UC Berkeley

Cytoplasmic Linker Associated Proteins (CLASPs) comprise a class of microtubule (MT) plus end-binding proteins (+Tips) that contribute to the dynamics and organization of MTs during many cellular processes, including migration and division. Human CLASP contains multiple MT-binding domains, including Tumor Overexpressed Gene (TOG), as well as Ser-x-Ile-Pro (SxIP) motifs known to target some +Tips though interaction with End-Binding Protein 1 (EB1), but how individual domains contribute to CLASP function is poorly understood. We generated full-length recombinant human CLASP and a series of truncation mutants and found that the two N-terminal TOG domains make the strongest contribution to MT polymerization and bundling and mediate binding to the MT lattice, while plus end tracking requires interaction with EB1. The C-terminal coiled-coil mediates dimerization and association with many other factors, including kinetochore motor CENP-E, while other regions near the S/R region interact with the spindle mid-zone protein PRC1 and chromokinesin Xkid. Importantly, only the full-length protein was able to rescue spindle assembly in Xenopus egg extracts depleted of endogenous CLASP. Even minor amounts of CLASP that was missing its C-terminal domain, led to dramatic spindle phenotypes, indicating that proper localization of CLASP is essential for its activity during mitosis.