Cell division is one of the most fundamental events for self-reproduction of living organisms together with DNA-replication. However, it consists of a very complex system, which is coordinated with DNA replication, membrane biogenesis and various other cellular activities. Study of cell division and membrane proteins has broad implications for understanding not only the processes of normal growth of cells, but also those resulting in uncontrolled growth such as found in cancer. Our long term goal is to solve the molecular mechanism of cell division--precise sequence of events leading to cell division. However, because of its complexity, we will still concentrate our efforts to investigate individual key events such as the molecular mechanism of regulation of gene expression, and biosynthesis and assembly of membrane proteins in the present application. In particular, we will further explore a novel regulatory mechanism of gene expression by means of a small RNA molecule, mRNA interfering complementory RNA (micRNA), which was recently found in our laboratory. We will attempt to elucidate the precise molecular mechanism of the mic regulation and at the same time to apply this new concept to artificially regulate a specific gene in Escherichia coli as well as in yeast. This novel method to regulate a specific gene will provide a new, exciting approach for the study of many essential genes of unknown functions including those required for cell division. We will also continue to investigate the molecular regulatory mechanism of gene expression of the major outer membrane proteins, OmpA, ompC, and ompF. Second, we will continue to apply oligonucleotide-directed site-specific mutagenesis to the prolipoprotein signal peptide to elucidate the exact structural requirement of the signal peptide for protein secretion across the membrane. Third, we will explore to find components required for protein secretion (a) by characterizing the lepA gene product, which is known to be cosynthesized with signal peptidase I and (b) by isolating and characterizing the initial secretory complex formed in the cytoplasm in the case of some mutant prolipoproteins. Fourth, we will attempt to clone the gene for the new lipoproteins found in E. coli as well as the lipoprotein gene of Pseudomonas aeruginosa.