Formation of a single new centriole from a pre-existing centriole is strictly controlled to maintain correct centrosome number and spindle polarity in cells. However, the mechanisms that govern this process are incompletely understood. Here, using several human cell lines, immunofluorescence and structured illumination microscopy methods, and ubiquitination assays, we show that the E3 ubiquitin ligase F-box and WD repeat domain containing 7 (FBXW7), a subunit of the SCF ubiquitin ligase, down-regulates spindle assembly 6 homolog (HsSAS-6), a key protein required for pro-centriole cartwheel assembly, and thereby regulates centriole duplication. We found that FBXW7 abrogation stabilizes HsSAS-6 and increases its recruitment to the mother centriole at multiple sites, leading to supernumerary centrioles. Ultra-structural analyses revealed that FBXW7 is broadly localized on the mother centriole and that its presence is reduced at the site where the HsSAS-6-containing pro-centriole is formed. This observation suggested that FBXW7 restricts pro-centriole assembly to a specific site to generate a single new centriole. In contrast, during HsSAS-6 overexpression, FBXW7 strongly associated with HsSAS-6 at the centriole. We also found that SCFFBXW7 interacts with HsSAS-6 and targets it for ubiquitin-mediated degradation. Further, we identified putative phosphodegron sites in HsSAS-6, whose substitutions rendered it insensitive to FBXW7-mediated degradation and control of centriole number. In summary, SCFFBXW7 targets HsSAS-6 for degradation and thereby controls centriole biogenesis by restraining HsSAS-6 recruitment to the mother centriole, a molecular mechanism that controls supernumerary centrioles/centrosomes and the maintenance of bipolar spindles.

中文翻译：

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泛素连接酶FBXW7靶向中心粒装配蛋白HsSAS-6降解，从而调节中心粒重复。

严格控制从先前存在的中心粒形成单个新的中心粒，以维持细胞中正确的中心粒数和纺锤体极性。但是，控制此过程的机制尚未完全理解。在这里，我们使用几种人类细胞系，免疫荧光和结构照明显微镜方法以及泛素化测定法，发现E3泛素连接酶F-box和WD重复域包含7（FBXW7）（SCF泛素连接酶的亚基）下调。纺锤体装配6同源物（HsSAS-6），前中心轮装配所需的关键蛋白，从而调节中心体重复。我们发现FBXW7废除稳定了HsSAS-6，并在多个位置增加了其对母体的募集，从而导致了多余的母体。超结构分析表明，FBXW7广泛定位于母中心，并且在含有HsSAS-6的原中心形成的位点减少了它的存在。该观察结果表明，FBXW7将前中心体装配限制在特定位置以生成单个新的中心体。相反，在HsSAS-6过表达期间，FBXW7在中心处与HsSAS-6紧密相关。我们还发现SCFFBXW7与HsSAS-6相互作用并将其靶向泛素介导的降解。此外，我们在HsSAS-6中鉴定出假定的磷酸腺嘌呤位点，其取代使其对FBXW7介导的降解和中心粒数的控制不敏感。总之，SCFFBXW7靶向HsSAS-6进行降解，从而通过限制将HsSAS-6募集到母体中心体来控制中心体的生物发生，