Abstract B14: Feedback regulation between atypical E2Fs and APC/CCdh1 coordinates cell cycle progression.

Abstract

E2F transcription factors control the oscillating expression pattern of multiple target genes during the cell cycle. Activator E2Fs, E2F1-3, induce an upswing of E2F targets, which is essential for the G1-to-S phase transition, whereas atypical E2Fs, E2F7 and E2F8, mediate a downswing of the same targets during late S, G2, and M phase. Expression of atypical E2Fs is induced by E2F1-3, but it is unknown how atypical E2Fs are inactivated in a timely manner. Using molecular assays, time lapse microscopy, and flow cytometry we now demonstrate that E2F7 and E2F8 are substrates of the Anaphase Promoting Cyclosome/Complex (APC/C). Removal of CDH1, or mutating the CDH1-interacting KEN boxes, stabilized E2F7/8 from anaphase onwards and during G1. Expressing KEN-mutant E2F7 during G1 severely impairs S-phase entry and eventually results in cell death. Furthermore, we show that E2F8, but not E2F7, interacts also with APC/CCdc20. Importantly, atypical E2Fs can activate the APC/CCdh1 by repressing its inhibitors Emi1, cyclin A, and cyclin E. In conclusion, we uncovered a feedback loop between atypical E2Fs and APC/CCdh1, which ensures balanced expression of cell cycle genes and normal cell cycle progression. Citation Format: Michiel Boekhout, Ruixue Yuan, Annelotte P. Wondergem, Hendrika A. Segeren, Elsbeth A. van Liere, Nesibu Awol, Imke Jansen, Rob MF Wolthuis, Alain de Bruin, Bart Westendorp. Feedback regulation between atypical E2Fs and APC/CCdh1 coordinates cell cycle progression.. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Cancer Cell Cycle - Tumor Progression and Therapeutic Response; Feb 28-Mar 2, 2016; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(11_Suppl):Abstract nr B14.

Chk1 and 14‐3‐3 proteins inhibit atypical E2Fs to prevent a permanent cell cycle arrest

“…Here, we show a novel mechanism for how the repressor activity of atypical E2Fs can be inhibited through direct phosphorylation by Chk1 which provides a docking station for 14‐3‐3 proteins to interact with E2F7 and E2F8 (Fig D). Previous studies have demonstrated that increased repressor activity of atypical E2Fs downregulates the expression of a large number of cell cycle genes resulting in a severe cell cycle arrest accompanied by increased DNA damage and apoptotic events (de Bruin et al , ; Maiti et al , ; Westendorp et al , ; Boekhout et al , ). Our findings provide strong evidence for a regulatory mechanism that suppresses the atypical E2F activity in order to avoid detrimental effects on cell cycle progression and cell survival under DNA‐damaging conditions.…”

“…E2F7 and E2F8 levels peak during S phase, to mediate the downswing of many oscillating target genes involved in DNA replication, metabolism, and DNA repair. We previously found that E2F7 and E2F8 are subjected to degradation via APC/C Cdh1 during G1 phase, to allow a time window for upregulation of E2F target genes by activator E2Fs, and subsequent S‐phase entry (Boekhout et al , ). Meanwhile, ectopic expression of E2F7 and E2F8 is sufficient to stall S‐phase progression, indicating that their activity must be carefully regulated to ensure unperturbed proliferation (Westendorp et al , ; Boekhout et al , ).…”

“…We previously found that E2F7 and E2F8 are subjected to degradation via APC/C Cdh1 during G1 phase, to allow a time window for upregulation of E2F target genes by activator E2Fs, and subsequent S-phase entry (Boekhout et al, 2016). Meanwhile, ectopic expression of E2F7 and E2F8 is sufficient to stall S-phase progression, indicating that their activity must be carefully regulated to ensure unperturbed proliferation (Westendorp et al, 2012;Boekhout et al, 2016). Interestingly, there is accumulating evidence suggesting that atypical E2Fs also play an important role in the DNA damage response.…”

Cyclin F‐dependent degradation of E2F7 is critical for DNA repair and G2‐phase progression

“…Ectopic expression of atypical E2Fs leads to downregulation of these target genes accompanied by a permanent Sphase arrest and severe DNA damage (Westendorp et al, 2012;Yuan et al, 2018). Recently, we have shown that APC/C Cdh1 targets E2F7 and E2F8 for degradation during the G1 phase of the cell cycle and that inhibition of the APC/C Cdh1 -mediated degradation of E2F7 and E2F8 impairs S-phase entry, eventually resulting in cell death (Boekhout et al, 2016). As such, activity of E2F7 and E2F8 must be tightly regulated during the cell cycle and in response to DNA damage.…”

“…However, the regulatory mechanism behind the downregulation in G2 is unknown (Boekhout et al, 2016). However, the regulatory mechanism behind the downregulation in G2 is unknown (Boekhout et al, 2016).…”