Cdc7 activates replication checkpoint by phosphorylating the Chk1-binding domain of Claspin in human cells.

Chi-Chun Yang,Mayumi Shindo,Hiroyuki Kato,Hisao Masai

doi:10.7554/elife.50796

Chi-Chun Yang, Mayumi Shindo + Show 2 more

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https://doi.org/10.7554/elife.50796

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Journal: eLife	Publication Date: Dec 31, 2019
Citations: 36	License type: CC BY 4.0

Affiliation: Tokyo Metropolitan Institute of Medical Science

Abstract

Replication checkpoint is essential for maintaining genome integrity in response to various replication stresses as well as during the normal growth. The evolutionally conserved ATR-Claspin-Chk1 pathway is induced during replication checkpoint activation. Cdc7 kinase, required for initiation of DNA replication at replication origins, has been implicated in checkpoint activation but how it is involved in this pathway has not been known. Here, we show that Cdc7 is required for Claspin-Chk1 interaction in human cancer cells by phosphorylating CKBD (Chk1-binding-domain) of Claspin. The residual Chk1 activation in Cdc7-depleted cells is lost upon further depletion of casein kinase1 (CK1γ1), previously reported to phosphorylate CKBD. Thus, Cdc7, in conjunction with CK1γ1, facilitates the interaction between Claspin and Chk1 through phosphorylating CKBD. We also show that, whereas Cdc7 is predominantly responsible for CKBD phosphorylation in cancer cells, CK1γ1 plays a major role in non-cancer cells, providing rationale for targeting Cdc7 for cancer cell-specific cell killing.

Highlights

Eukaryotic DNA replication depends on the formation of pre-Replicative Complex on chromatin during the G1 phase of cell cycle, which is mediated by assembly of Orc, Cdc6, Cdt1 and Mcm proteins
Cdc7 is required for replication checkpoint activation in human cells
We reported that conditional knockout of Cdc7 in mouse ES cells resulted in loss of Chk1 activation in response to hydroxyl urea (HU) or UV irradiation

Summary

Introduction

Eukaryotic DNA replication depends on the formation of pre-Replicative Complex on chromatin during the G1 phase of cell cycle, which is mediated by assembly of Orc, Cdc, Cdt and Mcm proteins. Cdc kinase plays a crucial role in initiation of DNA replication by phosphorylating Mcm proteins, essential as a part of the replicative helicase (Masai and Arai, 2002; Masai et al, 2010; Labib, 2010). Inhibition of DNA replication by hydroxyl urea (HU) or UV triggers cellular responses known as replication stress checkpoint (Branzei and Foiani, 2009). Conserved checkpoint kinases (Mec1Rad3-ATR) are activated in response to replication stress, which activates Rad53-Cds1Chk effector kinases, that inhibits progression of S phase as well as entry into M phase to reduce the genomic instability potentially caused by replication fork arrest. ATR phosphorylates Chk in the Clapsin-Chk complex more efficiently than Chk alone in the absence of Claspin in virto (Lindsey-Boltz et al, 2009)

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