Cdc45 is limiting for replication initiation in humans

Cdc45 is an essential protein that together with Mcm2-7 and GINS forms the eukaryotic replicative helicase CMG. Cdc45 seems to be rate limiting for the initial unwinding or firing of replication origins. In line with this view, Cdc45-overexpressing cells fired at least twice as many origins as contr...

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Published in:Cell cycle (Georgetown, Tex.) Tex.), 2016-01, Vol.15 (7), p.974-985
Main Authors: Köhler, Carsten, Koalick, Dennis, Fabricius, Anja, Parplys, Ann Christin, Borgmann, Kerstin, Pospiech, Helmut, Grosse, Frank
Format: Article
Language:English
Subjects:
Apoptosis
Cell Cycle Proteins - metabolism
DNA Replication
DNA, Single-Stranded - analysis
HeLa Cells
Histones - metabolism
Humans
Replication Origin
S Phase Cell Cycle Checkpoints
Signal Transduction
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Summary:Cdc45 is an essential protein that together with Mcm2-7 and GINS forms the eukaryotic replicative helicase CMG. Cdc45 seems to be rate limiting for the initial unwinding or firing of replication origins. In line with this view, Cdc45-overexpressing cells fired at least twice as many origins as control cells. However, these cells displayed an about 2-fold diminished fork elongation rate, a pronounced asymmetry of replication fork extension, and an early S phase arrest. This was accompanied by H2AX-phosphorylation and subsequent apoptosis. Unexpectedly, we did not observe increased ATR/Chk1 signaling but rather a mild ATM/Chk2 response. In addition, we detected accumulation of long stretches of single-stranded DNA, a hallmark of replication catastrophe. We conclude that increased origin firing by upregulated Cdc45 caused exhaustion of the single-strand binding protein RPA, which in consequence diminished the ATR/Chk1 response; the subsequently occurring fork breaks led to an ATM/Chk2 mediated phosphorylation of H2AX and eventually to apoptosis.
ISSN:1538-4101
1551-4005
DOI:10.1080/15384101.2016.1152424