Ethan Garner, Harvard, Watching Spheres Convert To Rods: Dissecting The Regulation And Spatial Organization Of Bacterial Cell Wall Synthesis
Short reception follows seminar
Rod shaped bacteria elongate by the action of cell-wall synthetic complexes that move circumferentially around the cell width. These directional motions are thought to reflect the insertion of new cell wall material as motions stop upon addition of cell wall inhibiting antibiotics. These synthetic complexes on the outside of the cell are linked to MreB filaments bound to the cytoplasmic surface of the membrane. Each filament/enzyme complex moves around the rod independently, with adjacent complexes moving in opposing directions. We work to understand how the nanometer scale synthetic activity of these filament/enzyme complexes, in combination with other proteins, create the micron scale order of rod shaped bacteria. We are currently focused on understanding the regulation, formation, and orientation of motion of MreB.
1) We have been working to understand how bacteria regulate the assembly of MreB. We have created strains where all expressed MreB proteins are fluorescently tagged at the native locus. These strains show no defects on growth rate, suggesting complete complementation and functionality. High-resolution imaging and single molecule tracking indicates that MreB exists in vivo as a discrete number of very short filaments (~220nm) above a diffusing pool of cytoplasmic MreB. We find that the cell regulates the membrane association of MreB. Membreane association of MreB is is controlled by the amount of lipid II, the membrane bound precursor used to build new cell wall. Our data suggests that there is feedback between the metabolic flux within the cell and the number of active enzymatic complexes.
2) To understand how the discrete activities of these disconnected, independent enzymes create rod shaped cells, we observe the dynamics of MreB filaments as we deform and reform cells from rods into spheres. While the directional motions of MreB are radially organized in rods, motions become isotropic in spherical cells. Radial organization falls apart beneath a given aspect ratio, indicating this system can sense the short axis of the cell. Spheres convert into rods by directly emitting rods of the correct width, suggesting that this machinery is tuned to both sense and propagate a given cellular width. These emerging rods are only maintained as long as the cell wall is rigid enough to support the deformation. Furthermore, we can convert and maintain spherical cells into rods by confining their geometry. These results suggest that the elongation machinery encodes an intrinsic sensor of cell width, orienting their motion of activity along a given curvature. This property allows feedback between the synthesis machinery and the pre-existing macro scale shape, independent of any previous cell wall material.
Location: Carl Icahn Lab 101
Date/Time: 03/24/14 at 4:15 pm - 03/24/14 at 5:15 pm
Category: Home & Away
Department: Lewis-Sigler Institute