The Rad[4.sup.TopBP1] ATR-Activation Domain functions in G1/S phase in a chromatin-dependent manner

DNA damage checkpoint activation can be subdivided in two steps: initial activation and signal amplification. The events distinguishing these two phases and their genetic determinants remain obscure. TopBP1, a mediator protein containing multiple BRCT domains, binds to and activates the ATR/ATRIP co...

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Bibliographic Details
Published in:PLoS genetics 2012-06, Vol.8 (6)
Main Authors: Lin, Su-Jiun, Wardlaw, Christopher P, Morishita, Takashi, Miyabe, Izumi, Chahwan, Charly, Caspari, Thomas, Schmidt, Ulrike, Carr, Antony M, Garcia, Valerie
Format: Article
Language:English
Subjects:
Chromatin
DNA damage
Genetic aspects
Microbial genetics
Physiological aspects
Saccharomyces
Online Access:Get full text
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Summary:DNA damage checkpoint activation can be subdivided in two steps: initial activation and signal amplification. The events distinguishing these two phases and their genetic determinants remain obscure. TopBP1, a mediator protein containing multiple BRCT domains, binds to and activates the ATR/ATRIP complex through its ATR-Activation Domain (AAD). We show that Schizosaccharomyces pombe Rad[4.sup.TopBP1] AAD-defective strains are DNA damage sensitive during Gl/S-phase, but not during G2. Using /acO-Laci tethering, we developed a DNA damage-independent assay for checkpoint activation that is Rad4 pBP1 AAD-dependent. in this assay, checkpoint activation requires histone H2A phosphorylation, the interaction between TopBPl and the 9-1-1 complex, and is mediated by the phospho-binding activity of Crb[2.sup.53BP1]. Consistent with a model where Rad[4.sup.TopBP1] AAD-dependent checkpoint activation is ssDNA/RPA-independent and functions to amplify otherwise weak checkpoint signals, we demonstrate that the Rad[4.sup.TopBP1] AAD is important for Chk1 phosphorylation when resection is limited in G2 by ablation of the resecting nuclease, Exol. We also show that the Rad[4.sup.TopBP1] AAD acts additively with a Rad9 AAD in Gl/S phase but not G2. We propose that AAD-dependent Rad[3.sup.ATR] checkpoint amplification is particularly important when DNA resection is limiting. in S. pombe, this manifests in Gl/S phase and relies on proteinchromatin interactions.
ISSN:1553-7390
1553-7404
DOI:10.1371/journal.pgen.1002801