Brun Cell division is a complex process that involves many proteins whose action must be coordinated with the DNA replication cycle. It is clear that an understanding of the timing of assembly of the different components of the division machinery and how and when the different components interact will be required to explain how division is controlled and how it is coordinated with the DNA replication cycle. This research will specifically examine the function of FtsA and its interaction with FtsZ. FtsZ is a tubulin-like GTPase that regulates the decision of when and where to divide. FtsZ is a highly conserved protein that polymerizes into a ring structure associated with the cytoplasmic membrane at the site of cell division. FtsZ recruits other cell division proteins to the site of cell division and may constrict, providing mechanical force for division. FtsA is localized to the division site in a FtsZ-dependent manner. FtsA is not required for the initiation of cytokinesis but is required for completion of cytokinesis. The long-term goal of this project is to understand the function of cell division proteins and to build a picture of protein-protein interactions in the cell division machinery as they occur during the cell cycle. This work will be performed in Caulobacter crescentus where each cell division gives rise to two morphologically and functionally different progeny cells: a motile swarmer cell and a sessile stalked cell. Caulobacter cells can be easily synchronized and specific morphological changes during the differentiation pathway provide a convenient indication of the stage of the cell cycle. Because there is only one DNA replication cycle per developmental cycle, events can be unambiguously assigned to a specific stage of the cell cycle. FtsZ and FtsA have been identified in Caulobacter and genetic interaction between the two proteins has already been established using the yeast two-hybrid system. The effect of mutations in different regions of FtsA will be analyzed to gain a bet ter understanding of its function and the domains that contribute to it. The domains of interaction between FtsZ and FtsA will be defined using the yeast two-hybrid system and fusions to glutathlone S-transferase. The localization of FtsA and FtsZ and their interaction will be analyzed during the cell cycle using immunofluorescence and in vivo cross-linking. This will allow a determination of the timing of the localization and the interaction of these two important cell division proteins within one cell cycle and its correlation with the stage of chromosome replication and the timing of nucleoid segregation. The mechanisms of cell division constitute one of the most fundamental problems in microbiology. This investigation will lead to a better understanding of cell division and will allow a picture of protein-protein interactions in the cell division machinery as they occur during the cell cycle to be established. A better understanding of the mechanisms of cell division will help the development of novel drug targets. The research will be done both in a research laboratory by senior investigators and in a new Independent Research in Molecular and Genetics course that will provide much needed independent research opportunities for a large number of undergraduate students. Students will learn about the different stages and aspects of scientific investigation by performing research on cell division and cell differentiation in Caulobacter and in sessions on critical analysis of the literature, scientific writing, oral presentation, and research ethics.
