Abstract
The Tau family microtubule-associated proteins (MAPs) promote microtubule stabilization and regulate microtubule-based motility. They share the C-terminal microtubule-binding domain, which includes three to five tubulin-binding repeats. Different numbers of repeats formed by alternative splicing have distinct effects on the activities of these proteins, and the distribution of these variants regulates fundamental physiological phenomena in cells. In this study, using cryo-EM, we visualized the MAP4 microtubule complex with the molecular motor kinesin-1. MAP4 bound to the C-terminal domains of tubulins along the protofilaments stabilizes the longitudinal contacts of the microtubule. The strongest bond of MAP4 was found around the intertubulin-dimer interface such that MAP4 coexists on the microtubule with kinesin-1 bound to the intratubulin-dimer interface as well. MAP4, consisting of five repeats, further folds and accumulates above the intertubulin-dimer interface, interfering with kinesin-1 movement. Therefore, these cryo-EM studies reveal new insight into the structural basis of microtubule stabilization and inhibition of kinesin motility by the Tau family MAPs.