A model of the microtubule cytoskeleton of the axon initial segment in mixtures of tubulin, tau, and GTP.

This review is part of a special memorial issue in honor of Erich Sackmann's remarkable research career, in particular on the biophysics of equilibrium and living membranes. Erich's large body of discoveries firmly established him as a giant in the field of biophysics. In our review, we describe structural studies of microtubules (MTs) in the presence of the microtubule-associated protein tau, an intrinsically disordered protein confined to the axon of mature neurons in vertebrates. We start with a brief review of experiments where the inherently dynamical MT is fixed with the cancer chemotherapy drug molecule paclitaxel. At a paclitaxel/tubulin-dimer molar ratio of 1, tau is found to modulate the protofilament number of MTs with a tendency to increase the MT diameter. Under these high paclitaxel/tubulin molar ratios, where a vast amount of comprehensive literature on the high-resolution structure of tubulin and MTs exists, tau does not mediate bundling interactions between MTs. The review further describes more recent structural studies, which combine synchrotron small-angle x-ray scattering (SAXS) with electron microscopy of plastic-embedded preparations. The studies focus on elucidating the steady-state assembled structures emerging from reaction mixtures of tubulin, tau, and GTP at 37°C, all in the "absence" of paclitaxel. The mixtures contained either 1 millimolar (mM) Mg2+ present in the buffer or up to an additional few mM of Mg2+ comparable to cellular concentrations. The latter mixtures are intended to provide samples for studies to clarify the effects of cellular metal ions on tubulin/tau assemblies. The combined paclitaxel-free studies represent a minimal cell-free model of the MT cytoskeleton found in the central core of the axon initial segment of mature neurons. SAXS and TEM revealed out-of-equilibrium steady-state structures with hexagonal and linear symmetries (in cross section) and large MT wall-to-wall spacing greater than the diameter of MTs. The spacings and "linear MT architecture" resemble the morphology of MT fascicles found in the axon initial segment. SAXS of reaction mixtures combined with TEM imaging of plastic-embedded samples provided evidence that microtubules are bundled and stabilized by an "intervening network" of complexes of tubulin oligomers and tau.