Nucleosome assembly activity and intracellular localization of human CAF-1 changes during the cell division cycle.
We characterized changes of nucleosome assembly activity, intracellular localization, and reversible phosphorylation of the human chromatin assembly factor CAF-1 during the somatic cell division cycle. HeLa cells were synchronized in the G1, S, G2, and M phases of the cell cycle. All three subunits of human CAF-1 (p150, p60, and p48) are present during the entire cell cycle. In interphase, p150 and p60 are bound to the nucleus, but they predominantly dissociate from chromatin during mitosis. During S phase, p150 and p60 are concentrated at sites of intranuclear DNA replication. Only a fraction of total p48 is associated with p150 and p60, and the majority is present in other high molecular weight complexes. The other nucleosome assembly protein, NAP-1, is predominantly cytosolic throughout the cell cycle. Human CAF-1 efficiently mediates nucleosome assembly during complementary DNA strand synthesis in G1, S, and G2 phase cytosolic extracts. Active CAF-1 can be isolated as a 6.5 S complex from G1, S, and G2 phase nuclei. In contrast, CAF-1 isolated from mitotic cytosol does not support nucleosome assembly during DNA synthesis. In mitosis, the p60 subunit of inactive CAF-1 is hyperphosphorylated, whereas active CAF-1 in interphase contains hypophosphorylated and/or phosphorylated forms of p60.
Marheineke
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K
Wellcome/Cancer Research Campaign Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, United Kingdom and the Department of Zoology, University of Cambridge, United Kingdom.
Krude
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T
D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins; D002843:Chromatin / Q000378:metabolism; D056504:Chromatin Assembly Factor-1; D002868:Chromosomal Proteins, Non-Histone; D004261:DNA Replication / Q000502:physiology; D004268:DNA-Binding Proteins / Q000737:chemistry*; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D008836:Micrococcal Nuclease / Q000378:metabolism; D008938:Mitosis / Q000502:physiology; D009687:Nuclear Proteins / Q000032:analysis; D056490:Nucleosome Assembly Protein 1; D009707:Nucleosomes / Q000378:metabolism*; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism; D014157:Transcription Factors
C533593:CHAF1B protein, human; C489780:CNOT8 protein, human; D018797:Cell Cycle Proteins; D002843:Chromatin; D056504:Chromatin Assembly Factor-1; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C533557:NAP1L1 protein, human; D009687:Nuclear Proteins; D056490:Nucleosome Assembly Protein 1; D009707:Nucleosomes; D011506:Proteins; D014157:Transcription Factors; D008836:Micrococcal Nuclease