Nucleosomes represent a challenge in regard to transcription. Histone eviction enables RNA polymerase II (RNAPII) progression through DNA, but compromises chromatin integrity. Here, we used the SNAP-tag system to distinguish new and old histones and monitor chromatin reassembly coupled to transcription in human cells. We uncovered a transcription-dependent loss of old histone variants H3.1 and H3.3. At transcriptionally active domains, H3.3 enrichment reflected both old H3.3 retention and new deposition. Mechanistically, we found that the histone regulator A (HIRA) chaperone is critical to processing both new and old H3.3 via different pathways. De novo H3.3 deposition is totally dependent on HIRA trimerization as well as on its partner ubinuclein 1 (UBN1), while antisilencing function 1 (ASF1) interaction with HIRA can be bypassed. By contrast, recycling of H3.3 requires HIRA but proceeds independently of UBN1 or HIRA trimerization and shows absolute dependency on ASF1–HIRA interaction. We propose a model whereby HIRA coordinates these distinct pathways during transcription to fine-tune chromatin states.
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The raw image dataset on which this paper is based is too large to be rendered available via public repositories, but is fully available upon request. Associated analysis code is also available upon request. All measurements reported in the figures are available as Source data files.
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We thank C. Domrane for generation of preliminary data with transcription inhibitors. We thank J.-P. Quivy and D. Jeffery for critical reading of the manuscript, and the PICT-IBiSA@Pasteur Imaging Facility of the Institut Curie, a member of the France Bioimaging National Infrastructure (no. ANR-10-INBS-04). This work was supported by la Ligue Nationale contre le Cancer (Equipe labellisée Ligue): ANR-11-LABX-0044_DEEP and ANR-10-IDEX-0001-02 PSL, ANR-12-BSV5-0022-02 ‘CHAPINHIB’, ANR-14-CE16-0009 ‘Epicure’, ANR-14-CE10-0013 ‘CELLECTCHIP’, EU project 678563 ‘EPOCH28’, ERC-2015-ADG-694694 ‘ChromADICT’, ANR-16-CE15-0018 ‘CHRODYT’, ANR-16-CE12-0024 ‘CHIFT’, ANR-16-CE11-0028 ‘REPLICAF’ and PSL-AFdL ‘TRACK’.
The authors declare no competing interests.
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Torné, J., Ray-Gallet, D., Boyarchuk, E. et al. Two HIRA-dependent pathways mediate H3.3 de novo deposition and recycling during transcription. Nat Struct Mol Biol 27, 1057–1068 (2020). https://doi.org/10.1038/s41594-020-0492-7
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