Functional Interaction of SKA and NDC80 Complexes at Kinetochores Promoting Anaphase Onset in Mitosis

Abstract

The kinetochore and spindle complex (SKA) and NDC80 complexes are essential kinetochore elements that ensure highly accurate chromosome segregation and successful progression through mitosis. The SKA heterodimer complex consists of SKA1, SKA2, and SKA3 subunits, and the NDC80 complex contains NDC80, NUF2, SPC24, and SPC25 subunits. Through live cell fluorescence timelapse imaging assays and expression of RNAi-resistant SKA3 constructs, we rescue SKA complex function in cells lacking endogenous SKA3. These assays reveal a critical span within SKA3s C-terminus required for successful mitotic progression. Structural protein modeling shows that this span encompasses the majority of a roughly 40 amino acid SKA3 C-terminal structural element that promotes interaction with the coiled-coil NDC80 and NUF2 subunits of the NDC80 complex. Thus, although spindle and kinetochore concentration of the SKA complex is mediated in part by the tubulin and tip-tracking capabilities provided by the SKA1 component of the SKA complex, transition from metaphase to anaphase requires the contribution of SKA3s C-terminal structural interface to mediate interaction between the SKA and NDC80 complexes. Significance StatementAccurate chromosome segregation is essential for genomic stability, and its failure underlies developmental defects, and cancer. The kinetochore-microtubule interface, where the SKA and NDC80 complexes converge, is central, yet the of these two complexes is incompletely defined. We identify a critical segment within the C-terminus of SKA3 required for the metaphase-to-anaphase transition. Using live-cell imaging with RNAi-resistant rescue constructs and structural modeling, we demonstrate that this region mediates SKA engagement with the coiled-coil domains of NDC80 and NUF2. Our findings establish that a small segment of SKA3s C-terminus provides an essential physical bridge between the SKA and NDC80 complexes. This work refines understanding kinetochore-microtubule interaction, functionally identifying a discrete contact whose disruption may be relevant to chromosomal instability in disease.