Polycomb mediated epigenetic silencing and replication timing at the INK4a/ARF locus during senescence.
BACKGROUND
The INK4/ARF locus encodes three tumor suppressor genes (p15(Ink4b), Arf and p16(Ink4a)) and is frequently inactivated in a large number of human cancers. Mechanisms regulating INK4/ARF expression are not fully characterized.


RESULTS
Here we show that in young proliferating embryonic fibroblasts (MEFs) the Polycomb Repressive Complex 2 (PRC2) member EZH2 together with PRC1 members BMI1 and M33 are strongly expressed and localized at the INK4/ARF regulatory domain (RD) identified as a DNA replication origin. When cells enter senescence the binding to RD of both PRC1 and PRC2 complexes is lost leading to a decreased level of histone H3K27 trimethylation (H3K27me3). This loss is accompanied with an increased expression of the histone demethylase Jmjd3 and with the recruitment of the MLL1 protein, and correlates with the expression of the Ink4a/Arf genes. Moreover, we show that the Polycomb protein BMI1 interacts with CDC6, an essential regulator of DNA replication in eukaryotic cells. Finally, we demonstrate that Polycomb proteins and associated epigenetic marks are crucial for the control of the replication timing of the INK4a/ARF locus during senescence.


CONCLUSIONS
We identified the replication licencing factor CDC6 as a new partner of the Polycomb group member BMI1. Our results suggest that in young cells Polycomb proteins are recruited to the INK4/ARF locus through CDC6 and the resulting silent locus is replicated during late S-phase. Upon senescence, Jmjd3 is overexpressed and the MLL1 protein is recruited to the locus provoking the dissociation of Polycomb from the INK4/ARF locus, its transcriptional activation and its replication during early S-phase. Together, these results provide a unified model that integrates replication, transcription and epigenetics at the INK4/ARF locus.
Agherbi
Hanane
H
Centre d'Immunologie INSERM-CNRS de Marseille Luminy, Marseille, France.
Gaussmann-Wenger
Anne
A
Verthuy
Christophe
C
Chasson
Lionel
L
Serrano
Manuel
M
Djabali
Malek
M
D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D016922:Cellular Senescence; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D042522:DNA Replication Timing; D004622:Embryo, Mammalian / Q000166:cytology; D000071221:Enhancer of Zeste Homolog 2 Protein; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism*; D020868:Gene Silencing; D011495:Histone-Lysine N-Methyltransferase / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D008745:Methylation; D051379:Mice; D008954:Models, Biological; D051788:Myeloid-Lymphoid Leukemia Protein / Q000378:metabolism; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins / Q000378:metabolism; D063150:Polycomb Repressive Complex 1; D063151:Polycomb Repressive Complex 2; D063146:Polycomb-Group Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*
C489421:Bmi1 protein, mouse; C502132:CDC6 protein, mouse; C570426:Cbx2 protein, mouse; D018797:Cell Cycle Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D006657:Histones; D009687:Nuclear Proteins; D063146:Polycomb-Group Proteins; D011518:Proto-Oncogene Proteins; D012097:Repressor Proteins; D051788:Myeloid-Lymphoid Leukemia Protein; D000071221:Enhancer of Zeste Homolog 2 Protein; C498410:Ezh2 protein, mouse; D011495:Histone-Lysine N-Methyltransferase; D063151:Polycomb Repressive Complex 2; D063150:Polycomb Repressive Complex 1