RBM14 prevents assembly of centriolar protein complexes and maintains mitotic spindle integrity.
Bottom Line: Here, we identify RBM14 as a novel suppressor of assembly of centriolar protein complexes.Intriguingly, the formation of such structures seems not to require the cartwheel structure that normally acts as a scaffold for centriole formation, whereas they can retain pericentriolar material and microtubule nucleation activity.We further demonstrate that such structures assemble in the cytoplasm even in the presence of pre-existing centrioles.
Affiliation: Centrosome Biology Laboratory, Center for Frontier Research, National Institute of Genetics, Mishima Shizuoka, Japan.Show MeSH
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Mentions: Since the ectopic centriolar protein complexes induced by RBM14 depletion could recruit PCM components (Fig 1E), we next tested whether they are potent to nucleate microtubules. Given that formation of the centriolar protein complexes frequently resulted in pseudo-bipolar spindle formation in which an abnormal number of centriolar protein foci clustered into spindle poles, we sought to establish an experimental setup in which it is possible to decluster the centriolar protein complexes and pre-existing centrioles and evaluate their ability to nucleate microtubules in mitosis. To this end, we carried out a microtubule regrowth assay using mitotic U2OS cells (Fig 5A). As expected, in control mitotic cells, we detected efficient microtubule regrowth and subsequent spindle formation within 30 min after cold treatment (Fig 5B). Importantly, we found that, upon RBM14 depletion, the ectopic centriolar protein complexes with PCM cloud formed extra spindle poles, which often induced formation of multipolar spindles at 30 min after cold treatment (Fig 5C–E). 46 ± 5% of the centriolar protein complexes appeared to harbor microtubule nucleation activity in mitosis (n = 150 in triplicate). However, we observed that extra small spindle poles in RBM14-depleted cells tended to be clustered into a pseudo-bipolar spindle assay at 60 min after cold treatment (Fig 5C). Taken together, these results indicate that the ectopic centriolar protein complexes induced by RBM14 depletion are at least in part potent to nucleate microtubules.
Affiliation: Centrosome Biology Laboratory, Center for Frontier Research, National Institute of Genetics, Mishima Shizuoka, Japan.