Agherbi Hanane, Gaussmann-Wenger Anne, Verthuy Christophe, Chasson Lionel, Serrano Manuel, Djabali Malek
Centre d'Immunologie INSERM-CNRS de Marseille Luminy, Marseille, France.
PLoS One. 2009 May 20;4(5):e5622. doi: 10.1371/journal.pone.0005622.
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.
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.
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.
INK4/ARF基因座编码三种肿瘤抑制基因（p15(Ink4b)、Arf和p16(Ink4a)），在大量人类癌症中经常失活。调节INK4/ARF表达的机制尚未完全阐明。
我们发现，在年轻的增殖胚胎成纤维细胞（MEF）中，多梳抑制复合物2（PRC2）成员EZH2与PRC1成员BMI1和M33强烈表达，并定位于被确定为DNA复制起点的INK4/ARF调控域（RD）。当细胞进入衰老状态时，PRC1和PRC2复合物与RD的结合丧失，导致组蛋白H3K27三甲基化（H3K27me3）水平降低。这种丧失伴随着组蛋白去甲基化酶Jmjd3表达的增加以及MLL1蛋白的募集，并与Ink4a/Arf基因的表达相关。此外，我们表明多梳蛋白BMI1与CDC6相互作用，CDC6是真核细胞中DNA复制的关键调节因子。最后，我们证明多梳蛋白和相关的表观遗传标记对于衰老过程中INK4a/ARF基因座复制时间的控制至关重要。
我们确定复制许可因子CDC6是多梳蛋白家族成员BMI1的新伙伴。我们的结果表明，在年轻细胞中，多梳蛋白通过CDC6被募集到INK4/ARF基因座，由此产生的沉默基因座在S期后期复制。衰老时，Jmjd3过表达，MLL1蛋白被募集到该基因座，促使多梳蛋白从INK4/ARF基因座解离，其转录激活并在S期早期复制。总之，这些结果提供了一个统一的模型，该模型整合了INK4/ARF基因座处的复制、转录和表观遗传学。
