Are minichromosomes valid model systems for DNA replication control? Lessons learned from Escherichia coli

Abstract

Initiation of chromosome replication is a key event in the life cycle of any organism. Little is known, however, about the regulatory mechanisms of this vital process. Conventionally, the initiation mechanism of chromosome replication in microorganisms has been studied using plasmids in which an origin of chromosome replication has been cloned, rather than using the chromosome itself. The reason for this is that even bacterial chromosomes are so large that biochemical and genetic manipulations become difficult and cumbersome. Recently, the combination of flow cytometry and genetic methods, in which modifications of the replication origin are systematically introduced onto the chromosome, has made possible detailed studies of the molecular events involved in the control of replication initiation in Escherichia coli. The results indicate that requirements for initiation at the chromosomal origin, oriC, are drastically different from those for initiation at cloned oriC.

Figure 1

The minimal oriC and its flanking regions. The positions of the five DnaA boxes, R1-R4 and M, the 13-mer repeats L, M, and R, the AT-rich sequence (AT), and binding sites for IHF and Fis are indicated. Arrows represent the location and direction of promoters.