Proteolysis mediated by the anaphase-promoting complex (APC) triggers chromosome segregation and exit from mitosis, yet its regulation is poorly understood. The conserved Cdc20 and Cdh1 proteins were identified as limiting, substrate-specific activators of APC-dependent proteolysis. CDC20 was required for the degradation of the APC substrate Pds1 but not for that of other APC substrates, such as Clb2 and Ase1. Conversely, cdh1Delta mutants were impaired in the degradation of Ase1 and Clb2 but not in that of Pds1. Overexpression of either CDC20 or CDH1 was sufficient to induce APC-dependent proteolysis of the appropriate target in stages of the cell cycle in which substrates are normally stable.

Exit from mitosis requires the inactivation of mitotic cyclin-dependent kinases (CDKs) by an unknown mechanism. We show that the Cdc14 phosphatase triggers mitotic exit by three parallel mechanisms, each of which inhibits Cdk activity. Cdc14 dephosphorylates Sic1, a Cdk inhibitor, and Swi5, a transcription factor for SIC1, and induces degradation of mitotic cyclins, likely by dephosphorylating the activator of mitotic cyclin degradation, Cdh1/Hct1. Feedback between these pathways may lead to precipitous collapse of mitotic CDK activity and help coordinate exit from mitosis.

There are remarkable differences in the regulation of Cdc20 and Cdh1/Hct1. Furthermore, the APC activator Cdc20 is itself a substrate of the Cdc27 have a role in the degradation of Cdc20 during S Phase and early mitosis that is not mediated by its destruction box.

No abstract

Meiotic cohesin serves in sister chromatid linkage and DNA repair until its subunit Rec8 is cleaved by separase. Separase is activated when its inhibitor, securin, is polyubiquitinated by the Cdc20 regulated anaphase-promoting complex (APC(Cdc20)) and consequently degraded. Differently regulated APCs (APC(Cdh1), APC(Ama1)) have not been implicated in securin degradation at meiosis I. We show that Mnd2, a factor known to associate with APC components, prevents premature securin degradation in meiosis by APC(Ama1). mnd2Delta cells lack linear chromosome axes and exhibit precocious sister chromatid separation, but deletion of AMA1 suppresses these defects. Besides securin, Sgo1, a protein essential for protection of centromeric cohesion during anaphase I, is also destabilized in mnd2delta cells. Mnd2's disappearance prior to anaphase II may activate APC(Ama1). Human oocytes may spend many years in meiotic prophase before maturation. Inhibitors of meiotic APC variants could prevent loss of chiasmata also in these cells, thereby guarding against aberrant chromosome segregation.

On solid growth media with limiting nitrogen source, diploid budding-yeast cells differentiate from the yeast form to a filamentous, adhesive, and invasive form. Genomic profiles of mRNA levels in Saccharomyces cerevisiae yeast-form and filamentous-form cells were compared. Disparate data types, including genes implicated by expression change, filamentation genes known previously through a phenotype, protein-protein interaction data, and protein-metabolite interaction data were integrated as the nodes and edges of a filamentation-network graph. Application of a network-clustering method revealed 47 clusters in the data. The correspondence of the clusters to modules is supported by significant coordinated expression change among cluster co-member genes, and the quantitative identification of collective functions controlling cell properties. The modular abstraction of the filamentation network enables the association of filamentous-form cell properties with the activation or repression of specific biological processes, and suggests hypotheses. A module-derived hypothesis was tested. It was found that the 26S proteasome regulates filamentous-form growth.

Molecular interactions provide paths for information flows. Genetic interactions reveal active information flows and reflect their functional consequences. We integrated these complementary data types to model the transcription network controlling cell differentiation in yeast. Genetic interactions were inferred from linear decomposition of gene expression data and were used to direct the construction of a molecular interaction network mediating these genetic effects. This network included both known and novel regulatory influences, and predicted genetic interactions. For corresponding combinations of mutations, the network model predicted quantitative gene expression profiles and precise phenotypic effects. Multiple predictions were tested and verified.

A key aspect of meiotic chromosome segregation is that cohesin, the protein complex that holds sister chromatids together, dissociates from chromosome arms during meiosis I and from centromeric regions during meiosis II. The budding yeast protein Spo13 plays a key role in preventing centromeric cohesin from being lost during meiosis I. We have determined the molecular basis for the metaphase arrest obtained when SPO13 is overexpressed during the mitotic cell cycle. Overexpression of SPO13 inhibits anaphase onset by at least two mechanisms. First, Spo13 causes a transient delay in degradation of the anaphase inhibitor Pds1. Second, Spo13 inhibits cleavage of the cohesin subunit Scc1/Mcd1 or its meiosis-specific homolog, Rec8, by the separase Esp1. The finding that Spo13 did not prevent cleavage of another Esp1 substrate, Slk19, suggests that overexpression of SPO13 is sufficient to prevent cohesin cleavage by protecting specific substrates from separase activity.[Key Words: Spo13; separin; cohesin; sister-chromatid; segregation; meiosis; S. cerevisiae] To generate two daughter cells with the exact same complement of chromosomes, it is critical that sister chromatids remain tightly associated prior to chromosome segregation and promptly dissociate as nuclear division commences during anaphase. Essential to this process is the multisubunit protein complex known as cohesin, which provides the physical linkage between sister chromatids. In budding yeast, the cohesin complex consists of at least four subunits, Smc1, Smc3, Scc1/ Mcd1, and Scc3 (for review, see Uhlmann 2001).Cohesion between sister chromatids is established during DNA replication (Uhlmann and Nasmyth 1998) and is maintained until the onset of anaphase, when chromosome segregation is initiated by the loss of cohesion. At the onset of anaphase, the cohesin subunit Scc1/ Mcd1 is cleaved by a protease known as separase (Esp1 in budding yeast), allowing sister chromatids to separate (for review, see Nasmyth et al. 2000). The activity of Esp1 is controlled by the anaphase inhibitor securin (Pds1 in budding yeast), which binds to and inhibits Esp1. A ubiquitin ligase called the Anaphase Promoting Complex or Cyclosome (APC/C), in association with the WD40 repeat protein Cdc20, targets Pds1 for degradation at the metaphase-anaphase transition (for review, see Peters 1998;Cohen-Fix 2001), thereby freeing Esp1 to initiate chromosome segregation and mitotic exit.In contrast to mitosis, where cohesion is lost along the entire length of the chromosome by the onset of anaphase, sister-chromatid cohesion is lost in a stepwise manner during meiosis (for review, see Miyazaki and Orr-Weaver 1994; Lee and Orr-Weaver 2001). Sister-chromatid cohesion is lost along chromosome arms at the onset of anaphase I, but centromeric cohesion is retained until the onset of anaphase II. Loss of cohesion on chromosome arms is required for homologs to segregate from each other , whereas retention of centromeric cohesion between sister chromatids beyond anaphase I and its subsequent loss a...