E2F transcription factors are implicated in diverse cellular functions. The founding member, E2F-1, is endowed with contradictory activities, being able to promote cell-cycle progression and induce apoptosis. However, the mechanisms that underlie the opposing outcomes of E2F-1 activation remain largely unknown. We show here that E2F-1 is directly methylated by PRMT5 (protein arginine methyltransferase 5), and that arginine methylation is responsible for regulating its biochemical and functional properties, which impacts on E2F-1-dependent growth control. Thus, depleting PRMT5 causes increased E2F-1 protein levels, which coincides with decreased growth rate and associated apoptosis. Arginine methylation influences E2F-1 protein stability, and the enhanced transcription of a variety of downstream target genes reflects increased E2F-1 DNA-binding activity. Importantly, E2F-1 is methylated in tumour cells, and a reduced level of methylation is evident under DNA damage conditions that allow E2F-1 stabilization and give rise to apoptosis. Significantly, in a subgroup of colorectal cancer, high levels of PRMT5 frequently coincide with low levels of E2F-1 and reflect a poor clinical outcome. Our results establish that arginine methylation regulates the biological activity of E2F-1 activity, and raise the possibility that arginine methylation contributes to tumourigenesis by influencing the E2F pathway.

Summary The mechanisms that underlie and dictate the different biological outcomes of E2F-1 activity have yet to be elucidated. We describe the residue-specific methylation of E2F-1 by the asymmetric dimethylating protein arginine methyltransferase (PRMT) 1 and symmetric dimethylating PRMT5, and relate the marks to different functional consequences of E2F-1 activity. Methylation by PRMT1 hinders methylation by PRMT5, which augments E2F-1-dependent apoptosis, whereas PRMT5-dependent methylation favours proliferation by antagonising methylation by PRMT1. The ability of E2F-1 to prompt apoptosis in DNA damaged cells coincides with enhanced PRMT1 methylation. In contrast, cyclin A binding to E2F-1 impedes PRMT1 methylation and augments PRMT5 methylation, thus ensuring that E2F-1 is locked into its cell cycle progression mode. The Tudor domain protein p100-TSN reads the symmetric methylation mark, and binding of p100-TSN down-regulates E2F-1 apoptotic activity. Our results define an exquisite level of precision in the reader-writer interplay that governs the biological outcome of E2F-1 activity.

Deregulation of the pRB/E2F pathway, which occurs frequently in human malignancy, is often associated with inappropriate proliferation and/or apoptosis. While the role of E2F1 in apoptosis induction has been well-established, it remains unclear how this pro-apoptotic activity is regulated in cancer. Here we describe EZH2, an oncogenic polycomb histone methyltransferase and an E2F1 target, as an important regulator of E2F1-dependent apoptosis. We show that E2F1 induces EZH2 expression, which in turn antagonizes the induction of E2F1 pro-apoptotic target Bim expression. RNAi-mediated gene depletion of EZH2 enhances E2F1-dependent Bim expression, thereby promoting the pro-apoptotic activity of E2F1. Hence, the concomitant induction of EZH2 and Bim by E2F1 constitutes a fail-safe mechanism to allow tumor cells with aberrant E2F1 activity to evade apoptosis. These findings reveal a novel mechanism by which the apoptotic activity of E2F1 is restrained in human cancer and also provide the first evidence that EZH2 directly regulates apoptotic process in cancer cells.