Nek2 is a mitotic kinase whose activity varies during the cell cycle. It is well known that Nek2 is involved in centrosome splitting, and a number of studies have indicated that Nek2 is crucial for maintaining the integrity of centrosomal structure and microtubule nucleation activity. In the present study, we report that NIP2, previously identified as centrobin, is a novel substrate of Nek2. NIP2 was daughter-centriole-specific, but was also found in association with a stable microtubule network of cytoplasm. Ectopic NIP2 formed aggregates but was dissolved by Nek2 into small pieces and eventually associated with microtubules. Knockdown of NIP2 showed significant reduction of microtubule organizing activity, cell shrinkage, defects in spindle assembly and abnormal nuclear morphology. Based on our results, we propose that NIP2 has a role in stabilizing the microtubule structure. Phosphorylation may be crucial for mobilization of the protein to a new microtubule and stabilizing it.
SummaryPericentriolar satellites are electron-dense granules that are concentrated around the centrosome. They are involved in the recruitment of centrosomal proteins and microtubule organization in interphase cells, but their mitotic functions are largely unknown. In this study, we characterize CEP90 as a component of pericentriolar satellites. CEP90 is present both in the centrosome and in the cytoplasm, but is transiently concentrated at the centrosome once cells enter mitosis. Depletion of CEP90 caused mitotic arrest with misaligned chromosomes. Spindle pole fragmentation was the most characteristic phenotype in CEP90-depleted cells. Spindle poles were fragmented as soon as the spindles attached, suggesting that the mechanical forces of spindle microtubules physically stress the structure of CEP90-depleted spindle poles. Based on these results, we propose that CEP90 is crucial for maintaining the integrity of spindle poles during mitosis.
Centriolar satellites are PCM-1-positive granules surrounding centrosomes. Proposed functions of the centriolar satellites include protein targeting to the centrosome, as well as communication between the centrosome and surrounding cytoplasm. CEP90 is a centriolar satellite protein that is critical for spindle pole integrity in mitotic cells. In this study, we examined the biological functions of CEP90 in interphase cells. CEP90 physically interacts with PCM-1 at centriolar satellites, and this interaction is essential for centrosomal accumulation of the centriolar satellites and eventually for primary cilia formation. CEP90 is also required for BBS4 loading on centriolar satellites and its localization in primary cilia. Our results imply that the assembly and transport of centriolar satellites are critical steps for primary cilia formation and ciliary protein recruitment.
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