Abstract
Fission yeast Atf1 is a member of the ATF/CREB basic leucine zipper (bZIP) family of transcription factors with strong homology to mammalian ATF2. Atf1 regulates transcription in response to stress stimuli and also plays a role in controlling heterochromatin formation and recombination. However, its DNA binding independent role is poorly studied. Here, we report that Atf1 has a distinct role in regulating the anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase. We have identified atf1(+) as a dose-dependent suppressor of apc5-1, a mutation causing mitotic arrest. Remarkably, the suppression is not dependent upon the bZIP domain and is therefore independent of the ability of Atf1 to bind DNA. Interestingly, Atf1 physically binds the APC/C in vivo. Furthermore, we show that addition of purified Atf1 proteins into a cell-free system stimulates ubiquitylation of cyclin B and securin by the APC/C. These results reveal a novel role for Atf1 in cell cycle control through protein-protein interaction.
Highlights
The anaphase-promoting complex/cyclosome (APC/C)2 is a large (1.5 MDa) multisubunit E3 ubiquitin ligase, which has essential functions in key events during the cell cycle, such as chromosome segregation and mitotic exit by degrading securin/Cut2/Pds1 and cyclin B/Cdc13/Clb2, respectively [2,3,4]
Apart from the well established role of the C-terminal WD40 domain in substrate recognition, we have recently shown that the interaction between the C-box of the Fizzy family of activators and the APC/C stimulates the activity, thereby providing an additional role in triggering ubiquitylation [13]
Our results indicate that the transcription factor Atf1 functions through its ability to bind DNA, and has a role in activating the APC/C ubiquitin ligase
Summary
HϪ leu ura ade hϪ apc5ϩ-3HA::kanMX leu hϪ apc5-1-3HA::kanMX leu ura ade hϪ apc5-1-3HA::kanMX ste11::ura4ϩ leu ura ade hϪ apc5-1-3HA::kanMX atf1::ura4ϩ leu ura ade hϪ atf1::ura4ϩ leu ura ade hϪ ste11::ura4ϩ leu ura ade hϪ atf1-bZIP⌬-2Pk::hphMX leu ura ade hϪ apc5-96::kanMX leu ura ade hϪ cut4-18::kanMX leu ura ade hϪ nuc2-72::kanMX leu ura ade hϪ cut100 leu hϪ cut665 leu hϪ apc10-433::kanMX leu ura ade hϪ lid1-TAP::kanMX mts leu ura h90 leu ura ade h90 atf1-bZIP⌬-2Pk::hphMX leu ura ade h90 atf1::ura4ϩ leu ura. Our results indicate that the transcription factor Atf functions through its ability to bind DNA, and has a role in activating the APC/C ubiquitin ligase
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