Omega (ParB) binding sites together with the RNA polymerase-recognized sequence are essential for centromeric functions of the Pω region in the partition system of pSM19035

Abstract

Partition systems contribute to stable plasmid inheritance in bacteria through the active separation of DNA molecules to daughter cells, and the centromeric sequence located either upstream or downstream of canonical partition operons plays an important role in this process. A specific DNAbinding protein binds to this sequence and interacts with the motor NTPase protein to form a nucleoprotein complex. The inc18-family plasmid pSM19035 is partitioned by products of d and ! genes, with d encoding a Walker-type ATPase and ! encoding a DNA-binding protein. As the two genes are transcribed separately, this system differs from others in its organization; nonetheless, expression of these genes is regulated by Omega, which also regulates the copy number of the plasmid (by controlling copS gene expression). Protein Omega specifically recognizes WATCACW heptad repeats. In this study, we constructed a synthetic d! operon to enable an analysis of the centromeric functions of Omega-binding sites P d , P ! and P copS , discrete from their promoter functions. Our results show that these three regions do not support plasmid stabilization equally. We demonstrate that the P ! site alone can simultaneously drive the expression of partition genes from the synthetic d! operon and act as a unique centromeric sequence to promote the most efficient plasmid partitioning. Moreover, P ! can support the centromeric function in concert with the synthetic d! operon expressed from a heterologous promoter demonstrating that P ! is the main centromeric sequence of the d-! partition system. Additionally, the RNA polymeraserecognized sequence in P ! is essential for its centromeric function.

Segrosome Complex Formation during DNA Trafficking in Bacterial Cell Division

“…The full centromere contains 3 separate parS sites, consisting of 9, 7, and 10 iterons that occur twice in the plasmid genome ( parS1, parS1 ′, parS2, parS2 ′, parS3, parS3 ′, de la Hoz et al, 2000 , 2004 ; Dmowski et al, 2006 ). However, parS2 appears to be the main centromeric sequence (Dmowski and Kern-Zdanowicz, 2016 ). Interestingly, each parS overlaps with the promoters of genes involved in plasmid copy number and maintenance: parS1 with Pδ, parS2 with Pω and parS3 with P copS (de la Hoz et al, 2000 ).…”