Chk1 Inhibits E2F6 Repressor Function in Response to Replication Stress to Maintain Cell-Cycle Transcription

Abstract

Background In eukaryotic cells, detection of replication stress results in the activation of the DNA replication checkpoint, a signaling cascade whose central players are the kinases ATR and Chk1. The checkpoint response prevents the accumulation of DNA damage and ensures cell viability by delaying progression into mitosis. However, the role and mechanism of the replication checkpoint transcriptional response in human cells, which is p53-independent, is largely unknown. Results We show that, in response to DNA replication stress, the regular E2F-dependent cell cycle transcriptional program is maintained at high levels and we establish the mechanisms governing such transcriptional upregulation. E2F6, a repressor of E2F-dependent G1/S transcription, replaces the activating E2Fs at promoters to repress transcription in cells progressing into S-phase in unperturbed conditions. Following replication stress, the checkpoint kinase Chk1 phosphorylates E2F6 leading to its dissociation from promoters. This promotes E2F-dependent transcription, which mediates cell survival by preventing DNA damage and cell death. Conclusions This work reveals, for the first time, that the regular cell cycle transcriptional program is part of the DNA replication checkpoint response in human cells and establishes the molecular mechanism involved. We show that maintaining high levels of G1/S cell cycle transcription in response to replication stress contributes to two key functions of the DNA replication checkpoint response, namely preventing genomic instability and cell death. Given the critical role of replication stress in oncogene transformation, a detailed understanding of the molecular mechanisms involved in the checkpoint response will contribute to a better insight into cancer development.

Control of cell cycle transcription during G1 and S phases

“…As a result, they should be able to repress transcription during S phase and the latter part of the cell cycle when pocket proteins are inhibited by Cdk activity 97 . Consistent with this view, E2F6 was recently found to repress G1–S transcription in late S phase 98 (FIG. 2b).…”

“…The DNA replication checkpoint transcriptional response probably maintains G1–S transcription in order to prevent genomic instability 98,101,102,108 (FIG. 3).…”

Tolerance of Deregulated G1/S Transcription Depends on Critical G1/S Regulon Genes to Prevent Catastrophic Genome Instability

“…E2F6, E2F7, and E2F8 are repressors of the E2F transcriptional program and, much like Nrm1 and Yox1 in fission yeast, they are G1/S targets themselves. E2F6 was recently shown to be involved in a negative-feedback loop to turn off transcription during the G1-to-S transition (Bertoli et al, 2013a), and E2F7 and E2F8 have also been implicated (Di Stefano et al, 2003;Lyons et al, 2006). Future research will reveal whether deregulation of G1/S transcription by inactivation of these negative-feedback loops in mammalian cells also creates a specific requirement for genome protection and S-phase cyclin activities.…”

Topology and Control of the Cell-Cycle-Regulated Transcriptional Circuitry

“…Upon entry into the cell cycle, Rb is phosphorylated by a G1 cyclin/CDK, in this case CycD/Cdk4/6, functional analogs of Cln3/Cdk1, leading to activation of the E2F1/DP1 targets. Finally, the G1/S genes are repressed by the binding of the E2F6, a repressive form of E2F, as cells progress into S phase, leading to the dissociation of E2F1/ DP1 from promoters (Bertoli et al 2013a). This precisely parallels the regulation of SBF in budding yeast.…”