An EB1-Binding Motif Acts as a Microtubule Tip Localization Signal

Abstract

Microtubules are filamentous polymers essential for cell viability. Microtubule plus-end tracking proteins (+TIPs) associate with growing microtubule plus ends and control microtubule dynamics and interactions with different cellular structures during cell division, migration, and morphogenesis. EB1 and its homologs are highly conserved proteins that play an important role in the targeting of +TIPs to microtubule ends, but the underlying molecular mechanism remains elusive. By using live cell experiments and in vitro reconstitution assays, we demonstrate that a short polypeptide motif, Ser-x-Ile-Pro (SxIP), is used by numerous +TIPs, including the tumor suppressor APC, the transmembrane protein STIM1, and the kinesin MCAK, for localization to microtubule tips in an EB1-dependent manner. Structural and biochemical data reveal the molecular basis of the EB1-SxIP interaction and explain its negative regulation by phosphorylation. Our findings establish a general "microtubule tip localization signal" (MtLS) and delineate a unifying mechanism for this subcellular protein targeting process.

Functional and Genetic Analysis of Spectraplakins in Drosophila

“…The Gas2-related domain (GRD) binds MTs and protects them against destabilization. The flexible, positively charged Ctail region associates with MTs and contains one or two SxIP sites required for binding to end-binding proteins (EB1) which is essential for axon growth (Alves-Silva et al, 2012;Applewhite et al, 2010;Honnappa et al, 2009;Sun, Leung, & Liem, 2001). The C-terminus can be phosphorylated to regulate MT association (Wu et al, 2011) and is the region specifically deleted in human BPAG1/dystonin mutations linked to neurodegeneration (Edvardson et al, 2012).…”

Role and regulation of kinesin-8 motors through the cell cycle

“…By contrast plus end accumulation of Kif18B to the tips of astral microtubules relies on its ability to bind EB1, an evolutionary conserved ?TIP protein that tracks microtubule plus tips by recognising the GTP cap, and potentially GDP.Pi tubulin subunits (Carlier et al 1989;Maurer et al 2011;Kumar and Wittmann 2012;Zanic et al 2009). Many EB1 binding proteins contain a conserved T/S-X-IP motif (Honnappa et al 2009) (where x is any amino acid (Jiang et al 2012) with a preference for lysine and arginine (Buey et al 2012)) that is essential for their ability to bind EB1 and track the tips of growing microtubules. Association of EB1 is conferred by the presence of several variants of this motif in the Kif18B tail domain (Tanenbaum et al 2011;Stout et al 2011).…”

“…Association of Kif18B to the plus tips of astral microtubule microtubules relies not only on EB1, but also on MCAK, a member of the kinesin-13 family, which is a non-motile microtubule depolymerase and an EB1-binding protein (Lee et al 2008;Honnappa et al 2009). MCAK mutants that fail to bind EB1 lead to a diminished level of Kif18B at the plus tip of astral microtubules.…”

Spectraplakin family proteins – cytoskeletal crosslinkers with versatile roles

“…MACF1 directly binds to EB1 (also known as MAPRE1) through its SxIP motif and exhibits MT plus-end tracking (Slep et al, 2005). The presence of SxIP motifs in the C-terminal region of BPAG1 controls its MT plus-end localization (Honnappa et al, 2009).…”

“…SxIP motifs exist in multiple MT plus-end-tracking proteins (+TIPs), including the spectraplakins MACF1 and BPAG1 (Akhmanova and Steinmetz, 2008;Gupta et al, 2010;Honnappa et al, 2009). MACF1 directly binds to EB1 (also known as MAPRE1) through its SxIP motif and exhibits MT plus-end tracking (Slep et al, 2005).…”