Measurement of histone replacement dynamics with genetically encoded exchange timers in yeast thumbnail

Measurement of histone replacement dynamics with genetically encoded exchange timers in yeast

Abstract

Histone exchange between histones carrying position-specific marks and histones bearing general marks is important for gene regulation, but understanding of histone exchange remains incomplete. To overcome the poor time resolution of conventional pulse–chase histone labeling, we present a genetically encoded histone exchange timer sensitive to the duration that two tagged histone subunits co-reside at an individual genomic locus. We apply these sensors to map genome-wide patterns of histone exchange in yeast using single samples. Comparing H3 exchange in cycling and G1-arrested cells suggests that replication-independent H3 exchange occurs at several hundred nucleosomes (<1% of all nucleosomes) per minute, with a maximal rate at histone promoters. We observed substantial differences between the two nucleosome core subcomplexes: H2A-H2B subcomplexes undergo rapid transcription-dependent replacement within coding regions, whereas H3-H4 replacement occurs predominantly within promoter nucleosomes, in association with gene activation or repression. Our timers allow the in vivo study of histone exchange dynamics with minute time scale resolution.

Access options

Subscribe to Journal

Get full journal access for 1 year

$59.00

only $4.92 per issue

All prices are NET prices.

VAT will be added later in the checkout.

Tax calculation will be finalised during checkout.

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Data availability

All figures were generated from the raw sequencing data accessible at accession number GSE157402. Source data are provided with this paper.

References

  1. 1.

    Luger, K., Mäder, A. W., Richmond, R. K., Sargent, D. F. & Richmond, T. J. Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389, 251–260 (1997).

  2. 2.

    Gossett, A. J. & Lieb, J. D. In vivo effects of histone H3 depletion on nucleosome occupancy and position in Saccharomyces cerevisiae. PLoS Genet. 8, e1002771 (2012).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  3. 3.

    Kim, U. J., Han, M., Kayne, P. & Grunstein, M. Effects of histone H4 depletion on the cell cycle and transcription of Saccharomyces cerevisiae. EMBO J. 7, 2211–2219 (1988).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  4. 4.

    Han, M., Kim, U. J., Kayne, P. & Grunstein, M. Depletion of histone H4 and nucleosomes activates the PHO5 gene in Saccharomyces cerevisiae. EMBO J. 7, 2221–2228 (1988).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  5. 5.

    Bannister, A. J. & Kouzarides, T. Regulation of chromatin by histone modifications. Cell Res. 21, 381–395 (2011).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  6. 6.

    Rando, O. J. & Winston, F. Chromatin and transcription in yeast. Genetics 190, 351–387 (2012).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  7. 7.

    Unnikrishnan, A., Gafken, P. R. & Tsukiyama, T. Dynamic changes in histone acetylation regulate origins of DNA replication. Nat. Struct. Mol. Biol. 17, 430–437 (2010).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  8. 8.

    Owen-Hughes, T. & Gkikopoulos, T. Making sense of transcribing chromatin. Curr. Opin. Cell Biol. 24, 296–304 (2012).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  9. 9.

    Rodríguez-Navarro, S. Insights into SAGA function during gene expression. EMBO Rep. 10, 843–850 (2009).

    PubMed 
    PubMed Central 
    Article 
    CAS 

    Google Scholar
     

  10. 10.

    Benson, L. J. et al. Modifications of H3 and H4 during chromatin replication, nucleosome assembly, and histone exchange. J. Biol. Chem. 281, 9287–9296 (2006).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  11. 11.

    Sobel, R. E., Cook, R. G., Perry, C. A., Annunziato, A. T. & Allis, C. D. Conservation of deposition-related acetylation sites in newly synthesized histones H3 and H4. Proc. Natl Acad. Sci. USA 92, 1237–1241 (1995).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  12. 12.

    Han, J., Zhou, H., Li, Z., Xu, R. M. & Zhang, Z. The Rtt109-Vps75 histone acetyltransferase complex acetylates non-nucleosomal histone H3. J. Biol. Chem. 282, 14158–14164 (2007).

    CAS 
    PubMed 
    Article 
    PubMed Central 

    Google Scholar
     

  13. 13.

    Bar-Ziv, R., Voichek, Y. & Barkai, N. Chromatin dynamics during DNA replication. Genome Res. 26, 1245–1256 (2016).

  14. 14.

    Deal, R. B., Henikoff, J. G. & Henikoff, S. Genome-wide kinetics of nucleosome turnover determined by metabolic labelinq of histones. Science 328, 1161–1164 (2010).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  15. 15.

    Verzijlbergen, K. F. et al. Recombination-induced tag exchange to track old and new proteins. Proc. Natl Acad. Sci. USA 107, 64–68 (2010).

    CAS 
    PubMed 
    Article 
    PubMed Central 

    Google Scholar
     

  16. 16.

    Jamai, A., Imoberdorf, R. M. & Strubin, M. Continuous histone H2B and transcription-dependent histone H3 exchange in yeast cells outside of replication. Mol. Cell 25, 345–355 (2007).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  17. 17.

    Dion, M. F. et al. Dynamics of replication-independent histone turnover in budding yeast. Science 315, 1405–1408 (2007).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  18. 18.

    Rufiange, A., Jacques, P. É., Bhat, W., Robert, F. & Nourani, A. Genome-wide replication-independent histone H3 exchange occurs predominantly at promoters and implicates H3 K56 acetylation and Asf1. Mol. Cell 27, 393–405 (2007).

  19. 19.

    Molenaar, T. M., Pagès-Gallego, M., Meyn, V. & van Leeuwen, F. Application of recombination -induced tag exchange (RITE) to study histone dynamics in human cells. Epigenetics 15, 901–913 (2020).

    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  20. 20.

    Yi, L. et al. Engineering of TEV protease variants by yeast ER sequestration screening (YESS) of combinatorial libraries. Proc. Natl Acad. Sci. USA 110, 7229–7234 (2013).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  21. 21.

    Weiner, A. et al. High-resolution chromatin dynamics during a yeast stress response. Mol. Cell 58, 371–386 (2015).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  22. 22.

    Kaplan, T. et al. Cell cycle- and chaperone-mediated regulation of H3K56ac incorporation in yeast. PLoS Genet. 4, e1000270 (2008).

  23. 23.

    Ferrari, P. & Strubin, M. Uncoupling histone turnover from transcription-associated histone H3 modifications. Nucleic Acids Res. 43, 3972–3985 (2015).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  24. 24.

    Senshu, T., Fukuda, M. & Ohashi, M. Preferential association of newly synthiesized H3 and H4 histones with newly replicated DNA. J. Biochem. 84, 985–988 (1978).

  25. 25.

    Worcel, A., Han, S. & Wong, M. L. Assembly of newly replicated chromatin. Cell 15, 969–977 (1978).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  26. 26.

    Kireeva, M. L. et al. Nucleosome remodeling induced by RNA polymerase II: loss of the H2A/H2B dimer during transcription. Mol. Cell 9, 541–552 (2002).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  27. 27.

    Kulaeva, O. I., Hsieh, F. K. & Studitsky, V. M. RNA polymerase complexes cooperate to relieve the nucleosomal barrier and evict histones. Proc. Natl Acad. Sci. USA 107, 11325–11330 (2010).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  28. 28.

    Sheinin, M. Y., Li, M., Soltani, M., Luger, K. & Wang, M. D. Torque modulates nucleosome stability and facilitates H2A/H2B dimer loss. Nat. Commun. 4, 1–8 (2013).

    Article 
    CAS 

    Google Scholar
     

  29. 29.

    Ramachandran, S., Ahmad, K. & Henikoff, S. Transcription and remodeling produce asymmetrically unwrapped nucleosomal intermediates. Mol. Cell 68, 1038–1053 (2017).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  30. 30.

    Kimura, H. & Cook, P. R. Kinetics of core histones in living human cells. J. Cell Biol. 153, 1341–1354 (2001).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  31. 31.

    Segal, E. et al. A genomic code for nucleosome positioning. Nature 442, 772–778 (2006).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  32. 32.

    Murray, S. C. et al. Sense and antisense transcription are associated with distinct chromatin architectures across genes. Nucleic Acids Res. 43, 7823–7837 (2015).

  33. 33.

    Cheung, V. et al. Chromatin- and transcription-related factors repress transcription from within coding regions throughout the Saccharomyces cerevisiae genome. PLoS Biol. 6, e277 (2008).

  34. 34.

    Murawska, M. et al. The chaperone FACT and histone H2B ubiquitination maintain S. pombe genome architecture through genic and subtelomeric functions. Mol. Cell 77, 501–513 (2020).

  35. 35.

    Ivanovska, I., Jacques, P.-E., Rando, O. J., Robert, F. & Winston, F. Control of chromatin structure by Spt6: different consequences in coding and regulatory regions. Mol. Cell. Biol. 31, 531–541 (2011).

  36. 36.

    Jeronimo, C., Poitras, C. & Robert, F. Histone recycling by FACT and Spt6 during transcription prevents the scrambling of histone modifications. Cell Rep. 28, 1206–1218 (2019).

  37. 37.

    DeGennaro, C. M. et al. Spt6 regulates intragenic and antisense transcription, nucleosome positioning, and histone modifications genome-wide in fission yeast. Mol. Cell. Biol. 33, 4779–4792 (2013).

  38. 38.

    Boisnard, S. et al. H2O2 activates the nuclear localization of Msn2 and Maf1 through thioredoxins in Saccharomyces cerevisiae. Eukaryot. Cell 8, 1429–1438 (2009).

  39. 39.

    Rodrigues-Pousada, C. et al. Yeast AP-1 like transcription factors (Yap) and stress response: a current overview. Microb. Cell 6, 267–285 (2019).

  40. 40.

    Kassem, S. et al. Histone exchange is associated with activator function at transcribed promoters and with repression at histone loci. Sci. Adv. https://doi.org/10.1126/sciadv.abb0333 (2020).

    Article 
    PubMed 
    PubMed Central 

    Google Scholar
     

  41. 41.

    Lehner, B. Conflict between noise and plasticity in yeast. PLoS Genet. 6, e1001185 (2010).

  42. 42.

    Tirosh, I. & Barkai, N. Two strategies for gene regulation by promoter nucleosomes. Genome Res. 18, 1084–1091 (2008).

  43. 43.

    Spector, M. S., Raff, A., DeSilva, H., Lee, K. & Osley, M. A. Hir1p and Hir2p function as transcriptional corepressors to regulate histone gene transcription in the Saccharomyces cerevisiae cell cycle. Mol. Cell. Biol. 17, 545–552 (1997).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  44. 44.

    Sherwood, P. W., Tsang, S. V. & Osley, M. A. Characterization of HIR1 and HIR2, two genes required for regulation of histone gene transcription in Saccharomyces cerevisiae. Mol. Cell. Biol. 13, 28–38 (1993).

    CAS 
    PubMed 
    PubMed Central 

    Google Scholar
     

  45. 45.

    Sutton, A., Bucaria, J., Osley, M. A. & Sternglanz, R. Yeast ASF1 protein is required for cell cycle regulation of histone gene transcription. Genetics 158, 587–596 (2001).

  46. 46.

    Xu, H., Kim, U. J., Schuster, T. & Grunstein, M. Identification of a new set of cell cycle-regulatory genes that regulate S-phase transcription of histone genes in Saccharomyces cerevisiae. Mol. Cell. Biol. 12, 5249–5259 (1992).

    CAS 
    PubMed 
    PubMed Central 

    Google Scholar
     

  47. 47.

    Jackson, V. In vivo studies on the dynamics of histone–DNA interaction: evidence for nucleosome dissolution during replication and transcription and a low level of dissolution independent of both. Biochemistry 29, 719–731 (1990).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  48. 48.

    Schwabish, M. A. & Struhl, K. Asf1 mediates histone eviction and deposition during elongation by RNA polymerase II. Mol. Cell 22, 415–422 (2006).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  49. 49.

    Chen, X. et al. Histone chaperone Nap1 is a major regulator of histone H2A-H2B dynamics at the inducible GAL locus. Mol. Cell. Biol. 36, 1287–1296 (2016).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  50. 50.

    Chapal, M., Mintzer, S., Brodsky, S., Carmi, M. & Barkai, N. Resolving noise–control conflict by gene duplication. PLoS Biol. 17, e3000289 (2019).

  51. 51.

    Kemmeren, P. et al. Large-scale genetic perturbations reveal regulatory networks and an abundance of gene-specific repressors. Cell 157, 740–752 (2014).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  52. 52.

    Gietz, R. D. & Schiestl, R. H. High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method. Nat. Protoc. 2, 31 (2007).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  53. 53.

    Janke, C. et al. A versatile toolbox for PCR-based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes. Yeast 21, 947–962 (2004).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  54. 54.

    Schuldiner, M. et al. Exploration of the function and organization of the yeast early secretory pathway through an epistatic miniarray profile. Cell 123, 507–519 (2005).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  55. 55.

    Gutin, J. et al. Fine-resolution mapping of TF binding and chromatin interactions. Cell Rep. 22, 2797–2807 (2018).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  56. 56.

    Voichek, Y. et al. Epigenetic control of expression homeostasis during replication is stabilized by the replication checkpoint. Mol. Cell 70, 1121–1133 (2018).

    CAS 
    PubMed 
    Article 
    PubMed Central 

    Google Scholar
     

  57. 57.

    Quinlan, A. R. & Hall, I. M. BEDTools: a flexible suite of utilities for comparing genomic features. Bioinformatics 26, 841–842 (2010).

  58. 58.

    Chereji, R. V., Ramachandran, S., Bryson, T. D. & Henikoff, S. Precise genome-wide mapping of single nucleosomes and linkers in vivo. Genome Biol. 19, 1–20 (2018).

    Article 
    CAS 

    Google Scholar
     

  59. 59.

    Park, D., Morris, A. R., Battenhouse, A. & Iyer, V. R. Simultaneous mapping of transcript ends at single-nucleotide resolution and identification of widespread promoter-associated non-coding RNA governed by TATA elements. Nucleic Acids Res. 42, 3736–3749 (2014).

  60. 60.

    Yassoura, M. et al. Ab initio construction of a eukaryotic transcriptome by massively parallel mRNA sequencing. Proc. Natl Acad. Sci. USA.106, 3264–3269 (2009).

    Article 

    Google Scholar
     

  61. 61.

    Churchman, L. S. & Weissman, J. S. Nascent transcript sequencing visualizes transcription at nucleotide resolution. Nature 469, 368–373 (2011).

  62. 62.

    Kaplan, N. et al. The DNA-encoded nucleosome organization of a eukaryotic genome. Nature 458, 362–366 (2009).

    CAS 
    PubMed 
    Article 

    Google Scholar
     

  63. 63.

    Gasch, A. P. et al. Genomic expression programs in the response of yeast cells to environmental changes. Mol. Biol. Cell 11, 4241–4257 (2000).

    CAS 
    PubMed 
    PubMed Central 
    Article 

    Google Scholar
     

  64. 64.

    Basehoar, A. D., Zanton, S. J. & Pugh, B. F. Identification and distinct regulation of yeast TATA box-containing genes. Cell 116, 699–709 (2004).

  65. 65.

    Santos, A., Wernersson, R. & Jensen, L. J. Cyclebase 3.0: a multi-organism database on cell-cycle regulation and phenotypes. Nucleic Acids Res. 43, D1140–D1144 (2014).

    PubMed 
    PubMed Central 
    Article 
    CAS 

    Google Scholar
     

  66. 66.

    Brodsky, S. et al. Intrinsically disordered regions direct transcription factor in vivo binding specificity. Mol. Cell 79, 459–471 (2020).

Download references

Acknowledgements

We thank all lab members for critical comments throughout and careful reading of the manuscript. We also thank Y. Voichek, R. Bar-Ziv, E. Metzl-Raz and B. Shilo for comments on the manuscript. We are very grateful to R. Diskin for structural insights during the method development. This work was funded by the Israel Science Foundation FIRST Program (grant No. 966/19), ERC and the Minerva Foundation.

Author information

Author notes

  1. These authors contributed equally: Gilad Yaakov, Felix Jonas.

Affiliations

  1. Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel

    Gilad Yaakov, Felix Jonas & Naama Barkai

Contributions

G.Y. and N.B. conceived, developed and designed the sensors. G.Y. performed all experiments. F.J. and N.B. developed and performed all analyses. All authors designed the experiments, discussed the results and wrote the manuscript.

Corresponding authors

Correspondence to
Gilad Yaakov or Naama Barkai.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Additional information

Peer review information Nature Biotechnology thanks Frederic Berger and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Supporting figure for Fig. 1.

a, Pearson correlation across all nucleosomes for HA and myc ChIP-seq levels in strains with the HHT1 allele (median of 17 repeats), the alternate HHT2 H3 allele (3 repeats), or both H3 alleles (4 repeats) tagged with the sensor. b, Left: Pearson correlation between histone mark signal intensity on nucleosomes profiled from exponentially growing cells bearing the HHT1 H3 exchange sensor in this study (rows) with Weiner et al. (columns 21). Right: Mean histone mark (H3K79-trimethyl, H3K4-trimethyl and H3K9-acetyl) traces around the TSS of all genes in the H3 sensor strain. c, Ponceau staining corresponding to Western-blot in Fig. 1b. Three repeats were performed giving similar results. d, Myc and HA level of all nucleosomes for the non-cleavable (median of 3 repeats) and the slowly cleaved variant with a point mutation in the 7AA cleavage site (2 repeats). Rho indicates the Pearson correlation between both intensities. e, Comparison of measured H3 exchange (myc/HA) per nucleosome with an additional external dataset 18. f, Quantitative ChIP-qPCR measurements for absolute exchange (log2(myc/HA)) are plotted versus the relative ChIP-seq counterparts for alpha factor arrested cells (left) or asynchronously growing cells with both H3 alleles tagged with the sensor (right). Analysis was on selected nucleosomes as in Fig. 1f. 2 biological ChIP-qPCR repeats were performed, each consisting of 4 technical ChIP repeats and 2 technical qPCR repeats. The mean values with standard of error are shown.

Extended Data Fig. 2 Supporting figure for Fig. 2.

a, Comparison between HA occupancy signal of all nucleosomes in the H2A, H2B or H4 reporter strains with the H3 reporter strain. rho indicates Pearson correlation. b, Genome-wide Pearson correlation between myc and HA levels per nucleosome of all reporter strains. Median of all ChIP repeats is shown with the number of repeats ‘n’ for each strain indicated on the right. a indicates a strain with G133A in the HHT1 allele (see Methods), b indicates a strain in which TEV is fused to H2B rather than H2A, * indicates a strain in which the alternate allele was tagged (HHT2 for H3 and HHF1 for H4) and # indicates the H2B reporter in which both TEV and the sensor are fused to H2B alleles (HTB2 and HTB1 respectfully). c, Mean myc and HA signal around the TSS for the indicated sensors, binned by expression level (top) or plasticity (bottom). Note that both leftmost systems equally report on H3 dynamics, with fusing TEV to either H2A or H2B. myc/HA signal (log) is shown in the corresponding Fig. 2c. d, As in (c) for strains in which the H2A.Z variant (Htz1) has been deleted in the H2A (median of 4 repeats) or H3 (8 repeats) sensors.

Extended Data Fig. 3 Supporting figure for Fig. 3.

a, Calculated myc and HA nucleosome intensities across a broad kon versus koff parameter space (leftmost, kclv=1.5 and f = 1 as in Fig. 2a). Varying the fraction of myc in unbound histones (f = 0.5, second from left), or myc cleavage rate (kclv*=0.15, second from right) were further tested. HA levels in all cases remain unchanged (rightmost). All axes and color are in log-scale. b, HA signal intensity distribution of nucleosomes on A/T-rich (75 + /-2%) and A/T-poor (50 + /-2%) regions in the genome for H2A and H3 sensor strains. The average A/T content in yeast is 63%. c, Gene body (+2) nucleosome myc signal in H2A versus its respective HA signal (left) or versus H3 myc (right). Color indicates expression level of the gene corresponding to the given nucleosome. d, Mean myc and HA intensity as in Fig. 3c for nucleosomes -2 to +3 (n = 2561, 3500, 4778, 4768 and 4637 for the different positions) in the various reporter strains. For H2B, analyses are shown for strains in which TEV was fused to either H3 or the other allele of H2B as indicated. For H3, TEV was fused to either H2A or H2B. Moving average is calculated over 0.5 expression levels. Shaded area is standard error. e, Fold change in H2A (y-axis) and H3 myc (x-axis) nucleosome levels in Spt6 over-expressing versus wild type cells. f, Antisense transcription was ordered by expression level32. Mean myc and HA intensity are shown for nucleosomes -2 to +2. Shaded area is standard error. g, Candidate screen for chaperones involved in exchange. As in (d) for indicated histone chaperone mutants alongside wildtype for promoter (-1) and gene body (+2) nucleosomes. DAmP; Decreased Abundance by mRNA Perturbation 52, OE; overexpression by TEF1 promoter substitution of native promoter. Number of repeats for H2A/H3 systems correspondingly: OE-Spt6 n = 3/3, spt16DAmP n = 2/3, spt6DAmP 2/3, hir1 n = 2/3. Shaded area is standard error.

Extended Data Fig. 4 Supporting figure for Fig. 4.

a, HA and myc intensity dynamics for the indicated sensor at gene body nucleosomes (+2) during H2O2 exposure. Each column is a single (repressed or induced) gene that corresponds to the given nucleosome. Color indicates the signal intensity change relative to its median (log) at each nucleosome. b, Median gene expression change (log2) of annotated functional gene groups (stress genes, ribosomal biogenesis53) or internally derived expression clusters (induced, repressed). Corresponding mean myc and HA levels around the TSS of stress genes and ribosomal biogenesis genes. c, As in (a) for promoter nucleosomes (-2) in the H3 reporter system. d, Top: Yap1 and Msn2 ChEC binding signal on gene promoters from54. The dashed line indicates the chosen binding threshold for Yap1 and Msn2 target genes. Color indicates the gene’s maximal expression level change during H2O2 exposure. 9 Yap1 targets that do not induce expression are marked in red squares. Bottom: Median Msn2 and Yap1 ChEC signal around the determined Msn2 and Yap1 binding sites. e, Median intensities around the Yap1 and Msn2 binding sites on target gene promoters during H2O2 exposure in the H2A reporter system. The number in brackets indicates the number of target genes for each transcription factor.

Extended Data Fig. 5 Supporting figure for Fig. 5.

a, Gene state (induced or repressed, Fig. 5a) was defined by the median expression change (Y-axis) versus plasticity (standard deviation, X-axis) in a deletion collection 51. Dashed lines indicate the threshold (delta median over 2-fold standard error) used to define high confidence genes with increased (red) or decreased (green) mRNA in exponentially growing wild type cells. b, Histogram of plasticity scores derived in (a) for TATA and TATA-less gene groups55. c, Expression plasticity vs. expression level for promoter (-2) and gene body (+2) nucleosomes. Color indicates myc intensity in the H3 or H2A reporter system respectively. d, Mean exchange signals for all reporter strains at promoter nucleosomes (-2 and -1, n = 1503 and 2028), as a function of gene expression plasticity defined in (a). Shaded area is standard error. e, Mean HA intensity on promoters in Hir1-deleted versus wild-type cells, corresponds to Fig. 5b. Rho indicates the Pearson correlation, and cyan diamonds highlight histone promoters. f, HA and myc for Asf1-deleted cells as in (e). g, Mean HA intensity for wild type, Asf1, Hir1 and Hir2-delted cells around two histone loci, corresponds to Fig. 4c. Refer to main figure for number of repeats in (eg).

Supplementary information

About this article

Verify currency and authenticity via CrossMark

Cite this article

Yaakov, G., Jonas, F. & Barkai, N. Measurement of histone replacement dynamics with genetically encoded exchange timers in yeast.
Nat Biotechnol (2021). https://doi.org/10.1038/s41587-021-00959-8

Download citation

Read More

Leave a Reply

Your email address will not be published. Required fields are marked *