Centrosomal nucleolin is required for microtubule network organization

Nucleolin is a pleiotropic protein involved in a variety of cellular processes. Although multipolar spindle formation has been observed after nucleolin depletion, the roles of nucleolin in centrosome regulation and functions have not been addressed. Here we report using immunofluorescence and biochemically purified centrosomes that nucleolin co-localized only with one of the centrioles during interphase which was further identified as the mature centriole. Upon nucleolin depletion, cells exhibited an amplification of immature centriole markers surrounded by irregular pericentrin staining; these structures were exempt from maturation markers and unable to nucleate microtubules. Furthermore, the microtubule network was disorganized in these cells, exhibiting frequent non-centrosomal microtubules. At the mature centriole a reduced kinetics in the centrosomal microtubule nucleation phase was observed in live silenced cells, as well as a perturbation of microtubule anchoring. Immunoprecipitation experiments showed that nucleolin belongs to protein complexes containing 2 key centrosomal proteins, γ-tubulin and ninein, involved in microtubule nucleation and anchoring steps. Altogether, our study uncovered a new role for nucleolin in restricting microtubule nucleation and anchoring at centrosomes in interphase cells.

核仁素（nucleolin）是一种参与多种细胞过程的多效性蛋白质。尽管在核仁素耗竭后已观察到多极纺锤体形成，但核仁素在中心体调控及其功能中的作用尚未被阐明。本研究通过免疫荧光技术与生化纯化的中心体实验发现，核仁素仅在间期与其中一个中心粒共定位，该中心粒经进一步鉴定为成熟中心粒。在核仁素耗竭后，细胞会出现未成熟中心粒标志物扩增的现象，且这些未成熟中心粒周围伴有不规则的周质蛋白（pericentrin）染色；此类结构不携带成熟中心粒标志物，也无法启动微管成核。此外，这些细胞内的微管网络发生紊乱，频繁出现非中心体依赖的微管。在经核仁素沉默的活细胞中，可观察到成熟中心粒的中心体微管成核阶段动力学速率降低，同时微管锚定功能也受到干扰。免疫沉淀实验表明，核仁素属于包含两种关键中心体蛋白——γ微管蛋白（γ-tubulin）与九蛋白（ninein）——的蛋白质复合物，这两种蛋白分别参与微管成核与锚定过程。综上，本研究揭示了核仁素在间期细胞中限制中心体处微管成核与锚定的全新功能。