Project summaryA cell is like a city, with an organized yet dynamic infrastructure grouped into specialties. For thelast 25 years, my lab has investigated how the simplest cells?bacteria?organize themselves anddivide to make progeny cells. We mainly focus on how bacteria such as E. coli achieve thedaunting task of splitting themselves in two at just the right time (once their genetic material isduplicated) and place (exactly in the middle) every 20 minutes without making errors. The keys tothis success are ancient and universal versions of protein polymers of actin (FtsA) and tubulin(FtsZ), which our lab visualized for the first time in living bacteria over 20 years ago. Today, we usestate of the art super-resolution imaging, combined with molecular genetics, protein biochemistry,interaction studies, and in vitro reconstitution, to gain more detailed insights into the structure andregulation of these cytoskeletal polymers and their associated proteins, which comprise thedynamic membrane-associated protein nanomachine (divisome) that divides bacterial cells.Thanks in part to our characterization of bypass suppressors of essential divisome proteins, it isnow becoming clear that the divisome is highly flexible, and can remodel itself in response tovarious inputs and perturbations. Despite impressive contributions by many labs, there is much tobe learned about overall divisome structure, the interchangeability of its parts, and how it remodelsin response to temporal and environmental cues. We will address these fundamental questions by(1) obtaining more high-resolution information about protein-protein contacts during cytokinesis bycombining biophysical, cytological, and genetic approaches; (2) investigating the role of oligomericstate of FtsZ and FtsA in divisome function and regulation, using super-resolution microscopy ofwhole cells and reconstituted biomimetic protein-membrane systems; (3) taking advantage of thediversity of divisome proteins from other model bacterial species to distinguish between commonand specialized mechanisms; (4) understanding the interplay between the divisome and otherlarge-scale cellular processes such as cell wall biosynthesis. We will leverage these approaches bycontinuing our collaborations with several close colleagues who have complementaryinterdisciplinary expertise.Our ongoing investigation of how the simplest cells divide should pave the way for anunprecedented understanding of how an entire cell functions and reproduces. Having an accuratemap of that city-cell's dynamic infrastructure will allow predictions to be made about how it works,and how to disrupt it.