Project Summary/Abstract: Cellular membranes can be organized by the formation of discrete membrane microdomains (MMs), such as the cholesterol- and flotillin-rich “lipid rafts” found in eukaryotes. In bacteria, little is understood about the spatial organization of the cell membrane. Because this domain does not synthesize cholesterol or related eukaryotic lipids, it traditionally has been assumed that they do not contain raft-like compartments. Hopanoid lipids are the closest bacterial analogs of cholesterol, and my lab has found that they promote formation of MMs in the facultative endosymbiont Bradyrhizobium diazoefficiens. These MMs have similar biophysical properties as eukaryotic lipid rafts and are typically polarized, suggesting a link between raft-like MMs and cell polarity. Here, I propose to assess whether hopanoid-mediated MMs have similar compositions as eukaryotic lipid rafts and identify biological processes that are coordinated in these regions (Project 1). We also will examine cell cycle and polarity regulation in B. diazoefficiens, and whether it is affected by hopanoid-mediated MMs in both free-living and host-associated environments (Project 2). These projects will help discover fundamental paradigms of bacterial membrane organization that are likely to be shared with human-associated microbes, and may uncover new parallels between bacterial and eukaryotic subcellular organization.