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From their transcriptional birth to their degradation, cellular mRNAs are coated with proteins in messenger ribonucleoprotein (mRNP) complexes. The mRNP composition controls every aspect of the life of the mRNA, from pre-mRNA processing to mRNA localization, translation and turnover. Transitions

mRNA decapping is a central step in eukaryotic mRNA decay that simultaneously shuts down translation initiation and activates mRNA degradation. A major complex responsible for decapping consists of the decapping enzyme Dcp2 in association with decapping enhancers. An important question is how the activity and accumulation of Dcp2 are regulated at the cellular level to ensure the specificity and fidelity of the Dcp2 decapping complex. Here, we show that human Dcp2 levels and activity are controlled by a competition between decapping complex assembly and Dcp2 degradation. This is mediated by a regulatory domain in the Dcp2 C terminus, which, on the one hand, promotes Dcp2 activation via decapping complex formation mediated by the decapping enhancer Hedls and, on the other hand, targets Dcp2 for ubiquitin-mediated proteasomal degradation in the absence of Hedls association. This competition between Dcp2 activation and degradation restricts the accumulation and activity of uncomplexed Dcp2, which may be important for preventing uncontrolled decapping or for regulating Dcp2 levels and activity according to cellular needs.

Genome stability depends on faithful chromosome segregation, which relies on maintenance of chromatid cohesion during S phase. In eukaryotes, Pds1/securin is the only known inhibitor that can prevent loss of cohesion. However, pds1Δ yeast cells and securin-null mice are viable. We sought to identify redundant mechanisms that promote cohesion within S phase in the absence of Pds1 and found that cells lacking the S-phase cyclins Clb5 and Clb6 have a cohesion defect under conditions of replication stress. Similar to the phenotype of pds1Δ cells, loss of cohesion in cells lacking Clb5 and Clb6 is dependent on Esp1. However, Pds1 phosphorylation by Cdk-cyclin is not required for cohesion. Moreover, cells lacking Clb5, Clb6, and Pds1 are inviable and lose cohesion during an unperturbed S phase, indicating that Pds1 and specific B-type cyclins promote cohesion independently of one another. Consistent with this, we find that Mcd1/Scc1 is less abundant on chromosomes in cells lacking Clb5 and Clb6 during replication stress. However, clb5Δ clb6Δ cells do accumulate Mcd1/Scc1 at centromeres upon mitotic arrest, suggesting that the cyclin-dependent mechanism is S phase specific. These data indicate that Clb5 and Clb6 promote cohesion which is then protected by Pds1 and that both mechanisms are required during replication stress.

Topoisomerase II (Topo II) performs topological modifications on double-stranded DNA molecules that are essential for chromosome condensation, resolution, and segregation. In mammals, G2 and metaphase cell cycle delays induced by Topo II poisons have been proposed to be the result of checkpoint activation in response to the catenation state of DNA. However, the apparent lack of such controls in model organisms has excluded genetic proof that Topo II checkpoints exist and are separable from the conventional DNA damage checkpoint controls. But here, we define a Topo II-dependent G2/M checkpoint in a genetically amenable eukaryote, budding yeast, and demonstrate that this checkpoint enhances cell survival. Conversely, a lack of the checkpoint results in aneuploidy. Neither DNA damage-responsive pathways nor Pds1/securin are needed for this checkpoint. Unusually, spindle assembly checkpoint components are required for the Topo II checkpoint, but checkpoint activation is not the result of failed chromosome biorientation or a lack of spindle tension. Thus, compromised Topo II function activates a yeast checkpoint system that operates by a novel mechanism. © 2006 by Cold Spring Harbor Laboratory Press., This work was funded by NIH grant CA099033 (D.J.C.).