Regulation of the Yeast Cen CYde

In S. cerevisiae, a highly regulated and complex network of proteins governs the cell cycle process, although major events are controlled by a single CDK (Cdc28) whose regulation is achieved mainly through the synthesis and degradation of cyclins and inhibitors. At the beginning of G1, Cln3 cyclin is sequestered in the cytoplasm by the Whi3 retention factor (Belli et al., 2001; Gari et al., 2001); later, the nuclear concentration of Cln3

increases in relationship to the total cell mass (Chen et al., 2000). When the level of the Cln3/Cdc28 complex is higher than a certain threshold, it triggers the phosphorylation of Whi5 (an orthologue of the mammalian Rb protein). This event allows the transcription of a second wave of cyclins (Cln1, Cln2, Clb5, and Clb6) by the activation of the SBF and the MBF transcription complex. The activation and accumulation of Cln1,2/Cdc28 engage two processes: the initiation of bud formation and phosphorylation of Sic1, which leads to its ubiquitination by the SCF/Cdc4 complex (Verma et al., 1997) and its destruction by the proteasome. When Sic1 is degraded, Clb5,6/Cdc28 activity rises abruptly and drives cells into S phase.

The progression into G2 phase depends mainly on another wave of cyclin production: Clb1 and Clb2. The Mcm1/SFF (Mcm1/Fkh2/Ndd1) complex is the transcription factor of CLB1 and CLB2 (Althoefer et al., 1995; Jorgensen and Tyers, 2000; Maher et al., 1995). The transcription of CLB1 and CLB2 is autocatalytic because it promotes its own transcription.

The activity of the Clb2/Cdc28 complex is controlled by inhibitory phosphorylation at a conserved tyrosine in the N terminus of Cdc28 by the kinase Swe1 and the reverse phosphatase Mih1. Swe1 does not play an important role during vegetative growth (Amon et al., 1992), but it has been demonstrated that it responds to cytoskeletal perturbations (McNulty and Lew, 2005).

At the end of the cell cycle, when DNA is fully replicated and all chromosomes are aligned on the metaphase plate, Cdc20 is activated (Hwang et al., 1998) and activation of Cdc20/APC promotes degradation of part of the Clb2 protein (Baumer et al., 2000). Later, when the nucleus migrates into the daughter cell, the Cdh1/APC destroys the remaining Clb2 protein. This event represents the exit from mitosis and the start of a new cycle.

In the fission yeast S. pombe, cell cycle progression is also controlled by a single CDK (Cdc2) associated with several cyclins. During G1, Cdc2 is associated with the cyclin Puc1(+) (a close relative of Cln proteins in budding yeast), which regulates the length of G1 (Martin-Castellanos et al., 2000). It is worth noting that puc1 is not the unique cyclin that controls G1 because mitotic cyclins are also playing some role in G1 (see later). Also, Cdc2/Puc1 may be important in downregulating the rum1 Cdk inhibitor (the homologue of S. cerevisiae Sic1) at the end of G1 (Martin-Castellanos et al., 2000).

The onset ofthe S phase in fission yeast is regulated positively at Start by another cyclin: cig2, a B-type cyclin that performs similar functions to Clb5, Clb6 in S. cerevisiae (Mondesert et al., 1996). Similarly to S. cerevisiae, rum1 may transiently inhibit cig2-associated cdc2 activity until the critical cell size required for Start is reached (Martin-Castellanos et al., 1996).

During G2, two B-type cyclins promote progression of the cell cycle: cig1 and cdc13 (cig1 has no obvious homologues in S. cerevisiae, but cdc13

corresponds to the Clbl and the Clb2 mitotic B-type cyclins). Correspondingly, the Cdc2 mitotic complexes are regulated negatively by the Weel protein kinase. Phosphorylation of Cdc2 on tyrosine-15 by Weel is critical for the inhibition of the complex, whereas dephosphorylation of these residues by the Cdc25 phosphatase is the key event governing the initiation of mitosis (Gould and Nurse, 1989; Millar et al., 1991). Therefore, the balance between the levels of Wee1 and Cdc25 sets a threshold for the activity of the cdc13—cdc12 and cig1—cdc13 complexes that determines the correct timing ofM-phase initiation.

At the end of mitosis, Cdc13 and Cig1 are partially degraded, possibly by the Slp1—APC/C complex (the equivalent of Cdc20/APC complex in S. cerevisiae). The remaining activity is necessary for the progression in G1 phase and is fully degraded by the Ste9/Srw1 associated with the APC/C (the homologous complex of Cdh1/APC) (Blanco et al., 2000). Then, the complete destruction of mitotic cyclins occurs during the G1 phase, when cells start a new cycle.

Exposure of cells to osmostress results in rapid activation of a highly conserved family of MAPKs, known as stress-activated protein kinases. Activation of SAPKs elicits the program for cell adaptation, which includes transcriptional and translational regulation of the genome allowing long-term adaptation. Moreover, the same program also produces a short-term adaptation effect, such as glycerol accumulation (Albertyn et al., 1994), or the reestablishment of ionic balance (Proft and Struhl, 2004). The accumulation of compatible solutes, such as glycerol, is a ubiquitous mechanism in cellular osmoregulation. Yeast cells also control glycerol accumulation via the Fps1p-mediated export of glycerol (Tamas et al., 1999).

The SAPK pathways are extremely conserved among eukaryotes and are composed of a tier of three consecutively activated kinases: a MAP kinase kinase kinase (MAPKKK), a MAP kinase kinase (MAPKK), and a MAP kinase (MAPK). Once phosphorylated, the MAPK concentrates in the nucleus, where it can phosphorylate its protein targets on serine/threonine followed by a proline (S/T-P). However, a portion of activated MAP kinase is retained in the cytoplasm to regulate cytosolic events (Reiser et al., 1999). Pathways of MAP kinase are controlled negatively by protein phosphatases acting on two levels: the MAPKK (serine-threonine phosphatases) or only on the MAP kinase (serine-threonine phosphatases and tyrosine phosphatases) (Keyse, 2000). There are different sensing mechanisms upstream of

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