Shibahara, K., Verreault, A., Stillman, B. (July 2000) The N-terminal domains of histones H3 and H4 are not necessary for chromatin assembly factor-1-mediated nucleosome assembly onto replicated DNA in vitro. Proceedings of the National Academy of Sciences of the United States of America, 97 (14). pp. 7766-7771. ISSN 0027-8424
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Abstract
An in vitro reconstitution system for the analysis of replication-coupled nucleosome assembly is described. In this "two-step system," nucleosome assembly is performed in a separate reaction from DNA replication, wherein purified newly replicated DNA remains noncovalently marked for subsequent chromatin assembly factor-1 (CAF-1)-dependent nucleosome assembly. Because the nucleosome assembly is performed separately from the DNA replication step, this system is more versatile and biochemically tractable when compared with nucleosome assembly during simian virus 40 (SV40) DNA replication. The N-terminal domains of histones H3 and H4 play an important but redundant function in nucleosome assembly in the budding yeast, Saccharomyces cerevisiae. It had been proposed that at least one tail of histone H3 or H4 is required for replication-coupled nucleosome assembly. However, we demonstrate that the N-terminal domains of both histone H3 and H4 are dispensable for CAF-l-mediated formation of nucleosome cores onto newly replicated DNA in vitro. CAF-1 and each of its individual subunits stably bound to recombinant (H3.H4)(2) tetramers lacking the N-terminal domains of both H3 and H4. Therefore, the N-terminal tails of histone H3 and H4 that contain the specific acetylation sites are not necessary for CAF-1-dependent nucleosome assembly onto replicated DNA. We suggest that the histone acetylation may be required for a CAF-1 independent pathway or function after deposition, by marking of newly replicated chromatin.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | NEWLY SYNTHESIZED HISTONES newly synthesized histones SACCHAROMYCES-CEREVISIAE Saccharomyces Cerevisiae FACTOR-I Factor I REMODELING FACTOR Remodeling Factor FISSION YEAST Fission Yeast DEPOSITION Deposition ACETYLATION Acetylation INVITRO in vitro CELLS cells RECONSTITUTION reconstitution |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA replication bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > nucleosome |
| CSHL Authors: | |
| Communities: | CSHL labs > Stillman lab |
| Highlight: | Stillman, Bruce W. |
| Depositing User: | CSHL Librarian |
| Date: | July 2000 |
| Date Deposited: | 29 Feb 2012 20:09 |
| Last Modified: | 10 Sep 2019 19:18 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/25053 |
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