Spindle integrity is critical for efficient mitotic progression and accurate chromosome segregation. Deregulation of spindles often leads to structural and functional aberrations, ultimately promoting segregation errors and aneuploidy, a hallmark of most human cancers. Here we report the characterization of a previously identified human sarcoma antigen (gene located at 19p13.11), Hice1, an evolutionarily nonconserved 46-kDa coiled-coil protein. Hice1 shows distinct cytoplasmic localization and associates with interphase centrosomes and mitotic spindles, preferentially at the spindle pole vicinity. Depletion of Hice1 by RNA interference resulted in abnormal and unstable spindle configurations, mitotic delay at prometaphase and metaphase, and elevated aneuploidy. Conversely, loss of Hice1 had minimal effects on interphase centrosome duplication. We also found that both full-length Hice1 and Hice1-N1, which is composed of 149 amino acids of the N-terminal region, but not the mutant lacking the N-terminal region, exhibited activities of microtubule bundling and stabilization at a near-physiological concentration. Consistently, overexpression of Hice1 rendered microtubule bundles in cells resistant to nocodazole- or cold-treatment-induced depolymerization. These results demonstrate that Hice1 is a novel microtubule-associated protein important for maintaining spindle integrity and chromosomal stability, in part by virtue of its ability to bind, bundle, and stabilize microtubules.

The mitotic spindle is constructed from microtubules (MTs) nucleated from centrosomes, chromosome proximal regions, and preexisting spindle MTs. Augmin, a recently identified protein complex, is a critical factor in spindle MT-based MT generation in Drosophila S2 cells. Previously, we identified one subunit of human augmin. Here, by using mass spectrometry, we identified the full human augmin complex of 8 subunits and show that it interacts with the gamma-tubulin ring complex (gamma-TuRC). Unlike augmin-depleted S2 cells, in which the defect in spindle-mediated MT generation is mostly compensated by centrosomal MTs, augmin knockdown alone in HeLa cells triggers the spindle checkpoint, reduces tension on sister kinetochores, and severely impairs metaphase progression. Human augmin knockdown also reduces the number of central spindle MTs during anaphase and causes late-stage cytokinesis failure. A link between augmin and gamma-TuRC is likely critical for these functions, because a gamma-TuRC mutant that attenuates interaction with augmin does not restore function in vivo. These results demonstrate that MT generation mediated by augmin and gamma-TuRC is critical for chromosome segregation and cytokinesis in human cells.

BACKGROUND: The assembly of a robust microtubule-based mitotic spindle is a prerequisite for the accurate segregation of chromosomes to progeny. Spindle assembly relies on the concerted action of centrosomes, spindle microtubules, molecular motors, and nonmotor spindle proteins. RESULTS: Here we use an RNA-interference screen of the human centrosome proteome to identify novel regulators of spindle assembly. One such regulator is HAUS, an 8-subunit protein complex that shares homology to Drosophila Augmin. HAUS localizes to interphase centrosomes and to mitotic spindle microtubules, and its disruption induces microtubule-dependent fragmentation of centrosomes along with an increase in centrosome size. HAUS disruption results in the destabilization of kinetochore microtubules and the eventual formation of multipolar spindles. These severe mitotic defects are alleviated by codepletion of NuMA, indicating that both factors regulate opposing activities. HAUS disruption alters NuMA localization, suggesting that mislocalized NuMA activity contributes to the spindle and centrosome defects observed. CONCLUSION: The human Augmin complex (HAUS) is a critical and evolutionary conserved multisubunit protein complex that regulates centrosome and spindle integrity.