Bacillus subtilis is a bacterium with a simple developmental pathway leading to spore formation under conditions of nutrient limitation. Shortly after the formation of the two cells required for spore formation, the membrane of the larger mother cell begins to migrate around the smaller forespore in a phagocytosis-like process known as engulfment. Ultimately the migrating membrane meets and fuses, releasing the forespore into the mother cell cytoplasm (Figure 1). My lab uses genetic and cell biological approaches to understand the mechanism of engulfment, as a model system for understanding how bacteria move macromolecules within their cells. We have developed methods for the study of engulfment that are ideal for the study of cellular polarity, protein localization and cell division.
Timelapse microscopy showing polar septation, chromosome segregation and engulfment of the smaller forespore by the larger mother cell.
Our research is focused on the following questions:
What is the mechanism of engulfment? We have identified two distinct protein complexes required for engulfment (Figure 2). The first is a mother cell anchored protein complex containing enzymes that hydrolyze the cell wall. The second is comprised of two proteins that make a zipper-like interaction across the septum and that together recruit all known proteins required for engulfment dependent gene expression. Our current studies are focused on understanding the molecular and biochemical mechanisms by which engulfment is mediated and by which the cell couples the completion of engulfment to gene expression.
How are membrane proteins localized within bacterial cells? Subcellular protein targeting is an essential feature of bacterial cells, as it is required for cell division, chromosome segregation, as well as for development. However, in contrast to eukaryotic cells, virtually nothing is known about the mechanism by which bacterial proteins reach their correct subcellular address. Our studies have identified two distinct mechanisms by which proteins localize during engulfment (Figure 2). We are also investigating the mechanism by which the assembly of the SpoIIIE DNA translocase is restricted to one side of the septum and the mechanism by which membrane proteins are inserted into the bilayer.
Fig. 2. Two pathways for protein localization during sporulation, the septal landmark pathway (A-C) and the protein-protein zipper (D-F).
How do bacteria catalyze membrane fission? The final step of engulfment is a membrane fission event that releases the forespore into the mother cell cytoplasm. We have identified a conserved bacterial protein required for this membrane fission event, the SpoIIIE DNA translocase that is required for the completion of chromosome segregation into the forespore (Figure 1).
How does FtsZ localize and move within bacterial cells? We are applying our cell biological methods to the study of the mechanism by which the tubulin-like protein FtsZ and key proteins that regulate its assembly localize and move within bacterial cells.
Blaylock, B., X. Jiang, A. Rubio, C.P. Moran, Jr. and K. Pogliano. (2004) Zipper-like interaction between proteins in adjacent daughter cells mediates protein localization. Genes and Development 18:2916-2928.
Liu, N.-J. L., R.A. Dutton and K. Pogliano. (2006) Evidence that the SpoIIIE DNA translocase participates in membrane fusion during cytokinesis and engulfment. Molecular Microbiology 59:1097-1113.
Broder, D. and K. Pogliano. (2006) Forespore engulfment mediated by a ratchet-like mechanism. Cell 126:917-928.
Chiba, S. K. Coleman and K. Pogliano. (2007) Impact of membrane fusion and proteolysis on SpoIIQ dynamics and interaction with SpoIIIAH. J. Biol. Chem. 282:2576-2586.
Aung, S., J. Shum, A. Abanes-De Mello, D. Broder, J. Fredlund-Gutierrez, S. Chiba and K. Pogliano. (2007) Dual localization pathways for the engulfment proteins during Bacillus subtilis sporulation. Molecular Microbiology 65:1534-1546.
Becker, E.C. and K. Pogliano. (2007) Cell-specific SpoIIIE assembly and DNA translocation polarity are dictated by chromosome orientation. Molecular Microbiology 66:1066-1079.
Ptacin, J., M. Nollmann, E.C. Becker, N.R. Cozzarelli, K. Pogliano and C. Bustamante. (2008) Sequence-directed DNA export guides chromosome translocation during sporulation in Bacillus subtilis. Nature Structural and Molecular Biology 15:485-493.
Gregory, J.A., E.C. Becker and K. Pogliano. (2008) Bacillus subtilis MinC destabilizes FtsZ-rings at new cell poles and contributes to the timing of cell division. Genes and Development, in press.
Kit Pogliano received her Ph.D. from the Department of Microbiology and Molecular Genetics at Harvard Medical School and was a Damon Runyon-Walter Winchell postdoctoral fellow at Harvard University. She is a recipient of the Searle Scholar and Beckman Young Investigator awards. Dr. Pogliano was named in 2009 as a Fellow in the American Academy of Microbiology.