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Over the past few years we have developed an image-processing framework that allows us to extract precise 3D shapes of bacterial cells from fluorescence microscopy data. This active mesh approach minimizes the difference between an observed Z-stack and model shapes that have been convolved with the experimental point spread function. From these xyz coordinates, we calculate geometric parameters such as local curvatures, surface areas, and the relative enrichment of fluorescent signals. Our fluorescent signal is the concentration of the bacterial actin homolog MreB. We have integrated a fluorescent sandwich fusion of MreB with msfGFP at the native locus. This construct has unperturbed mass-doubling times and a proper rod-like shape.We have previously shown in both two and three dimensions that MreB is enriched at negative curvatures and away from positive curvatures [Ursell et al. PNAS 2014]. Here we show in the straight rod Escherichia coli and the curved rod Caulobacter crescentus that MreB senses local surface curvature and, as a result, helps ensure rod-like growth of the cell. The curvature sensed by Escherchia coli MreB is the Gaussian curvature of the surface, the product of the two principal curvatures. Using MreB point mutants that have altered curvature sensitivities, we are testing the hypothesis that cells straighten deformations by patterning growth at the proper geometry. To stably generate their comma shaped geometry, Caulobacter crescentus have developed ways to overcome this straightening mechanism. The enrichment profile of CcMreB shows a plateau in enrichment at small, positive Gaussian curvature values that depends on the protein crescentin. In addition to its structural role in bending the cell, we propose a mechanism whereby crescentin alters the curvature preference of MreB and allows the stabile propagation of the Caulobacter shape. |
