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J. Biol. Chem., Vol. 275, Issue 45, 35512-35521, November 10, 2000

This Article Full Text Full Text (PDF) All Versions of this Article: 275/45/35512    most recent M003073200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Keller, C. Articles by Krude, T. Search for Related Content PubMed PubMed Citation Articles by Keller, C. Articles by Krude, T. Social Bookmarking                      What's this?

Requirement of Cyclin/Cdk2 and Protein Phosphatase 1 Activity for Chromatin Assembly Factor 1-dependent Chromatin Assembly during DNA Synthesis^*

Christian Keller and Torsten Krude^§

From the Wellcome/Cancer Research Campaign Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, United Kingdom and the Department of Zoology, University of Cambridge, Downing Street, Cambridge CB3 7EJ, United Kingdom

The influence of reversible protein phosphorylation on nucleosome assembly during DNA replication was analyzed in extracts from human cells. Inhibitor studies and add-back experiments indicated requirements of cyclin A/Cdk2, cyclin E/Cdk2, and protein phosphatase type 1 (PP1) activities for nucleosome assembly during DNA synthesis by chromatin assembly factor 1 (CAF-1). The p60 subunit of CAF-1 is a molecular target for reversible phosphorylation by cyclin/Cdk complexes and PP1 during nucleosome assembly and DNA synthesis in vitro. Purified p60 can be directly phosphorylated by purified cyclin A/Cdk2, cyclin E/Cdk2, and cyclin B1/Cdk1, but not by cyclin D/Cdk4 complexes in vitro. Cyclin B1/Cdk1 triggers hyperphosphorylation of p60 in the presence of additional cytosolic factors. CAF-1 containing hyperphosphorylated p60 prepared from mitotic cells is inactive in nucleosome assembly and becomes activated by dephosphorylation in vitro. These data provide functional evidence for a requirement of the cell cycle machinery for nucleosome assembly by CAF-1 during DNA replication.

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