All life demands the temporal and spatial control of essential biological functions. In bacteria, the recent discovery of coordinating elements provides a framework to begin to explain cell growth and division. Here we present the discovery of a supramolecular structure in the membrane of the coccal bacterium Staphylococcus aureus, which leads to the formation of a largescale pattern across the entire cell body; this has been unveiled by studying the distribution of essential proteins involved in lipid metabolism (PlsY and CdsA). The organization is found to require MreD, which determines morphology in rod-shaped cells. The distribution of protein complexes can be explained as a spontaneous pattern formation arising from the competition between the energy cost of bending that they impose on the membrane, their entropy of mixing, and the geometric constraints in the system. Our results provide evidence for the existence of a self-organized and nonpercolating molecular scaffold involving MreD as an organizer for optimal cell function and growth based on the intrinsic self-assembling properties of biological molecules.T he perpetuation of all cellular life requires the temporal and spatial management of essential biological functions, within the morphological framework characteristic of a specific organism. The underlying processes, which determine cell shape, are intimately intertwined with cell division and constitute pivotal issues for cell biology; their coordination in prokaryotes is mediated through counterparts of eukaryotic actin, tubulin, and intermediate filaments in addition to other specific components (1, 2). Several of these apparent cytoskeletal elements capitalize on their membrane binding properties, assembling along the longitudinal axis, between the poles of rod-shaped cells; they participate in many processes, including selection of the division site via the Min system and other components (3, 4); guidance and control of the cell wall biosynthetic machinery responsible for cell size, polarity, and shape through the actin-like protein MreB (5-11); and chromosome partitioning into daughter cells using another actin-like filament, ParM (12).Despite this set of highly coordinated mechanisms, it has recently been shown that otherwise rod-and cocci-shaped bacteria can exist as largely spherical wall-less forms known as L-forms, with the capacity to divide (13). Importantly, the division of L-forms of the rod-shaped bacterium Bacillus subtilis is freed from the requirement of the classical tubulin-like division component FtsZ (14). L-forms appear to divide by scission after blebbing, tabulation, or vesiculation dependent on an altered rate of membrane biosynthesis (15); this harks back perhaps to a more evolutionary primitive mechanism permitting cellular proliferation. Thus, are there underlying organizational mechanisms that exist, independent of apparent cytoskeletal elements? The fluid mosaic model proposing the free diffusion of membrane proteins through the lipids has been challenged by growing e...

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The bacterial cytoplasmic membrane is the interface between the cell and its environment, with multiple membrane proteins serving its many functions. However, how these proteins are organised to permit optimal physiological processes is largely unknown. Based on our initial findings that 2 phospholipid biosynthetic enzymes (PlsY and CdsA) localise heterogeneously in the membrane of the bacterium Staphylococcus aureus, we have analysed the localisation of other key membrane proteins. A range of protein fusions were constructed and used in conjunction with quantitative image analysis. Enzymes involved in phospholipid biosynthesis as well as the lipid raft marker FloT exhibited a heterogeneous localisation pattern. However, the secretion associated SecY protein, was more homogeneously distributed in the membrane. A FRET-based system also identified novel colocalisation between phospholipid biosynthesis enzymes and the respiratory protein CydB revealing a likely larger network of partners. PlsY localisation was found to be dose dependent but not to be affected by membrane lipid composition. Disruption of the activity of the essential cell division organiser FtsZ, using the inhibitor PC190723 led to loss of PlsY localisation, revealing a link to cell division and a possible role for FtsZ in functions not strictly associated with septum formation.

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SummaryAn intracellular approach for monitoring protein production in Staphylococcus aureus is described. mCherry, fused to the dodecapeptide Tip, was capable of inducing tetracycline repressor (TetR). Time‐ and concentration‐dependent production of mCherry could be correlated to TetR‐controlled GFPmut2 activity. This approach can potentially be extended to native S. aureus proteins.