Two S-phase checkpoint systems, one involving the function of both BIME and Tyr15 phosphorylation of p34(cdc2), inhibit NIMA and prevent premature mitosis

Abstract

We demonstrate that there are at least two S-phase checkpoint mechanisms controlling mitosis in Aspergillus. The first responds to the rate of DNA replication and inhibits mitosis via tyrosine phosphorylation of p34(cdc2). Cells unable to tyrosine phosphorylate p34(cdc2) are therefore viable but are unable to tolerate low levels of hydroxyurea and prematurely enter lethal mitosis when S-phase is slowed. However, if the NIMA mitosis-promoting kinase is inactivated then non-tyrosine-phosphorylated p34(cdc2) cannot promote cells prematurely into mitosis. Lack of tyrosine-phosphorylated p34(cdc2) also cannot promote mitosis, or lethality, if DNA replication is arrested, demonstrating the presence of a second S-phase checkpoint mechanism over mitotic initiation which we show involves the function of BIME. In order to overcome the S-phase arrest checkpoint over mitosis it is necessary both to prevent tyrosine phosphorylation of p34(cdc2) and also to inactivate BIME. Lack of tyrosine phosphorylation of p34(cdc2) allows precocious expression of NIMA during S-phase arrest, and lack of BIME then allows activation of this prematurely expressed NIMA by phosphorylation. The mitosis-promoting NIMA kinase is thus a target for S-phase checkpoint controls.