Transposon A-generated mutations in the mercuric resistance genes of plasmid R100-1.
A series of 23 transposon 801(Tn801)-induced mutations of plasmid R100-1 from mercuric salts resistance to sensitivity was studied. Although Tn801 transposed frequently into the mer region of the plasmid, fine structural analysis showed that the site of insertion within mer varied. About one-half of the Tn801 insertion events also caused a deletion of greater than 1 megadalton. Genetic and restriction endonuclease EcoRI and BamHI analysis of the mutant plasmid deoxyribonucleic acid elucidated the organization of the mer operon and suggested the existence of a trans-acting regulatory factor governing resistance to mercuric salts. Tn801 insertions leading to mercuric sensitivity occurred in the restriction endonuclease fragments EcoRI-H and EcoRI-I. Regulatory mutations leading to a 50-fold-reduced synthesis of mercuric reductase enzyme occurred in two complementation classes thought to represent the gene for a trans-acting inducer molecule and a cis-acting operator-promoter sequence. Mutations leading to total loss of the enzyme mercuric reductase occurred on both the EcoRI-H and EcoRI-I fragments, showing that the structural gene for this enzyme (merA) bridges the EcoRI cleavage site separating the segments. Hypersensitivity to mercuric salts resulted when Tn801 insertion occurred in the reductase gene in the operatordistal portion of the operon. Hypersensitive cells inducibly bound three to five times more Hg2+ at low concentrations than did sensitive (plasmidless) cells. This finding led to the proposal that another gene (merT) controls uptake of Hg2+ by the cells. Transcription of the operon was deduced to start in the EcoRI-H fragment and to move into the EcoRI-I fragment of the plasmid genome.
D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005816:Genetic Complementation Test; D008628:Mercury / Q000378:metabolism / Q000494:pharmacology*; D009154:Mutation; D009941:Organomercury Compounds / Q000494:pharmacology; D011815:R Factors; D011995:Recombination, Genetic
D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D009941:Organomercury Compounds; D008628:Mercury