Linking PCNA-dependent replication and ATR by human Claspin

French National Centre for Scientific Research, Lutetia Parisorum, Île-de-France, France
Biochemical and Biophysical Research Communications (Impact Factor: 2.3). 04/2007; 354(4):1028-33. DOI: 10.1016/j.bbrc.2007.01.091
Source: PubMed


Recent studies in Xenopus have identified a new checkpoint protein called Claspin that is believed to transduce the checkpoint DNA damage signals to Chk1 kinase. Here we show that the human Claspin homolog is a chromatin bound protein either in the absence or in the presence of damaged DNA, independent of its association with ATR. Furthermore, we show that human Claspin is found in complex with PCNA, an essential component of the DNA replication machinery, and is released upon DNA replication arrest. Interfering with PCNA function by overexpression of p21 mutant, impaired in its interaction with Cdks but not with PCNA, leads to ATR-dependent Chk1 activation. These findings suggest that the dissociation of Claspin-PCNA could be part of the signal leading to Chk1 activation.

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    • "Cells were treated for 4 h with BL (5 mU/ml) (Calbiochem, San Diego, CA, USA). For cell cycle synchronization, cells were treated for 24 h with 2 mM of hydroxyurea (Sigma- Aldrich, St Louis, MO, USA) and released from hydroxyurea block as previously described (Brondello et al., 2007). G2/M cells were obtained by a 24 h treatment with 10 mg/ml of nocodazole (Sigma-Aldrich). "
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    • "By immunoprecipitating endogenous Claspin and Timeless, we found that these two proteins associate with each other and with PCNA even in undamaged cells (Fig. 4A,B). Consistent with our results, two independent studies reported recently the interactions of Claspin with PCNA and Timeless (Brondello et al. 2007; Gotter et al. 2007). "
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