A Novel Zinc Finger Protein Zfp277 Mediates Transcriptional Repression of the Ink4a/Arf Locus through Polycomb Repressive Complex 1

Masamitsu Negishi,Shinobu Mochizuki,Kristian Helin,Haruhiko Koseki,Atsushi Iwama,Atsunori Saraya

doi:10.1371/journal.pone.0012373

Masamitsu Negishi, Shinobu Mochizuki + Show 4 more

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https://doi.org/10.1371/journal.pone.0012373

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BackgroundPolycomb group (PcG) proteins play a crucial role in cellular senescence as key transcriptional regulators of the Ink4a/Arf tumor suppressor gene locus. However, how PcG complexes target and contribute to stable gene silencing of the Ink4a/Arf locus remains little understood.Methodology/Principal FindingsWe examined the function of Zinc finger domain-containing protein 277 (Zfp277), a novel zinc finger protein that interacts with the PcG protein Bmi1. Zfp277 binds to the Ink4a/Arf locus in a Bmi1-independent manner and interacts with polycomb repressor complex (PRC) 1 through direct interaction with Bmi1. Loss of Zfp277 in mouse embryonic fibroblasts (MEFs) caused dissociation of PcG proteins from the Ink4a/Arf locus, resulting in premature senescence associated with derepressed p16Ink4a and p19Arf expression. Levels of both Zfp277 and PcG proteins inversely correlated with those of reactive oxygen species (ROS) in senescing MEFs, but the treatment of Zfp277 −/− MEFs with an antioxidant restored the binding of PRC2 but not PRC1 to the Ink4a/Arf locus. Notably, forced expression of Bmi1 in Zfp277 −/− MEFs did not restore the binding of Bmi1 to the Ink4a/Arf locus and failed to bypass cellular senescence. A Zfp277 mutant that could not bind Bmi1 did not rescue Zfp277 −/− MEFs from premature senescence.Conclusions/SignificanceOur findings implicate Zfp277 in the transcriptional regulation of the Ink4a/Arf locus and suggest that the interaction of Zfp277 with Bmi1 is essential for the recruitment of PRC1 to the Ink4a/Arf locus. Our findings also highlight dynamic regulation of both Zfp277 and PcG proteins by the oxidative stress pathways.

Highlights

Cellular senescence is a fundamental cellular program triggered after a finite number of cell divisions or more rapidly in response to acute stress
Our study demonstrated that Zinc finger domain-containing protein 277 (Zfp277) is critical for the recruitment and/or stable accumulation of Bmi1-containing PRC1 at the Ink4a/Arf locus
These findings define Zfp277 as a platform for PRC1 and provide a novel mechanism for PRC1 to lodge at its target loci

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Introduction

Cellular senescence is a fundamental cellular program triggered after a finite number of cell divisions (replicative senescence) or more rapidly in response to acute stress (premature senescence). It functions as a cell-autonomous safeguard against severe genomic instability and carcinogenesis and contributes to aging in mammals as well This irreversible cytostatic state can be triggered by multiple mechanisms, including telomere shortening, oxidative stress, cancer-causing genetic alterations, and DNA damage [1,2]. P16Ink4a protein directly inhibits the activities of the cyclin D-dependent kinases, Cdk and Cdk, and maintains the pRb-E2f pathway in an anti-proliferative state, while p19Arf controls the stabilization and activation of p53 through binding to Mdm, and promotes cellular senescence in primary fibroblasts [5] Both p16Ink4a and p19Arf play an important role in cellular mortality [6,7,8], there seem to be species-specific differences in their function. How PcG complexes target and contribute to stable gene silencing of the Ink4a/Arf locus remains little understood

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