title: Cardiolipin microdomains localize to negatively curved regions of Escherichia coli membranes.
abstract: Many proteins reside at the cell poles in rod-shaped bacteria. Several hypotheses have drawn a connection between protein localization and the large cell-wall curvature at the poles. One hypothesis has centered on the formation of microdomains of the lipid cardiolipin (CL), its localization to regions of high membrane curvature, and its interaction with membrane-associated proteins. A lack of experimental techniques has left this hypothesis unanswered. This paper describes a microtechnology-based technique for manipulating bacterial membrane curvature and quantitatively measuring its effect on the localization of CL and proteins in cells. We confined Escherichia coli spheroplasts in microchambers with defined shapes that were embossed into a layer of polymer and observed that the shape of the membrane deformed predictably to accommodate the walls of the microchambers. Combining this technique with epifluorescence microscopy and quantitative image analyses, we characterized the localization of CL microdomains in response to E. coli membrane curvature. CL microdomains localized to regions of high intrinsic negative curvature imposed by microchambers. We expressed a chimera of yellow fluorescent protein fused to the N-terminal region of MinD--a spatial determinant of E. coli division plane assembly--in spheroplasts and observed its colocalization with CL to regions of large, negative membrane curvature. Interestingly, the distribution of MinD was similar in spheroplasts derived from a CL synthase knockout strain. These studies demonstrate the curvature dependence of CL in membranes and test whether these structures participate in the localization of MinD to regions of negative curvature in cells.
authors: Department of Biochemistry and Biomedical Engineering, University of Wisconsin, Madison, WI 53706, USA.
authors: Lars D
authors: LD
authors: Renner
authors: Douglas B
authors: DB
authors: Weibel
mesh_terms: D000251:Adenosine Triphosphatases / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D002308:Cardiolipins / Q000737:chemistry / Q000378:metabolism*; D002455:Cell Division; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D029968:Escherichia coli Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D055786:Gene Knockout Techniques; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D021962:Membrane Microdomains / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D008565:Membrane Proteins / Q000235:genetics; D008856:Microscopy, Fluorescence / Q000379:methods*; D008858:Microscopy, Phase-Contrast / Q000379:methods*; D013104:Spheroplasts / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D002308:Cardiolipins; D029968:Escherichia coli Proteins; D008164:Luminescent Proteins; D008565:Membrane Proteins; C054562:yellow fluorescent protein, Bacteria; D017855:Transferases (Other Substituted Phosphate Groups); C100091:cardiolipin synthetase; D000251:Adenosine Triphosphatases; C072069:MinD protein, E coli