EB1-binding–myomegalin protein complex promotes centrosomal microtubules functions

Significance Microtubule dynamics is tightly regulated during fundamental biological processes such as mitosis, thereby representing a major target for anticancer therapies. To better understand the molecular mechanisms underlying the organization of the microtubule network, we systematically investigated proteins interacting with EB1, a major regulator of microtubules dynamics. We identified a specific isoform of myomegalin, which we termed “SMYLE,” that assembles a macromolecular complex associated with the centrosome, the major microtubule-organizing center in cells, and also connected to the microtubule nucleating complex. SMYLE promoted microtubule assembly from the centrosome and subsequent stabilization of microtubules at the cell periphery. This had consequences on cell motility, mitosis, and cell-cycle progression, suggesting that SMYLE might be an important player in tumor progression. Control of microtubule dynamics underlies several fundamental processes such as cell polarity, cell division, and cell motility. To gain insights into the mechanisms that control microtubule dynamics during cell motility, we investigated the interactome of the microtubule plus-end–binding protein end-binding 1 (EB1). Via molecular mapping and cross-linking mass spectrometry we identified and characterized a large complex associating a specific isoform of myomegalin termed “SMYLE” (for short myomegalin-like EB1 binding protein), the PKA scaffolding protein AKAP9, and the pericentrosomal protein CDK5RAP2. SMYLE was associated through an evolutionarily conserved N-terminal domain with AKAP9, which in turn was anchored at the centrosome via CDK5RAP2. SMYLE connected the pericentrosomal complex to the microtubule-nucleating complex (γ-TuRC) via Galectin-3–binding protein. SMYLE associated with nascent centrosomal microtubules to promote microtubule assembly and acetylation. Disruption of SMYLE interaction with EB1 or AKAP9 prevented microtubule nucleation and their stabilization at the leading edge of migrating cells. In addition, SMYLE depletion led to defective astral microtubules and abnormal orientation of the mitotic spindle and triggered G1 cell-cycle arrest, which might be due to defective centrosome integrity. As a consequence, SMYLE loss of function had a profound impact on tumor cell motility and proliferation, suggesting that SMYLE might be an important player in tumor progression.