Temporal changes in Hec1 phosphorylation control kinetochore–microtubule attachment stability during mitosis

Abstract

SummaryPrecise control of the attachment strength between kinetochores and spindle microtubules is essential to preserve genomic stability. Aurora B kinase has been implicated in regulating the stability of kinetochore-microtubule attachments but its relevant kinetochore targets in cells remain unclear. Here, we identify multiple serine residues within the N-terminus of the kinetochore protein Hec1 that are phosphorylated in an Aurora-B-kinase-dependent manner during mitosis. On all identified target sites, Hec1 phosphorylation at kinetochores is high in early mitosis and decreases significantly as chromosomes bi-orient. Furthermore, once dephosphorylated, Hec1 is not highly rephosphorylated in response to loss of kinetochore-microtubule attachment or tension. We find that a subpopulation of Aurora B kinase remains localized at the outer kinetochore even upon Hec1 dephosphorylation, suggesting that Hec1 phosphorylation by Aurora B might not be regulated wholly by spatial positioning of the kinase. Our results define a role for Hec1 phosphorylation in kinetochore-microtubule destabilization and error correction in early mitosis and for Hec1 dephosphorylation in maintaining stable attachments in late mitosis.

Minimal model for collective kinetochore–microtubule dynamics

“…4 B and C), consistent with previous in vivo results (10,18). Our finding is also consistent with experiments showing decreased oscillation speed and amplitude when phosphorylation by Aurora B is suppressed (16).…”

“…Properly attached kinetochores undergo center-of-mass (CM) and breathing oscillations that are regulated by collective MT dynamics (7)(8)(9)(10)(11)(12). Incorrect attachments-such as syntelic attachment of both kinetochores to the same pole-must be corrected (13)(14)(15)(16)(17). Tension may cue this process because bioriented kinetochore pairs are under tension while syntelically attached kinetochores are not (7,9,15,17,18).…”

“…Tension may cue this process because bioriented kinetochore pairs are under tension while syntelically attached kinetochores are not (7,9,15,17,18). Error correction is also mediated by Aurora B kinase phosphorylating MT-binding kinetochore proteins (13)(14)(15)(16)(17)(19)(20)(21). A consistent theory of metaphase kinetochore-MT dynamics should capture CM and breathing oscillations for correctly attached pairs and elucidate the contributions of tension and phosphorylation to syntelic error correction.…”

Pre-anaphase chromosome oscillations are regulated by the antagonistic activities of Cdk1 and PP1 on Kif18A

“…4b,c). To further confirm that the interaction between PP1 and Kif18A is critical for the dephosphorylation of Kif18A rather than for the transport of PP1 to KTs, we analysed Hec1, a KT component whose phosphorylation state is highly sensitive to the activities of Aurora-B and PP1 at KTs 26,27 . Phospho-specific antibodies demonstrated that expression of GFP-Kif18A PP1D in Kif18A-RNAi cells did not affect the phosphorylation levels of Hec1 at KTs (Fig.…”

Complications dawn for kinetochore regulation by Aurora

“…Another interesting suggestion is that Aurora B may play an important role in normal dynamics and alignment of chromosomes, as well as in error correction. Previous studies have found that mutating phosphorylation sites in the Hec1 N-terminal tail to nonphosphorylatable alanines causes defects in chromosome alignment (10) and suppresses chromosome oscillations about the spindle's equator (9). Inhibiting Aurora B also suppresses oscillations, even though chromosomes are properly attached (17).…”