Role of tau in the spatial organization of axonal microtubules: keeping parallel microtubules evenly distributed despite macromolecular crowding.

Opposing views have been proposed regarding the role of tau, the principal microtubule-associated protein in axons. On the one hand, tau forms cross-bridges at the interface between microtubules and induces microtubule bundling in neurons. On the other hand, tau is also considered a polymer brush which efficiently separates microtubules. In mature axons, microtubules are indeed arranged in parallel arrays and are well separated from each other. To reconcile these views, we developed a mechanistic model based on in vitro and cellular approaches combined to analytical and numerical analyses. The results indicate that tau forms long-range cross-bridges between microtubules under macromolecular crowding conditions. Tau cross-bridges prevent the redistribution of tau away from the interface between microtubules, which would have occurred in the polymer brush model. Consequently, the short-range attractive force between microtubules induced by macromolecular crowding is avoided and thus microtubules remain well separated from each other. Interestingly, in this unified model, tau diffusion on microtubules enables to keep microtubules evenly distributed in axonal sections at low tau levels.

Probing protein interactions in living mammalian cells on a microtubule bench.

Microtubules are µm-long cylinders of about 25 nm in diameter which are present in the cytoplasm of eukaryotic cells. Here, we have developed a new method which uses these cylindrical structures as platforms to detect protein interactions in cells. The principle is simple: a protein of interest used as bait is brought to microtubules by fusing it to Tau, a microtubule-associated protein. The presence of a protein prey on microtubules then reveals an interaction between bait and prey. This method requires only a conventional optical microscope and straightforward fluorescence image analysis for detection and quantification of protein interactions. To test the reliability of this detection scheme, we used it to probe the interactions among three mRNA-binding proteins in both fixed and living cells and compared the results to those obtained by pull-down assays. We also tested whether the molecular interactions of Cx43, a membrane protein, can be investigated with this system. Altogether, the results indicate that microtubules can be used as platforms to detect protein interactions in mammalian cells, which should provide a basis for investigating pathogenic protein interactions involved in human diseases.