Signaling pathways can control the activation of gene expression programs and thereby regulate cell fate determination. In embryonic stem cells, certain gene expression programs allow the cells to self-renew whereas other programs trigger differentiation into specific cell types as a response to developmental signaling (58
). Elucidation of how temporal changes in transcription programs are coupled to control of cell growth and division is therefore of fundamental importance for our understanding of developmental processes.
Global gene transcription analysis in yeasts and higher eukaryotes has revealed that a significant proportion of the genome is transcribed in a periodic manner during cell cycle progression (5
). Correct periodic regulation is believed to play a critical role in normal cell proliferation, and the genes are often deregulated in different forms of cancer (6
). Depending on the organism, the number of periodically expressed genes ranges from ~400 to more than 1,000 (5
). These include genes with well-established roles in cell cycle progression, such as those encoding cyclins, transcription factors and protein kinases.
A cluster named CLB2
in budding yeast (35 genes) or cluster 1 in fission yeast (87 genes) is periodically expressed and activated at mitosis and repressed in G1
of the next cell cycle (4
). In budding yeast, transcription of the CLB2
cluster is controlled by the forkhead proteins Fkh1 and Fkh2, which cooperate with Mcm1 (a MADS box protein) and the Ndd1 coactivator (27
). In fission yeast, forkhead proteins Sep1 and Fkh2 and the MADS box protein Mbx1 regulate mitotic transcription (12
). Deletion of the sep1
gene results in reduced transcription, whereas overexpression of sep1
induces expression of the same genes. In contrast, deletion of fkh2
causes elevated levels of gene transcription, suggesting a role for this transcription factor in negative regulation of gene transcription (49
). Furthermore, the periodic binding of Sep1 to cluster 1 promoters coincides with gene activation, whereas Fkh2 is bound to those genes when they are repressed, supporting the idea that Sep1 promotes gene expression and Fkh2 represses it (43
Our understanding of how regulation of CLB2
or cluster 1 genes is coordinated with mitotic progression has increased in recent years, revealing the importance of phosphorylation of specific transcription factors by Cdk1 and the Polo kinase and dephosphorylation by the CDC14 phosphatase. In Saccharomyces cerevisiae
, the Cdc28 (Cdk1) kinase phosphorylates both Ndd1 and Fkh2 and these phosphorylation events stimulate direct interactions between Ndd1 and the FHA domain of Fkh2 (18
). The Polo kinase Cdc5 is also temporally recruited to CLB2
gene cluster promoters and phosphorylates Ndd1, which helps to establish a positive feedback loop for CLB2
cluster activation (17
). Similarly, in
, the Polo kinase Plo1 phosphorylates the Mbx1 MADS box protein to positively control gene expression in a feedback loop (43
). In vitro
, Mbx1 is also phosphorylated by Cdk1 and dephosphorylated by the Cdc14-like phosphatase Clp1, with Clp1 apparently having a repressive role in controlling gene expression (42
). Similar to Mbx1, the Fkh2 protein also becomes phosphorylated during mitosis, but the responsible kinase and the function of this modification have not been described.
Albeit Cdk1 kinase is the master regulator of cell cycle progression in eukaryotic cells and the protein is an important regulator of mitotic transcription, recent studies revealed that other factors are also required to establish global periodic transcription patterns in budding yeast (41
). A “transcription network oscillator” has been proposed to operate and couple periodic transcription to the Cdk oscillator (51
), even if the molecular basis for this new oscillator remains to be established.
The Mediator complex is a coregulator of eukaryotic transcription and functions as a bridge between gene-specific transcription regulators and the polymerase II (Pol II) machinery at the promoter (16
). Phylogenetic studies have suggested that the Mediator complex is present in most eukaryotic organisms (10
). Cyclin-dependent kinase 8 (Cdk8) is a conserved Mediator component found in most eukaryotes. In multicellular organisms, Cdk8 has been associated with signaling pathways related to cell differentiation and neuronal development (29
). Early studies identified Cdk8 as a negative regulator of global transcription, but later investigations have modified this conclusion somewhat, since Cdk8 is also required for stimulation of specific genes (24
). Cdk8 has been shown to phosphorylate specific transcription factors to regulate their activity (2
). Recently, Cdk8 has also been connected to cancer development (21
), but a specific role in cell cycle progression has not been demonstrated.
We here demonstrate that Fkh2 and Cdk8 are important regulators of cell cycle progression in fission yeast. We found that Cdk8 controls the levels and activity of Fkh2, which in turn regulates Cdk1 activity via the Wee1 kinase. Our studies reveal a novel pathway whereby cells may couple mitotic gene activation to the control of mitotic entry and suggest that Mediator may be an integrative hub for both transcription regulation and cell cycle progression (33