This proposal is concerned with the genetic analysis of genes required for cell division in prokaryotic cells. The ultimate goal of the project is to completely understand the molecular mechanism of this complex fundamental process. In particular, cytokinesis is studied in the filamentous soil bacterium Streptomyces coelicolor, a species readily amenable to genetic analysis. S. coelicolor is a particularly advantageous system to use to identify genes involved in bacterial cytokinesis because cell division, normally an essential process, is dispensable for growth and viability in this mycelial organism. The intent is to isolate critical genes that have not yet been identified in any prokaryotic organism. The products of genes identified by this project could be potential targets for novel compounds with biological activity. The need for such compounds is especially important today with the emergence of multiply drug-resistant pathogens. This project proposes to identify and characterize genes involved in cell division employing two mutagenesis and cloning strategies. In the first approach, division genes will be identified by chemical mutagenesis and cloned using ordered cosmid complementation libraries. In the second approach, division genes will be identified and cloned by insertional mutagenesis using a transposon. Immunofluorescence microscopy will be used to determine if the localization of FtsZ, a key division protein, is distributed in the new mutants. Genes that affect this early event are of particular interest because they have not been identified by any organism. Finally, experiments are proposed to analyze if the products of newly identified genes co-localize at the same subcellular position and pattern as FtsZ.
