Specificity ofE. coli K-12 chromosomal segment regulating the expression of systems inhibiting flac plasmid transfer

Abstract

Specificity of E. coli K-12 chromosomal Thr-Leu-segment (genetic locus tis) regulating the expression of systems inhibiting Flac plasmid transfer is revealed. The findings point to a complex (polygenous) structure of this locus. Key Words: plasmid; chromosomal genes; transfer inhibition system; gene expressionChromosome segments of bacterial host cells participate in the regulation of F plasrnid and some other F-like plasmid transfer along with plasmid genetic systems (f'm-systems) [1][2][3][4]. Previously, we revealed the role of E. coli K-12 Thr-Leu chromosome segment in the expression of fin V, an F-like plasmid pAP53 system [2]. Possible effect of chromosome locus, denoted as locus tis, on the function of other known systems regulating plasmid transfer is still unclear. In this study we compared the specificity of locus tis toward fm systems OP, Q, U, V, and W, capable of inhibiting conjugative transfer of Flac plasmid [5,6]. MATERIALS AND METHODSReference derepressed (drd) Flac plasmid and reference repressed (rd) plasmids R100 (fro OP), TP108 (fro Q), JR66a (fin U), R485 (fin V), and R455 (fro W) from N. Willetts' collection (UK) were used. Plasmid hosts were E. coli K-12 cells C600 (Lac, Thr, Leu, and Rif) and AP132 (Lac and Nal). E. coli strain Hfr C were donors of the tis chromosome locus. Department of BiolotEr and General Genetics, Russian University of Peoples' Friendship, MoscowConjugation crossing-over of bacteria and selection of genetic recombinants and plasmid transconjugates were carried out routinely. For assessing the capacity of Fin + plasmids to inhibit the functions of Flac plasmid transfer genes (Tra-function), the transfer inhibition index (TII) was calculated as the ratio of the rate of Flac plasmid transfer from singleplasmid donor cells to the rate of the same plaslnid transfer from diplasmid transconjugate cells. Functional activity of "sex" piles whose production is regulated by the Flac plasmid tra genes was assessed from the sensitivity of relevant bacterial cells to pilespecific MS2 phage. RESULTSConjugation crossing over of Hfr C (Tis +) cells with C600 (Tis-) recipient strain cells was performed to obtain genetic recombinants containing tis locus. The resultant Thr+Leu + recombinants were used as bacterial hosts containing drd Flac plasmid and one reference rd plasmid with a known fin type. The corresponding single-plasmid transconjugates (recombinant cells conraining Flac plasmid alone) were used as control.Flac plasmid TII for each reference Fin + plasmid was estimated from the results of subsequent cros-