Mechanisms of spindle-pole organization are influenced by kinetochore activity in mammalian cells

The spindle is a fusiform bipolar-microtubule array that is responsible for chromosome segregation during mitosis [1-4]. Focused poles are an essential feature of spindles in vertebrate somatic cells, and pole focusing has been shown to occur through a centrosome-independent self-organization mechanism where microtubule motors cross-link and focus microtubule minus ends [4-11]. Most of our understanding of this mechanism for pole focusing derives from studies performed in cell-free extracts devoid of centrosomes and kinetochores [5, 7, 9, 10]. Here, we examine how sustained force from kinetochores influences the mechanism of pole focusing in cultured cells. We show that the motor-driven self-organization activities associated with NuMA (i.e., cytoplasmic dynein) and HSET are not necessary for pole focusing if sustained force from kinetochores is inhibited in Nuf2- or Mis12-deficient cells. Instead, pole organization relies on TPX2 as it cross-links spindle microtubules to centrosome-associated mitotic asters. Thus, both motor-driven and static-cross-linking mechanisms contribute to spindle-pole organization, and kinetochore activity influences the mechanism of spindle-pole organization. The motor-driven self-organization of microtubule minus ends at spindle poles is needed to organize spindle poles in vertebrate somatic cells when kinetochores actively exert force on spindle microtubules.