CaMKIIγ-mediated inactivation of the Kin I kinesin MCAK is essential for bipolar spindle formation

Abstract

MCAK, a member of the kinesin-13 family, is a microtubule (MT) depolymerase that is necessary to ensure proper kinetochore MT attachment during spindle formation. Regulation of MCAK activity and localization is controlled in part by Aurora B kinase at the centromere. Here we analyzed human cells depleted of the ubiquitous Ca2+/calmodulin-dependent protein kinase IIγ isoform (CaMKIIγ) by RNA interference and found that CaMKIIγ was necessary to suppress MCAK depolymerase activity in vivo. A functional overlap with TOGp, a MT regulator known to counteract MCAK, was suggested by similar CaMKIIγ- and TOGp-depletion phenotypes, namely disorganized multipolar spindles. A replicating vector system, which permits inducible overexpression in cells that simultaneously synthesize interfering short hairpin RNAs, was used to dissect the functional interplay between CaMKIIγ, TOGp, and MCAK. Our results revealed two distinct but functionally overlapping mechanisms for negative regulation of the cytosolic/centrosomal pool of MCAK. These two mechanisms, involving CaMKIIγ and TOGp, respectively, are both essential for spindle bipolarity in a normal physiological context, but not in MCAK-depleted cells. © 2005 European Molecular Biology Organization | All Rights Reserved.