Dynamics of nucleosome remodelling by individual ACF complexes

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Abstract

The ATP-dependent chromatin assembly and remodelling factor (ACF) functions to generate regularly spaced nucleosomes, which are required for heritable gene silencing. The mechanism by which ACF mobilizes nucleosomes remains poorly understood. Here we report a single-molecule FRET study that monitors the remodelling of individual nucleosomes by ACF in real time, revealing previously unknown remodelling intermediates and dynamics. In the presence of ACF and ATP, the nucleosomes exhibit gradual translocation along DNA interrupted by well-defined kinetic pauses that occurred after approximately seven or three to four base pairs of translocation. The binding of ACF, translocation of DNA and exiting of translocation pauses are all ATP-dependent, revealing three distinct functional roles of ATP during remodelling. At equilibrium, a continuously bound ACF complex can move the nucleosome back-and-forth many times before dissociation, indicating that ACF is a highly processive and bidirectional nucleosome translocase.

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Figure 1: Monitoring ACF-catalysed nucleosome remodelling by single-molecule FRET.
Figure 2: Real-time dynamics of ACF-catalysed nucleosome translocation.
Figure 3: ACF-catalysed nucleosome translocation is interrupted by well defined kinetic pauses.
Figure 4: ACF catalyses processive and bidirectional nucleosome translocation.

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Acknowledgements

We thank J. Widom for providing the plasmid containing the 601 positioning sequence and R. E. Kingston for the plasmids containing the SNF2h and Acf1 genes. We also thank L. Racki and E. Abbondanzieri for helpful discussions, and W. Huang and B. Harada for help with some experiments. This work is supported in part by Howard Hughes Medical Institute (to X.Z.) and the National Institutes of Health (GM073767) and the Beckman Foundation (to G.J.N). X.Z. is a Howard Hughes Medical Institute investigator. M.D.S. was a NIH Ruth L. Kirschstein NSRA Fellow, G.J.N is a Leukemia and Lymphoma Society Scholar.

Author Contributions T.R.B. performed the experiments and analysis with help from M.D.S.; J.G.Y. made the enzymes and histone proteins. T.R.B., G.J.N. and X.Z. designed the experiments. X.Z. oversaw the project.

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Correspondence to Geeta J. Narlikar or Xiaowei Zhuang.

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Blosser, T., Yang, J., Stone, M. et al. Dynamics of nucleosome remodelling by individual ACF complexes. Nature 462, 1022–1027 (2009) doi:10.1038/nature08627

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