Regulation of Ace2-dependent genes requires components of the PBF complex in Schizosaccharomyces pombe.

The division cycle of unicellular yeasts is completed with the activation of a cell separation program that results in the dissolution of the septum assembled during cytokinesis between the 2 daughter cells, allowing them to become independent entities. Expression of the eng1(+) and agn1(+) genes, encoding the hydrolytic enzymes responsible for septum degradation, is activated at the end of each cell cycle by the transcription factor Ace2. Periodic ace2(+) expression is regulated by the transcriptional complex PBF (PCB Binding Factor), composed of the forkhead-like proteins Sep1 and Fkh2 and the MADS box-like protein Mbx1. In this report, we show that Ace2-dependent genes contain several combinations of motifs for Ace2 and PBF binding in their promoters. Thus, Ace2, Fkh2 and Sep1 were found to bind in vivo to the eng1(+) promoter. Ace2 binding was coincident with maximum level of eng1(+) expression, whereas Fkh2 binding was maximal when mRNA levels were low, supporting the notion that they play opposing roles. In addition, we found that the expression of eng1(+) and agn1(+) was differentially affected by mutations in PBF components. Interestingly, agn1(+) was a major target of Mbx1, since its ectopic expression resulted in the suppression of Mbx1 deletion phenotypes. Our results reveal a complex regulation system through which the transcription factors Ace2, Fkh2, Sep1 and Mbx1 in combination control the expression of the genes involved in separation at the end of the cell division cycle.

Removal of centrosomal PP1 by NIMA kinase unlocks the MPF feedback loop to promote mitotic commitment in S. pombe.

BACKGROUND: Activation of the Cdk1/cyclin B complex, also known as mitosis-promoting factor (MPF), drives commitment to mitosis. Interphase MPF is inhibited through phosphorylation of Cdk1 by Wee1-related kinases. Because Cdc25 phosphatases remove this phosphate, Cdc25 activity is an essential part of the switch that drives cells into mitosis. The generation of a critical "trigger" of active MPF promotes a positive feedback loop that employs Polo kinase to boost Cdc25 activity and inhibit Wee1, thereby ensuring that mitotic commitment is a bistable switch. Mutations in the spindle pole body (SPB) component Cut12 suppress otherwise lethal deficiencies in Cdc25. RESULTS: Cut12 harbors a bipartite protein phosphatase 1 (PP1) docking domain. Mutation of either element alone suppressed the temperature-dependent lethality of cdc25.22, whereas simultaneous ablation of both allowed cells to divide in the complete absence of Cdc25. Late G2 phase phosphorylation between the two elements by MPF and the NIMA kinase Fin1 blocked PP1(Dis2) recruitment, thereby promoting recruitment of Polo to Cut12 and the SPB and elevating global Polo kinase activity throughout the cell. CONCLUSIONS: PP1 recruitment to Cut12 sets a threshold for Polo's feedback-loop activity that locks the cell in interphase until Cdc25 pushes MPF activity through this barrier to initiate mitosis. We propose that events on the SPB (and, by inference, the centrosome) integrate inputs from diverse signaling networks to generate a coherent decision to divide that is appropriate for the particular environmental context of each cell. PP1 recruitment sets one or more critical thresholds for single or multiple local events within this switch.

Ace2p controls the expression of genes required for cell separation in Schizosaccharomyces pombe.

Schizosaccharomyces pombe cells divide by medial fission through contraction of an actomyosin ring and deposition of a multilayered division septum that must be cleaved to release the two daughter cells. Here we describe the identification of seven genes (adg1(+), adg2(+), adg3(+), cfh4(+), agn1(+), eng1(+), and mid2(+)) whose expression is induced by the transcription factor Ace2p. The expression of all of these genes varied during the cell cycle, maximum transcription being observed during septation. At least three of these proteins (Eng1p, Agn1p, and Cfh4p) localize to a ring-like structure that surrounds the septum region during cell separation. Deletion of the previously uncharacterized genes was not lethal to the cells, but produced defects or delays in cell separation to different extents. Electron microscopic observation of mutant cells indicated that the most severe defect is found in eng1Delta agn1Delta cells, lacking the Eng1p endo-beta-1,3-glucanase and the Agn1p endo-alpha-glucanase. The phenotype of this mutant closely resembled that of ace2Delta mutants, forming branched chains of cells. This suggests that these two proteins are the main activities required for cell separation to be completed.