The Rad4TopBP1 ATR-Activation Domain Functions in G1/S Phase in a Chromatin-Dependent Manner

摘要: DNA damage checkpoint activation can be subdivided in two steps: initial and signal amplification. The events distinguishing these phases their genetic determinants remain obscure. TopBP1, a mediator protein containing multiple BRCT domains, binds to activates the ATR/ATRIP complex through its ATR-Activation Domain (AAD). We show that Schizosaccharomyces pombe Rad4TopBP1 AAD–defective strains are sensitive during G1/S-phase, but not G2. Using lacO-LacI tethering, we developed damage–independent assay for is AAD–dependent. In this assay, requires histone H2A phosphorylation, interaction between TopBP1 9-1-1 complex, mediated by phospho-binding activity of Crb253BP1. Consistent with model where AAD–dependent ssDNA/RPA–independent functions amplify otherwise weak signals, demonstrate AAD important Chk1 phosphorylation when resection limited G2 ablation resecting nuclease, Exo1. also acts additively Rad9 G1/S phase propose Rad3ATR amplification particularly limiting. S. pombe, manifests relies on protein–chromatin interactions.