Structural insights into the cooperative binding of SeqA to a tandem GATC repeat.

SeqA is a negative regulator of DNA replication in Escherichia coli and related bacteria that functions by sequestering the origin of replication and facilitating its resetting after every initiation event. Inactivation of the seqA gene leads to unsynchronized rounds of replication, abnormal localization of nucleoids and increased negative superhelicity. Excess SeqA also disrupts replication synchrony and affects cell division.

Switching protein-DNA recognition specificity by single-amino-acid substitutions in the P1 par family of plasmid partition elements.

The P1, P7, and pMT1 par systems are members of the P1 par family of plasmid partition elements. Each has a ParA ATPase and a ParB protein that recognizes the parS partition site of its own plasmid type to promote the active segregation of the plasmid DNA to daughter cells. ParB contacts two parS motifs known as BoxA and BoxB, the latter of which determines species specificity.

Hda inactivation of DnaA is the predominant mechanism preventing hyperinitiation of Escherichia coli DNA replication.

Initiation of DNA replication from the Escherichia coli chromosomal origin is highly regulated, assuring that replication occurs precisely once per cell cycle. Three mechanisms for regulation of replication initiation have been proposed: titration of free DnaA initiator protein by the datA locus, sequestration of newly replicated origins by SeqA protein and regulatory inactivation of DnaA (RIDA), in which active ATP-DnaA is converted to the inactive ADP-bound form. DNA microarray analyses showed that the level of initiation in rapidly growing cells that lack datA was indistinguishable from that in wild-type cells, and that the absence of SeqA protein caused only a modest increase in initiation, in agreement with flow-cytometry data.

Crystal structure of a SeqA-N filament: implications for DNA replication and chromosome organization.

Escherichia coli SeqA binds clusters of transiently hemimethylated GATC sequences and sequesters the origin of replication, oriC, from methylation and premature reinitiation. Besides oriC, SeqA binds and organizes newly synthesized DNA at replication forks. Binding to multiple GATC sites is crucial for the formation of stable SeqA-DNA complexes.

Surprising dependence on postsegregational killing of host cells for maintenance of the large virulence plasmid of Shigella flexneri.

Low-copy-number plasmids all encode multiple systems to ensure their propagation, including replication, partition (active segregation), and postsegregational killing (PSK) systems. PSK systems kill those rare cells that lose the plasmid due to replication or segregation errors. PSK systems should not be used as the principle means of maintaining the plasmid.

Interplay between plasmid partition and postsegregational killing systems.

Active partition systems and postsegregational killing (PSK) systems are present together in naturally occurring low-copy-number plasmids. Theory suggests that PSK may act as the ultimate determinant of plasmid retention, whereas the partition system may minimize the growth penalty to the host, resulting in a near-ideal symbiosis when the systems combine. Here, we prove the validity of this principle for a specific case involving the P1par system and the mvp PSK system.

Transcriptional interference by a complex formed at the centromere-like partition site of plasmid P1.

The partition site, parS, promotes accurate segregation of the replicated P1 plasmid to daughter cells when the P1-encoded ParA and ParB proteins are supplied. The parS site was inserted into the Escherichia coli chromosome between the promoter and the structural gene for beta-galactosidase, lacZ. There was little interference with lacZ expression when ParA and ParB were supplied in trans.