A plasmid-encoded H-NS protein selectively binds its own plasmid

H-NS is an abundant nucleoid-associated protein found in Enterobacterales species. Some conjugative plasmids encode H-NS homologues, which are thought to facilitate plasmid maintenance and reduce the fitness costs associated with plasmid carriage. Here, we characterize HppX_CROD2, an H-NS homologue encoded by the IncX4 plasmid pCROD2 of Citrobacter rodentium. Our data indicate that HppX_CROD2 has a strong preference for binding pCROD2 over the chromosome or other plasmids. By contrast, chromosomally encoded H-NS displays no preference for plasmid sequence. When expressed from a heterologous plasmid in Escherichia coli, HppX_CROD2 showed similar DNA-sequence preference to chromosomally encoded H-NS. Moreover, HppX_CROD2 binding to a sequence from pCROD2 was much lower when that sequence was cloned in a laboratory plasmid. Thus, HppX_CROD2 preferentially binds DNA in the context of the plasmid where it is encoded, a phenomenon we term "cognate plasmid specificity". We propose that cognate plasmid specificity occurs through recognition of plasmid-specific DNA topology generated by plasmid-encoded topoisomerases. Cognate plasmid specificity may insulate regulation of plasmid genes from the effects of host DNA, while minimizing disruption of host chromosome regulation due to plasmid carriage.