Aurora-B kinase pathway controls the lateral to end-on conversion of kinetochore-microtubule attachments in human cells

Abstract

Human chromosomes are captured along microtubule walls (lateral attachment) and then tethered to microtubule-ends (end-on attachment) through a multi-step end-on conversion process. Upstream regulators that orchestrate this remarkable change in the plane of kinetochore-microtubule attachment in human cells are not known. By tracking kinetochore movements and using kinetochore markers specific to attachment status, we reveal a spatially defined role for Aurora-B kinase in retarding the end-on conversion process. To understand how Aurora-B activity is counteracted, we compare the roles of two outer-kinetochore bound phosphatases and find that BubR1-associated PP2A, unlike KNL1-associated PP1, plays a significant role in end-on conversion. Finally, we uncover a novel role for Aurora-B regulated Astrin-SKAP complex in ensuring the correct plane of kinetochore-microtubule attachment. Thus, we identify Aurora-B as a key upstream regulator of end-on conversion in human cells and establish a late role for Astrin-SKAP complex in the end-on conversion process.

PP1 and PP2A use opposite phospho-dependencies to control distinct processes at the kinetochore

“…As shown previously by others (Shrestha et al, 2017), removal of the PP2A-B56 SLiM in BubR1 (BubR1 PP2A ) causes severe defects in chromosome alignment, whereas inactivation of the PP1 SLiM in Knl1 (Knl1 PP1 ) does not ( fig.1a). These defects are associated with enhanced phosphorylation of the Ndc80 tail region ( fig.1b and supp.…”

Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments

“…This raises the question of how and when Polo activity and Ser499 phosphorylation are antagonized to allow timely formation of stable end‐on attachments. PP2A‐B56 phosphatase may have a role in this process, since impairing its association with BubR1 was recently shown to dramatically increase the frequency of laterally attached KTs in human cells (Shrestha et al , ). However, because BubR1‐PP2A‐B56 is already present at high levels on early mitotic KTs, we reason that additional mechanisms must operate to prevent premature end‐on conversion (Moura & Conde, ).…”

Molecular determinants of the Ska-Ndc80 interaction and their influence on microtubule tracking and force-coupling

“…Upon entry into M-phase (mitosis or meiosis) and spindle assembly, chromosomes are initially transported to the spindle poles, where the microtubule density is highest, and from there to the spindle's equatorial plane, forming lateral attachments to the microtubule lattice. CENP-E, a kinetochore-localized, microtubule-plus-end-directed motor plays an essential function in this process (Bancroft et al, 2015;Barisic et al, 2014;Cai et al, 2009;Chakraborty et al, 2019;Kapoor et al, 2006;Kim et al, 2008;Kitajima et al, 2011;Magidson et al, 2011;Shrestha et al, 2017;Tanaka et al, 2005). In a poorly understood process of "endconversion", kinetochores engage the microtubule-binding interface of the KMN network and transition from binding to the lattice to binding to the dynamic plus ends of the microtubules, which become embedded into the kinetochore's outer plate (Dong et al, 2007;Kuhn and Dumont, 2017;McIntosh et al, 2013;Wan et al, 2009).…”

“…At one end of Ndc80, two closely interacting calponin-homology (CH) domains near the N-terminal ends of NDC80 and NUF2 form a globular structure that binds the microtubule. An ~80-residue basic tail preceding the NDC80 CH domain (Ndc80-tail) has also been implicated in microtubule binding, and phosphorylation by Aurora kinase activity has been proposed to modulate electrostatic interactions with the negatively charged MT lattice (Alushin et al, 2012;Cheerambathur et al, 2017;Cheeseman et al, 2002;Cheeseman et al, 2006;Ciferri et al, 2008;DeLuca et al, 2006;DeLuca et al, 2011;DeLuca et al, 2018;Guimaraes et al, 2008;Long et al, 2017;Miller et al, 2008;Shrestha et al, 2017;Tooley et al, 2011;Umbreit et al, 2012;Wei et al, 2007;Ye et al, 2015;Zaytsev et al, 2015;Zaytsev et al, 2014).…”