound to DNA non-specifically, and (ii) that non-specific binding shares common characteristics between vastly different DNA-bound proteins and thus may have a common underlying mechanism. We develop new and robust procedure for evaluation of adverse effects of labeling, and new quantitative analysis procedures that significantly improve residence time measurements by accounting for fluorophore blinking. Our results provide a framework for the reliable performance and analysis of single molecule TF experiments in yeast.

Keywords : blinking chromatin dna yeasts transcription factor molecule

Document type :

Journal articles

Domain :

Life Sciences [q-bio]

Complete list of metadatas

Cited literature [31 references] Display Hide Download

https://hal-pasteur.archives-ouvertes.fr/pasteur-01622680
Contributor : Florian Mueller <>
Submitted on : Tuesday, October 24, 2017 - 3:37:00 PM
Last modification on : Saturday, April 28, 2018 - 9:02:01 PM

2016_Ball_NAR_SMT-in-yeast.pdf

Publication funded by an institution

Single molecule tracking of Ace1p in Saccharomyces cerevisiae defines a characteristic residence time for non-specific interactions of transcription factors with chromatin

File

Licence

Identifiers

Collections

Citation

Export

Metrics

107

26

Contact

Single molecule tracking of Ace1p in Saccharomyces cerevisiae defines a characteristic residence time for non-specific interactions of transcription factors with chromatin

File

Licence

Identifiers

Collections

Citation

Export

Share

Metrics

107

26

Contact