The genomic island SGI1 and its variants, the important vehicles of multi-resistance in Salmonella strains, are integrative elements mobilized exclusively by the conjugative IncA/C plasmids. Integration and excision of the island are carried out by the SGI1-encoded site-specific recombinase Int and the recombination directionality factor Xis. Chromosomal integration ensures the stable maintenance and vertical transmission of SGI1, while excision is the initial step of horizontal transfer, followed by conjugation and integration into the recipient. We report here that SGI1 not only exploits the conjugal apparatus of the IncA/C plasmids but also utilizes the regulatory mechanisms of the conjugation system for the exact timing and activation of excision to ensure efficient horizontal transfer. This study demonstrates that the FlhDC-family activator AcaCD, which regulates the conjugation machinery of the IncA/C plasmids, serves as a signal of helper entry through binding to SGI1 xis promoter and activating SGI1 excision. Promoters of int and xis genes have been identified and the binding site of the activator has been located by footprinting and deletion analyses. We prove that expression of xis is activator-dependent while int is constitutively expressed, and this regulatory mechanism is presumably responsible for the efficient transfer and stable maintenance of SGI1.
Many organisms control initiation of DNA replication by limiting supply or activity of initiator proteins. In plasmids, such as P1, initiators are limited primarily by transcription and dimerization. However, the relevance of initiator limitation to plasmid copy number control has appeared doubtful, because initiator oversupply increases the copy number only marginally. Copy number control instead has been attributed to initiator-mediated plasmid pairing (''handcuffing''), because initiator mutations to handcuffing deficiency elevates the copy number significantly. Here, we present genetic evidence of a role for initiator limitation in plasmid copy number control by showing that autorepression-defective initiator mutants also can elevate the plasmid copy number. We further show, by quantitative modeling, that initiator dimerization is a homeostatic mechanism that dampens active monomer increase when the protein is oversupplied. This finding implies that oversupplied initiator proteins are largely dimeric, partly accounting for their limited ability to increase copy number. A combination of autorepression, dimerization, and handcuffing appears to account fully for control of P1 plasmid copy number.autorepression ͉ DNA replication control ͉ homeostatic control ͉ plasmid copy number control
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