Phosphorylation and inactivation of the mitotic inhibitor Weel by the nim1/cdr1 kinase

Abstract

The G2-M phase transition in eukaryotes is regulated by the synergistic and opposing activities of a cascade of distinct protein kinases and phosphatases. This cascade converges on Cdc2, a serine/threonine protein kinase required for entry into mitosis (reviewed in ref. 1). In the fission yeast Schizosaccharomyces pombe, inactivation of the Cdc2/cyclin B complex is achieved by phosphorylation of tyrosine 15 by Wee1 (refs 2,3). The action of the Wee1 kinase is opposed by the action of the Cdc25 phosphatase, which dephosphorylates Cdc2 on tyrosine 15, thereby activating the Cdc2/cyclin B complex. Much less is known about the regulatory signals upstream of cdc25 and wee1. Genetics indicate that the mitotic inducer nim1/cdr1 acts upstream of wee1, possibly as a negative regulator of wee1 (refs 10, 11). To characterize the nim1/cdr1 protein (Nim1), we have overproduced it in both bacterial and baculoviral expression systems. We report that Nim1 possesses intrinsic serine-kinase, threonine-kinase and tyrosine-kinase activities. Co-expression of the Nim1 and Wee1 kinases in insect cells results in the phosphorylation of Wee1 and therefore a shift in its electrophoretic mobility on SDS-polyacrylamide gels. When Wee1 is phosphorylated, its ability to phosphorylate Cdc2 on tyrosine 15 is inhibited; treatment with phosphatase restores this kinase activity. Furthermore, purified bacterially produced Nim1 kinase directly phosphorylates and inactivates Wee1 in vitro. These results show that nim1/cdr1 functions as a positive regulator of mitosis by directly phosphorylating and inactivating the mitotic inhibitor Wee1.

Turning off the G2 DNA damage checkpoint

“…Two kinases in fission yeast, Cdr1 and Cdr2, were originally identified by the "changed division response"; when nitrogen becomes limiting, Cdr1 and Cdr2 inactivate Wee1 by phosphorylation of the catalytic domain, advancing the point of mitotic entry [39][40][41][42][43]. Under cycling conditions, deletion of cdr1, cdr2 or both has only a minor impact on cell cycle progression.…”

Yeast polo-like kinases: functionally conserved multitask mitotic regulators

“…In fission yeast and higher eucaryotes, Cdc2 is phosphorylated at Tyr15 and negatively regulated by Wee1 (Russell and Nurse, 1987;Parker and Piwnica-Worms, 1992;McGowan and Russell, 1993), an event that is reversed by the activity of Cdc25C phosphatase (Draetta and Eckstein, 1997;Nilsson and Hoffmann, 2000). Wee1, in turn, is phosphorylated and negatively regulated by a pair of related kinases, Nim1/Cdr1 (Russell and Nurse, 1987;Coleman et al, 1993;Parker et al, 1993) and Cdr2 (Breeding et al, 1998;Kanoh and Russell, 1998). In budding yeast, Clb (cyclin B homolog)-bound Cdc28 is phosphorylated at the equivalent Tyr19 and negatively regulated by the Wee1 ortholog Swe1 (Booher et al, 1993), which is antagonized by the action of the Cdc25 ortholog Mih1 (Russell et al, 1989;Sia et al, 1996).…”

Radiation-induced apoptosis of tumor cells is facilitated by inhibition of the interaction between Survivin and Smac/DIABLO

“…[32], Wee1 [33], Rb [34] and Sld1 [35] were reported to induce deceleration and acceleration in the migration in the gel, respectively. Next, we investigated the effects of (Fig.…”