The Rad4TopBP1 ATR-Activation domain functions in G1/S phase in a chromatin-dependent manner

Lin, Su-Jiun, Wardlaw, Christopher Paul, Morishita, Takashi, Miyabe, Izumi, Chahwan, Charly, Caspari, Thomas, Schmidt, Ulrike, Carr, Antony and Garcia, Valerie (2012) The Rad4TopBP1 ATR-Activation domain functions in G1/S phase in a chromatin-dependent manner. PLoS Genetics, 8 (6). ISSN 1553-7390

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Abstract

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 Rad4TopBP1 AAD–defective strains are DNA damage sensitive during G1/S-phase, but not
during G2. Using lacO-LacI tethering, we developed a DNA damage–independent assay for checkpoint activation that is
Rad4TopBP1 AAD–dependent. In this assay, checkpoint activation requires histone H2A phosphorylation, the interaction
between TopBP1 and the 9-1-1 complex, and is mediated by the phospho-binding activity of Crb253BP1. Consistent with a
model where Rad4TopBP1 AAD–dependent checkpoint activation is ssDNA/RPA–independent and functions to amplify
otherwise weak checkpoint signals, we demonstrate that the Rad4TopBP1 AAD is important for Chk1 phosphorylation when
resection is limited in G2 by ablation of the resecting nuclease, Exo1. We also show that the Rad4TopBP1 AAD acts additively
with a Rad9 AAD in G1/S phase but not G2. We propose that AAD–dependent Rad3ATR checkpoint amplification is
particularly important when DNA resection is limiting. In S. pombe, this manifests in G1/S phase and relies on protein–
chromatin interactions.

Item Type: Article
Schools and Departments: School of Life Sciences > Sussex Centre for Genome Damage and Stability
Subjects: Q Science > QH Natural history > QH0301 Biology > QH0426 Genetics
Depositing User: Valerie Garcia
Date Deposited: 01 Nov 2012 11:21
Last Modified: 07 Mar 2017 04:39
URI: http://sro.sussex.ac.uk/id/eprint/41768

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