Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

The RCAF complex mediates chromatin assembly during DNA replication and repair

Nature volume 402, pages 555–560 (1999)Cite this article

Abstract

Chromatin assembly is a fundamental biological process that is essential for the replication and maintenance of the eukaryotic genome1,2,3,4. In dividing cells, newly synthesized DNA is rapidly assembled into chromatin by the deposition of a tetramer of the histone proteins H3 and H4, followed by the deposition of two dimers of histones H2A and H2B to complete the nucleosome—the fundamental repeating unit of chromatin5. Here we describe the identification, purification, cloning, and characterization of replication-coupling assembly factor (RCAF), a novel protein complex that facilitates the assembly of nucleosomes onto newly replicated DNA in vitro. RCAF comprises the Drosophila homologue of anti-silencing function 1 protein ASF16 and histones H3 and H4. The specific acetylation pattern of H3 and H4 in RCAF is identical to that of newly synthesized histones. Genetic analyses in Saccharomyces cerevisiae demonstrate that ASF1 is essential for normal cell cycle progression, and suggest that RCAF mediates chromatin assembly after DNA replication and the repair of double-strand DNA damage in vivo.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Purification of RCAF activity.
Figure 2: RCAF cannot be replaced by histones and acts synergistically with CAF-1 in the assembly of chromatin on to newly replicated DNA.
Figure 3: Defects in growth and transcriptional silencing after the loss of both RCAF and CAF-1 activity.
Figure 4: Effects of disruptions to ASF1 and CAC1 on sensitivity to mutagens.

Similar content being viewed by others

References

  1. Grunstein,M. Histone acetylation in chromatin structure and transcription. Nature 389, 349–352 (1997).

    Article  ADS  CAS  Google Scholar 

  2. Kornberg,R. D. & Lorch,Y. Chromatin-modifying and -remodelling complexes. Curr. Opin. Genet. Dev. 9, 148–151 (1999).

    Article  CAS  Google Scholar 

  3. Ito,T., Tyler,J. K. & Kadonaga,J. T. Chromatin assembly factors: a dual function in nucleosome formation and mobilization? Genes Cells 2, 593–600 (1997).

    Article  CAS  Google Scholar 

  4. Adams,C. R. & Kamakaka,R. T. Chromatin assembly: biochemical identities and genetic redundancy. Curr. Opin. Genet. Dev. 9, 185–190 (1999).

    Article  CAS  Google Scholar 

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

    Article  ADS  CAS  Google Scholar 

  6. Le,S., Davis,C., Konopka,J. B. & Sternglanz,R. Two new S-phase specific genes from Saccharomyces cerevisiae. Yeast 13, 1029–1042 (1997).

    Article  CAS  Google Scholar 

  7. Stillman,B. W. & Gluzman,Y. Replication and supercoiling of simian virus 40 DNA in cell extracts from human cells. Mol. Cell. Biol. 5, 2051–2060 (1985).

    Article  CAS  Google Scholar 

  8. Stillman,B. Chromatin assembly during SV40 DNA replication in vitro. Cell 45, 555–565 (1986).

    Article  CAS  Google Scholar 

  9. Smith,S. & Stillman,B. Purification and characterization of CAF-I, a human cell factor required for chromatin assembly during DNA replication in vitro. Cell 58, 15–25 (1989).

    Article  CAS  Google Scholar 

  10. Tyler,J. K., Bulger,M., Kamakaka,R. T., Kobayashi,R. & Kodonaga,J. T. The p55 subunit of Drosophila chromatin assembly factor 1 is homologous to a histone deacetylase-associated protein. Mol. Cell. Biol. 16, 6149–6159 (1996).

    Article  CAS  Google Scholar 

  11. Ito,T., Bulger,M., Pazin,M. J., Kobayashi,R. & Kadonaga,J. T. ACF, an ISWI-containing and ATP-utilizing chromatin assembly and remodeling factor. Cell 90, 145–155 (1997).

    Article  CAS  Google Scholar 

  12. Ito,T., Bulger,M. Kobayashi,R. & Kadonaga,J. T. Drosophila NAP-1 is a core histone chaperone that functions in ATP-facilitated assembly fo regularly spaced nucleosomal arrays. Mol. Cell. Biol. 16, 3112–3124 (1996).

    Article  CAS  Google Scholar 

  13. Ito,T., Tyler,J. K., Bulger,M., Kobayashi,R. & Kadonaga,J. T. ATP-facilitated chromatin assembly with a nucleoplasmid-like protein from Drosophila melanogaster. J. Biol. Chem. 271, 25041–25048 (1996).

    Article  CAS  Google Scholar 

  14. Singer,M. S. et al. Identification of high-copy disruptors of telomeric silencing in Saccharomyces cerevisiae. Genetics 150, 613–632 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Spellman,P. K. et al. Comprehensive identification of cell cycle-regulated genes of the yeast Saccharomyces cerevisiae by microarray hybridization. Mol. Biol. Cell 9, 3273–3287 (1998).

    Article  CAS  Google Scholar 

  16. 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).

    Article  ADS  CAS  Google Scholar 

  17. McIntosh,E. M. MCB elements and the regulation of DNA replication genes in yeast. Curr. Genet. 24, 185–192 (1993).

    Article  CAS  Google Scholar 

  18. Verreault,A., Kaufman,P. D., Kobayashi,R. & Stillman,B. Nucleosome assembly by a complex of CAF-I and acetylated histones H3/H4. Cell 87, 95–104 (1996).

    Article  CAS  Google Scholar 

  19. Kaufman,P. D., Kobayashi,R. & Stillman,B. Ultraviolet radiation sensitivity and reduction of telomeric silencing in Saccharomyces cerevisiae cells lacking chromatin assembly factor-I. Genes Dev. 11, 345–357 (1997).

    Article  CAS  Google Scholar 

  20. Ling,X., Harkness,T. A. A., Schultz,M. C., Fisher-Adams,G. & Grunstein,M. Yeast histone H3 and H4 amino termini are important for nucleosome assembly in vivo and in vitro: redundant and position-independent functions in assembly but not in gene regulation. Genes Dev. 10, 686–699 (1996).

    Article  CAS  Google Scholar 

  21. Gottschling,D. E., Aparicio,O. M., Billington,B. L. & Zakian,V. A. Position effect at S. cerevisiae telomeres: reversible repression of Pol II transcription. Cell 63, 751–762 (1990).

    Article  CAS  Google Scholar 

  22. Monson,E. K., Bruin,D. de & Zakian,V. A. The yeast cac 1 protein is required for the stable inheritance of transcriptionally repressed chromatin at telomeres. Proc. Natl Acad. Sci. USA 94, 13081–13086 (1997).

    Article  ADS  CAS  Google Scholar 

  23. Schwartz,J. L. Monofunctional alkylating agent-induced S-phase-dependent DNA damage. Mutant. Res. 216, 111–118 (1989).

    Article  CAS  Google Scholar 

  24. Povirk,L. F. DNA damage and mutagenesis by radiomimetic DNA-cleaving agents: bleomycin, neocarzinostatin and other enediynes. Mutat. Res. 355, 71–89 (1996).

    Article  Google Scholar 

  25. Elledge,S. J. Cell cycle checkpoints: preventing an identity crisis. Science 274, 1664–1672 (1996).

    Article  ADS  CAS  Google Scholar 

  26. Han,M., Chang,M., Kim,U.-J. & Grunstein,M. Histone H2B repression causes cell-cycle-specific arrest in yeast: effects on chromosomal segregation, replication, and transcription. Cell 48, 589–597 (1987).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  28. Kamakaka,R. T., Bulger,M. & Kadonaga,J. T. Potentiation of RNA polymerase II transcription by Gal4-VP16 during but not after DNA replication and chromatin assembly. Genes Dev. 7, 1779–1795 (1993).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank P. Willy, D. Fyodorov, X. Huang, J. Butler, M. Levenstein, N. Dhillon, D. Donze, S. Ghidelli, L. Krushel and E. Stone for critical reading of the manuscript; L. Pillus for helpful suggestions and discussions; T. Laverty and G. Rubin for determination of the cytological location of dASF1; K. Collins for generating recombinant dASF1 protein; M. Levenstein for H3 antibodies and purified histones H3–H4; D. Allis for H4 antibodies; and R. Sternglanz, P. Kaufman, D. Shore, D. Gottschling and J. Rine for providing various strains and plasmids used in this study. This work was supported by grants from the NIH to J.T.K., R.K. and R.T.K.; J.K.T. is a Leukemia Society of America Special Fellow.

Author information

Author notes

  1. Shaw-Ree Chen

    Present address: Department of Biochemistry, University of Washington, Seattle, Washington, 98195, USA

Authors and Affiliations

  1. Department of Biology and Center for Molecular Genetics, University of California at San Diego, La Jolla, 92093-0347, California, USA

    Jessica K. Tyler, Shaw-Ree Chen & James T. Kadonaga

  2. National Institutes of Health, NICHD, 18 Library Drive, Bethesda, 20892-5430, Maryland, USA

    Christopher R. Adams

  3. Cold Spring Harbor Laboratory, P.O. Box 100, Cold Spring Harbor, 11724, New York, USA

    Ryuji Kobayashi & Rohinton T. Kamakaka

Authors
  1. Jessica K. Tyler
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Christopher R. Adams
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shaw-Ree Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ryuji Kobayashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Rohinton T. Kamakaka
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. James T. Kadonaga
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jessica K. Tyler.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tyler, J., Adams, C., Chen, SR. et al. The RCAF complex mediates chromatin assembly during DNA replication and repair. Nature 402, 555–560 (1999). https://doi.org/10.1038/990147

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/990147

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing