CHARACTERIZATION OF HURP FUNCTION IN MITOSIS

Abstract

Chromosome biorientation and congression during mitosis require precise control of microtubule dynamics. The dynamics of kinetochore microtubules (K-MTs) are regulated by a variety of microtubule associated proteins (MAPs). Recently, a MAP known as HURP (hepatoma up-regulated protein) was identified. During mitosis, RanGTP releases HURP from the Importin ß inhibitory complex and allows it to localize to the kinetochore fiber (K-fiber). HURP bundles and stabilizes K-fibers and promotes chromosome congression. However, the mechanism underlying the role of HURP in regulating chromosome congression and proper mitotic progression remains elusive. This study investigates the role of HURP in mitosis by characterizing the domain function of HURP. By studying truncation and deletion mutant of HURP, we identify a functional microtubule binding domain, a Kif18A binding domain, an Importin ß regulatory domain and two D-boxes. We show that overexpression of the N-terminal microtubule binding domain (1-278 aa, HURP278) of HURP induces a series of mitotic defects including lagging chromosomes during the late prometaphase to metaphase stage, prolonged mitosis and increased kinetochore oscillation amplitude, mimicking the effects of Kif18A depletion. In addition, we identify Kif18A as a novel interaction partner of HURP. We reveal that HURP regulates Kif18A localization, movement and dynamics at the plus-end of K-MTs. The specificity of the effect of HURP on Kif18A is further confirmed by rescue experiment as the misaligned chromosomes in HURP278-overexpressing cells can be partially rescued by the overexpression of Kif18A. Our results demonstrate in part the regulatory mechanism for Kif18A at the K-MTs during chromosome congression and provide new insights into the role of HURP in regulating chromosome congression and mitotic progression.