title: Sensing of replication stress and Mec1 activation act through two independent pathways involving the 9-1-1 complex and DNA polymerase ε.
abstract: Following DNA damage or replication stress, budding yeast cells activate the Rad53 checkpoint kinase, promoting genome stability in these challenging conditions. The DNA damage and replication checkpoint pathways are partially overlapping, sharing several factors, but are also differentiated at various levels. The upstream kinase Mec1 is required to activate both signaling cascades together with the 9-1-1 PCNA-like complex and the Dpb11 (hTopBP1) protein. After DNA damage, Dpb11 is also needed to recruit the adaptor protein Rad9 (h53BP1). Here we analyzed the mechanisms leading to Mec1 activation in vivo after DNA damage and replication stress. We found that a ddc1Δdpb11-1 double mutant strain displays a synthetic defect in Rad53 and H2A phosphorylation and is extremely sensitive to hydroxyurea (HU), indicating that Dpb11 and the 9-1-1 complex independently promote Mec1 activation. A similar phenotype is observed when both the 9-1-1 complex and the Dpb4 non-essential subunit of DNA polymerase ε (Polε) are contemporarily absent, indicating that checkpoint activation in response to replication stress is achieved through two independent pathways, requiring the 9-1-1 complex and Polε.
authors: Department of Biomolecular Sciences and Biotechnology, Università degli Studi di Milano, Milano, Italy.
authors: Fabio
authors: F
authors: Puddu
authors: Gabriele
authors: G
authors: Piergiovanni
authors: Paolo
authors: P
authors: Plevani
authors: Marco
authors: M
authors: Muzi-Falconi
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism; D004257:DNA Polymerase II / Q000378:metabolism*; D004261:DNA Replication; D004789:Enzyme Activation; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012264:Ribonucleotide Reductases / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D015398:Signal Transduction; D014466:Ultraviolet Rays
chemical_list: D018797:Cell Cycle Proteins; C097185:DPB11 protein, S cerevisiae; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; D029701:Saccharomyces cerevisiae Proteins; D012264:Ribonucleotide Reductases; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D004257:DNA Polymerase II