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A conformational switch in HP1 releases auto-inhibition to drive heterochromatin assembly

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Abstract

A hallmark of histone H3 lysine 9 (H3K9)-methylated heterochromatin, conserved from the fission yeast Schizosaccharomyces pombe to humans, is its ability to spread to adjacent genomic regions1,2,3,4,5,6. Central to heterochromatin spread is heterochromatin protein 1 (HP1), which recognizes H3K9-methylated chromatin, oligomerizes and forms a versatile platform that participates in diverse nuclear functions, ranging from gene silencing to chromosome segregation1,2,3,4,5,6. How HP1 proteins assemble on methylated nucleosomal templates and how the HP1–nucleosome complex achieves functional versatility remain poorly understood. Here we show that binding of the key S. pombe HP1 protein, Swi6, to methylated nucleosomes drives a switch from an auto-inhibited state to a spreading-competent state. In the auto-inhibited state, a histone-mimic sequence in one Swi6 monomer blocks methyl-mark recognition by the chromodomain of another monomer. Auto-inhibition is relieved by recognition of two template features, the H3K9 methyl mark and nucleosomal DNA. Cryo-electron-microscopy-based reconstruction of the Swi6–nucleosome complex provides the overall architecture of the spreading-competent state in which two unbound chromodomain sticky ends appear exposed. Disruption of the switch between the auto-inhibited and spreading-competent states disrupts heterochromatin assembly and gene silencing in vivo. These findings are reminiscent of other conditionally activated polymerization processes, such as actin nucleation, and open up a new class of regulatory mechanisms that operate on chromatin in vivo.

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Figure 1: Dissecting Swi6 self-association equilibria.
Figure 2: Impact of disrupting H3 tail mimic–CD interaction.
Figure 3: Electron microscopy studies of Swi6 and Swi6–H3K C 9me3 nucleosome complex.
Figure 4: Nucleosome recognition and in vivo impact of disrupting loop–CD interaction.

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Acknowledgements

We thank J. Tretyakova for preparation of histone proteins and J. Leonard for sample preparation for cryo-electron microscopy of nucleosome alone. We thank W. Lim, M. Simon, K. Armache, J. Zalatan, L. Racki and members of the Narlikar laboratory for discussions. D.C. would like to thank I. Ortiz Torres and K. M. Kuchenbecker for scientific discussions and members of the Schuck laboratory for advice on AUC approaches. This work was supported by a grant from the Hillblom foundation to D.C., by grants from the American Cancer Society and Leukemia and Lymphoma Society to G.J.N., National Institutes of Health (NIH) grant R01GM071801 to H.D.M. and by a New Technology Award to Y.C. from the UCSF Program for Breakthrough Biomedical Research. P.S. was supported by the Intramural Research Program of the NIBIB, NIH. N.N. and E.P. were supported by the NIH grant AR053720.

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Contributions

D.C. and G.J.N. identified, developed and addressed the core questions. D.C. performed the bulk of the experiments. P.S. trained D.C. in the use of AUC approaches and was instrumental in interpreting the AUC data. N.N. performed the EPR experiments. D.B.M. trained D.C. in strain construction and in the use of S. pombe assays. J.F.G. constructed some of the S. pombe strains and performed initial in vivo experiments. E.P., R.C., A.L. and D.C. deconvolved the EPR spectra. S.W. generated the cryo-electron-microscopy reconstruction of the nucleosome alone. M.L. generated the EM reconstructions of the Swi6–nucleosome complex and the two-dimensional reconstructions of the CFP–Swi6 and Swi6–CFP constructs. M.L., S.W. and Y.C. analysed the electron microscopy data. Y.C. oversaw all of the electron microscopy studies. H.D.M. oversaw the design and interpretation of the in vivo experiments. R.C. oversaw the EPR analysis and interpretation. D.C. and G.J.N. wrote the bulk of the manuscript with substantial intellectual contributions from R.C.

Corresponding author

Correspondence to Geeta J. Narlikar.

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The authors declare no competing financial interests.

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Canzio, D., Liao, M., Naber, N. et al. A conformational switch in HP1 releases auto-inhibition to drive heterochromatin assembly. Nature 496, 377–381 (2013). https://doi.org/10.1038/nature12032

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