Molecular Basis of the Essential S Phase Function of the Rad53 Checkpoint Kinase
| Autor: | Ming-Daw Tsai, Eric S.-W. Chen, Alessandro Fazio, Nicolas C. Hoch, Shun-Chung Wang, Andrew Hammet, Achille Pellicioli, Robert J. Buckland, Andrei Chabes, Jörg Heierhorst |
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| Rok vydání: | 2013 |
| Předmět: |
Saccharomyces cerevisiae Proteins
DNA damage Mutant Cell Cycle Proteins Saccharomyces cerevisiae Protein Serine-Threonine Kinases Biology medicine.disease_cause S Phase medicine CHEK1 Phosphorylation DNA Fungal Molecular Biology Checkpoint Kinase 2 Mutation Kinase Intracellular Signaling Peptides and Proteins DNA replication Articles Cell Biology G2-M DNA damage checkpoint Molecular biology Protein Structure Tertiary Enzyme Activation Proteolysis Gene Deletion DNA Damage |
| Zdroj: | Molecular and Cellular Biology. 33:3202-3213 |
| ISSN: | 1098-5549 |
| DOI: | 10.1128/mcb.00474-13 |
| Popis: | The essential yeast kinases Mec1 and Rad53, or human ATR and Chk1, are crucial for checkpoint responses to exogenous genotoxic agents, but why they are also required for DNA replication in unperturbed cells remains poorly understood. Here we report that even in the absence of DNA-damaging agents, the rad53-4AQ mutant, lacking the N-terminal Mec1 phosphorylation site cluster, is synthetic lethal with a deletion of the RAD9 DNA damage checkpoint adaptor. This phenotype is caused by an inability of rad53-4AQ to activate the downstream kinase Dun1, which then leads to reduced basal deoxynucleoside triphosphate (dNTP) levels, spontaneous replication fork stalling, and constitutive activation of and dependence on S phase DNA damage checkpoints. Surprisingly, the kinase-deficient rad53-K227A mutant does not share these phenotypes but is rendered inviable by additional phosphosite mutations that prevent its binding to Dun1. The results demonstrate that ultralow Rad53 catalytic activity is sufficient for normal replication of undamaged chromosomes as long as it is targeted toward activation of the effector kinase Dun1. Our findings indicate that the essential S phase function of Rad53 is comprised by the combination of its role in regulating basal dNTP levels and its compensatory kinase function if dNTP levels are perturbed. |
| Databáze: | OpenAIRE |
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