The Transcription Factor Atf1 Binds and Activates the APC/C Ubiquitin Ligase in Fission Yeast

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 anaphasepromoting 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.Ubiquitylation is a post-translational modification that occurs through the action of an enzymatic cascade consisting of three enzymes E1, E2, and E3, and typically controls the proteolysis, localization, or activity of a protein (1). It is a tightly regulated, highly specific and temporally controlled process, and thus plays an important role in many processes such as the cell cycle, signal transduction, transcription, DNA repair, development, and regulation of the immune response.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 (2-4). The APC/C belongs to the RING finger family of E3 ubiquitin ligases, which include Ubr1, c-Cbl, and Skp1-Cullin 1-F-box (SCF) complex (5), but unlike other members of this family, it is exceptionally large and complex, consisting of at least 11 conserved subunits. The APC/C appears to consist of two separable subcomplexes which associate independently with the scaffold subunit, Apc1/Cut4 (6). The first subcomplex includes a RING finger subunit Apc11 and a cullin domain subunit Apc2. This subcomplex is capable of ubiquitylating proteins to some extent, but lacks substrate specificity (7-9). The specificity seems to rely on the second subcomplex, consisting of three subunits Apc3/Nuc2/Cdc27, Apc6/Cut9/Cdc16, and Apc8/Cut23/Cdc23, which contain a tetratricopeptide repeat (TPR) domain. A member of the Fizzy family of APC/C activators, Cdh1, has been shown to bind Apc3/Cdc27 via its C-terminal IR motifs and recruit a substrate to the APC/C (10 -12). However, the exact roles of the remaining subunits and other TPR subunits, such as Apc5 and Apc7, are unknown.The activity of the APC/C is regulated in part by the association of the Fizzy family of WD40-containing activator proteins (2-4). There are two major activators during the cell cycle, Fizzy/Slp1/Cdc2...

The Saccharomyces cerevisiae Anaphase-Promoting Complex Interacts with Multiple Histone-Modifying Enzymes To Regulate Cell Cycle Progression

“…It seems that the link between chromatin and the APC in budding and fission yeasts is conserved, but the mechanisms involved may have diverged. It is not clear whether the APC influences chromatin modifications and gene expression in fission yeast, but a recent demonstration that the Atf1 transcription factor genetically and physically interacted with the fission yeast Apc5 suggests that the link between the APC and transcription is indeed conserved (45). Additional work will be required to work out the details and differences between the two yeast species and the relevance of these differences as far as human biology is concerned.…”

Ubiquitin and Proteasomes in Transcription

“…This concept is at the heart of our model of how Ub-mediated proteolysis controls TAs (Figure 6), but examples also exist in which an E3 can directly interpret the CTD code on Pol II (98) or in which the CTD is read together with other structural changes in polymerase that only occur when the molecule is active (99). Second, components of the UPS can be activated specifically within a transcriptional context (Figure 8 b ) (100, 101). One of the best examples of this is the Ub ligase TIF1 γ , which is activated by modified histone tails (102), allowing it to select only its chromatin-bound pool of substrate (Smad4) for ubiquitylation.…”

The Basic Leucine Zipper Domain Transcription Factor Atf1 Directly Controls Cdc13 Expression and Regulates Mitotic Entry Independently of Wee1 and Cdc25 in Schizosaccharomyces pombe

“…Atf1 is also necessary for accumulation of cells in G 1 after nitrogen starvation (1). It has been shown to be important for degradation of the mitotic cyclin Cdc13 by activating the APC/cyclosome (APC/C) ubiquitin ligase (7).…”