Contact-dependent cell-cell signaling is essential for development of organisms from prokaryotes to humans though the mechanisms are poorly understood. The objective of this project is to examine the molecular basis of C-signaling, which is essential for fruiting body development of Myxococcus xanthus. C-signaling is initiated by contact between cells and leads to changes in gene expression that mediate multicellular morphogenesis. CsgA, the only known protein involved in C-signal production, contains consensus motifs found in the short chain alcohol dehydrogenase (SCAD) family. Serine appears to be a substrate for the enzyme. It is likely that either CsgA or its enzymatic product is transferred between cells to initiate C-signaling, but the compartmentalization of the signaling components remains unclear. The enzymology of CsgA will be examined with regard to the range of substrates utilized. The chemical product(s) of each reaction will be assayed, identified, and purified. The biological activity of the products will be examined to determine whether they rescue development of csgA mutants. Finally, NAD+ analogs that inhibit CsgA enzyme activity in vitro will be used to determine whether CsgA enzyme activity is essential for C-signaling in vivo. The subcellular location of CsgA and its interaction with other proteins will be determined by biochemical and genetic approaches including inner and outer membrane separation, microscopy, and the use of the yeast two hybrid system to identify interacting proteins. Finally, a model will be developed to explain C-signaling. This project is expected to advance our knowledge in several important areas including prokaryotic development and contact-dependent cell interactions. The project will also result in laboratory training of graduate and undergraduate students, some of whom will be minority and female students. The results will be disseminated through presentations at meetings and publication in journals. The major benefits will be an improvement in the scientific literacy of a segment of the population, and an improved understanding of prokaryotic biology and diversity.
