Toxin-antitoxin systems are defined as a group of plasmid- and chromosome-encoded loci that specify a cell toxin and a protein antitoxin. Plasmid-encoded toxin-antitoxin systems stabilize their replicons by killing plasmid-free cells. Here, we show that the relBE genes of Escherichia coli K-12 have all the basic features previously connected with toxin-antitoxin systems: (i) relE encodes a cytotoxin lethal or inhibitory to host cells; (ii) relB encodes an antitoxin that prevents the lethal action of the relE-encoded toxin; (iii) the relBE genes stabilize a mini-R1 test plasmid; and (iv) the RelB antitoxin autoregulates the relBEF operon at the level of transcription. Using database searching, we found relBE homologues on the chromosomes of E. coli K-12, Haemophilus influenzae and Vibrio cholerae. A fifth relBE homologue was identified on the enterotoxin encoding E. coli plasmid P307. Indirect evidence suggests that the toxicity of RelE may be related to the inhibition of protein synthesis. Based on these observations, we propose a model that explains the delayed relaxed phenotype associated with mutations in relB.