DESCRIPTION (provided by applicant): Programmed cell death (PCD), defined as an active process which results in cell suicide, is recognized as an essential mechanism in multicellular organisms. In 1996, we reported on the first chromosomal toxin-antitoxin system discovered to be responsible for PCD in bacteria. This was the Escherichia coli (E. coli) mazEF regulateable module in which mazF encodes a stable toxin, MazF, and mazE encodes a labile antitoxin, MazE, that overcomes the lethal effect of MazF. Since 1996, we have made considerable progress in understanding the genetic, biochemical, structural and physiological properties of this system, and its relation to stressful conditions and the action of some well known antibiotics. In continuation of this work, here we propose to extend our research by a comprehensive characterization of a novel quorum sensing Extra-cellular Death Factor (EDF) that we were excited to discover as being a signaling peptide molecule required for E. coli mazEF-mediated cell death. Our five specific aims are as follows: 1 to characterize the exact chemical composition of E. coli EDF; 2. to elucidate the mechanism of EDF action; 3.To elucidate the relation of EDF to mazEF and to E. coli other toxin-antitoxin systems; 4.To characterize the EDF receiver(s) system; 5.To characterize EDFs of bacteria other than E. coli. The integrative research on EDF and its relation to bacterial death that we propose here should contribute to the understanding of a unique novel quorum sensing signaling molecule and its importance to bacterial PCD. Moreover, our results should also help to clarify a fundamental biological phenomenon of bacterial communication and thereby the multicellular behavior of bacterial populations. Practically, the results of our study may be useful as a basis for a novel strategy for generating a new class of antibiotics that trigger bacterial PCD, and thereby to help what has become a public health problem.