Reconstitution of a hierarchical +TIP interaction network controlling microtubule end tracking of dynein.

Growing microtubule end regions recruit a variety of proteins collectively termed +TIPs, which confer local functions to the microtubule cytoskeleton. +TIPs form dynamic interaction networks whose behaviour depends on a number of potentially competitive and hierarchical interaction modes. The rules that determine which of the various +TIPs are recruited to the limited number of available binding sites at microtubule ends remain poorly understood. Here we examined how the human dynein complex, the main minus-end-directed motor and an important +TIP (refs , , ), is targeted to growing microtubule ends in the presence of different +TIP competitors. Using a total internal reflection fluorescence microscopy-based reconstitution assay, we found that a hierarchical recruitment mode targets the large dynactin subunit p150Glued to growing microtubule ends via EB1 and CLIP-170 in the presence of competing SxIP-motif-containing peptides. We further show that the human dynein complex is targeted to growing microtubule ends through an interaction of the tail domain of dynein with p150Glued. Our results highlight how the connectivity and hierarchy within dynamic +TIP networks are orchestrated.