The incC korB region of RK2 repositions a mini-RK2 replicon in Escherichia coli

Analysis by fluorescence microscopy has established that plasmid RK2 in Escherichia coli and other gram-negative bacteria is present as discrete clusters that are located inside the nucleoid at the mid- or quarter-cell positions. A mini-RK2 replicon containing an array of tetO repeats was visualized in E. coli cells that express a TetR-EYFP fusion protein. Unlike intact RK2, the RK2 mini-replicon (pCVI) was localized as a cluster at the cell poles outside of the nucleoid. Insertion of the O-B1 incC korB partitioning (par) region of RK2 into pCVI resulted in a shift of the mini-replicon to within the nucleoid region at the mid- and quarter-cell positions. Despite the repositioning of the mini-RK2 replicon to the cellular positions where intact RK2 is normally located, the insertion of the intact O-B1 incC korB region did not significantly stabilize the mini-RK2 plasmid during cell growth. Deletions within the O-B1 incC or the korB region resulted in a failure of this par region to move pCVI out of its polar position. The insertion of the par system of plasmid F into pCVI resulted in a similar shift in the location of pCV1 to the nucleoid region. Unlike O-B1 incC korB, the insertion of the RK2 parABC resolvase system into pCVI did not affect the polar positioning of pCVI. This effect Of O-B1 incC korB on the location of pCV1 provides additional evidence for a partitioning role of this region of plasmid RK2. However, the failure of this region to significantly increase the stability of the mini-RK2 plasmid indicates that the localization of the plasmid to the mid- and quarter cell positions in E coli is not in itself sufficient for the stable maintenance of plasmid RK2. (c) 2007 Elsevier Inc. All rights reserved.