The mammalian kinetochore independently regulates its passive and active force-generating interfaces with microtubules

Abstract

The kinetochore links chromosomes to dynamic spindle microtubules and drives both chromosome congression and segregation. To do so, the kinetochore must hold on to depolymerizing and polymerizing microtubules. At metaphase, one sister kinetochore couples to depolymerizing microtubules, pulling its sister along polymerizing microtubules [1,2]. Distinct kinetochore-microtubule interfaces mediate these behaviors: active interfaces transduce microtubule depolymerization into mechanical work, and passive interfaces generate friction as the kinetochore slides along microtubules [3,4]. We do not know the physical and molecular nature [5-7] of these interfaces, or how they are regulated to support diverse mitotic functions in mammalian cells. To address this question, we focus on the mechanical role of the essential load-bearing protein Hec1 [8-11]. Hec1s affinity for microtubules is regulated by Aurora B phosphorylation on its N-terminal tail [12-15], but its role at the passive and active interfaces remains unclear. Here, we use laser ablation to trigger cellular pulling on mutant kinetochores and decouple sisters in vivo, and thereby separately probe Hec1s role as it moves on polymerizing versus depolymerizing microtubules. We show that Hec1 phosphorylation tunes passive friction along polymerizing microtubules, modulating both the magnitude and timescale of responses to force. In contrast, we find that Hec1 phosphorylation does not affect the kinetochores ability to grip depolymerizing microtubules, or switch to this active force-generating state. Together, the data suggest that different kinetochore interfaces engage with growing and shrinking microtubules, and that passive friction can be regulated without disrupting active force generation. Through this mechanism, the kinetochore can modulate its grip on microtubules as its functional needs change during mitosis, and yet retain its ability to couple to microtubules powering chromosome movement.