期刊
GENES & DEVELOPMENT
卷 23, 期 10, 页码 1221-1233出版社
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1101/gad.1775809
关键词
DnaA reactivation; nucleotide exchange; initiation regulation; functional DNA sequence; protein complex; replication cycle
资金
- Ministry of Education, Culture, Sports, Technology, and Science of Japan
In Escherichia coli, ATP-DnaA, unlike ADP-DnaA, can initiate chromosomal replication at oriC. The level of cellular ATP-DnaA fluctuates, peaking at around the time of replication initiation. However, it remains unknown how the ATP-DnaA level increases coordinately with the replication cycle. In this study, we show that two chromosomal intergenic regions, herein termed DnaA-reactivating sequence 1 (DARS1) and DnaA-reactivating sequence 2 (DARS2), directly promote regeneration of ATP-DnaA from ADP-DnaA by nucleotide exchange, resulting in the promotion of replication initiation in vitro and in vivo. Coordination of initiation with the cell cycle requires DARS activity and its regulation. Oversupply of DARSs results in increase in the ATP-DnaA level and enhancement of replication initiation, which can inhibit cell growth in an oriC-dependent manner. Deletion of DARSs results in decrease in the ATP-DnaA level and inhibition of replication initiation, which can cause synthetic lethality with a temperature-sensitive mutant dnaA and suppression of overinitiation by the lack of seqA or datA, negative regulators for initiation. DARSs bear a cluster of DnaA-binding sites. DnaA molecules form specific homomultimers on DARS1, which causes specific interactions among the protomers, reducing their affinity for ADP. Our findings reveal a novel regulatory pathway that promotes the initiation of chromosomal replication via DnaA reactivation.
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