The kfrA gene is the first in a tricistronic operon required for survival of IncP-1 plasmid R751

Abstract

ThekfrAgene of the IncP-1 broad-host-range plasmids is the best-studied member of a growing gene family that shows strong linkage to the minimal replicon of many low-copy-number plasmids. KfrA is a DNA binding protein with a long, alpha-helical, coiled-coil tail. Studying IncP-1βplasmid R751, evidence is presented thatkfrAand its downstream genesupf54.8andupf54.4were organized in a tricistronic operon (renamed herekfrA kfrB kfrC), expressed from autoregulatedkfrAp, that was also repressed by KorA and KorB. KfrA, KfrB and KfrC interacted and may have formed a multi-protein complex. Inactivation of eitherkfrAorkfrBin R751 resulted in long-term accumulation of plasmid-negative bacteria, whereas wild-type R751 itself persisted without selection. Immunofluorescence studies showed that KfrAR751formed plasmid-associated foci, and deletion of the C terminus of KfrA caused plasmid R751ΔC2kfrAfoci to disperse and mislocalize. Thus, the KfrABC complex may be an important component in the organization and control of the plasmid clusters that seem to form the segregating unit in bacterial cells. The studied operon is therefore part of the set of functions needed for R751 to function as an efficient vehicle for maintenance and spread of genes in Gram-negative bacteria.