Asterless is a scaffold for the onset of centriole assembly.

Cancer Research UK Cell Cycle Genetics Group, University of Cambridge, Department of Genetics, Downing Street, Cambridge CB2 3EH, UK.
Nature (Impact Factor: 42.35). 10/2010; 467(7316):714-8. DOI: 10.1038/nature09445
Source: PubMed

ABSTRACT Centrioles are found in the centrosome core and, as basal bodies, at the base of cilia and flagella. Centriole assembly and duplication is controlled by Polo-like-kinase 4 (Plk4): these processes fail if Plk4 is downregulated and are promoted by Plk4 overexpression. Here we show that the centriolar protein Asterless (Asl; human orthologue CEP152) provides a conserved molecular platform, the amino terminus of which interacts with the cryptic Polo box of Plk4 whereas the carboxy terminus interacts with the centriolar protein Sas-4 (CPAP in humans). Drosophila Asl and human CEP152 are required for the centrosomal loading of Plk4 in Drosophila and CPAP in human cells, respectively. Depletion of Asl or CEP152 caused failure of centrosome duplication; their overexpression led to de novo centriole formation in Drosophila eggs, duplication of free centrosomes in Drosophila embryos, and centrosome amplification in cultured Drosophila and human cells. Overexpression of a Plk4-binding-deficient mutant of Asl prevented centriole duplication in cultured cells and embryos. However, this mutant protein was able to promote microtubule organizing centre (MTOC) formation in both embryos and oocytes. Such MTOCs had pericentriolar material and the centriolar protein Sas-4, but no centrioles at their core. Formation of such acentriolar MTOCs could be phenocopied by overexpression of Sas-4 in oocytes or embryos. Our findings identify independent functions for Asl as a scaffold for Plk4 and Sas-4 that facilitates self-assembly and duplication of the centriole and organization of pericentriolar material.

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    ABSTRACT: Centrioles are 9-fold symmetrical structures at the core of centrosomes and base of cilia whose dysfunction has been linked to a wide range of inherited diseases and cancer [ 1 ]. Their duplication is regulated by a protein kinase of conserved structure, the C. elegans ZYG-1 or its Polo-like kinase 4 (Plk4) counterpart in other organisms [ 2–4 ]. Although Plk4’s centriolar partners and mechanisms that regulate its stability are known, its crucial substrates for centriole duplication have never been identified. Here we show that Drosophila Plk4 phosphorylates four conserved serines in the STAN motif of the core centriole protein Ana2 to enable it to bind and recruit its Sas6 partner. Ana2 and Sas6 normally load onto both mother and daughter centrioles immediately after their disengagement toward the end of mitosis to seed procentriole formation. Nonphosphorylatable Ana2 still localizes to the centriole but can no longer recruit Sas6 and centriole duplication fails. Thus, following centriole disengagement, recruitment of Ana2 and its phosphorylation by Plk4 are the earliest known events in centriole duplication to recruit Sas6 and thereby establish the architecture of the new procentriole engaged with its parent.
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    ABSTRACT: ZYG-1 is a polo-like kinase essential for centriole assembly in Caenorhabditis elegans. The targeting of ZYG-1 to nascent centrioles is via its central cryptic polo-box (CPB) domain. To shed light on the molecular basis of ZYG-1 recruitment, it is necessary to obtain structural knowledge of the ZYG-1 CPB. Here, the expression, purification and preliminary crystallographic analysis of the ZYG-1 CPB are reported. The protein was overexpressed in Escherichia coli strain BL21 (DE3), purified by multi-step chromatography and crystallized using the vapour-diffusion method. Crystals of the wild-type protein exhibited an order–disorder pathology, which was solved by reductive lysine methylation. A complete anomalous data set was collected to 2.54 Å resolution at the Se K edge (λ = 0.9792 Å). The crystal belonged to space group P2, with unit-cell parameters a = 53.3, b = 60.09, c = 87.51 Å, β = 93.31°. There were two molecules in the asymmetric unit.
    Acta Crystallographica Section F. 10/2014; 70(10).
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    ABSTRACT: Formation of a new centriole adjacent to a pre-existing centriole occurs only once per cell cycle. Despite being crucial for genome integrity, the mechanisms controlling centriole biogenesis remain elusive. Here, we identify RBM14 as a novel suppressor of assembly of centriolar protein complexes. Depletion of RBM14 in human cells induces ectopic formation of centriolar protein complexes through function of the STIL/CPAP complex. 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. Moreover, we find that, upon RBM14 depletion, a part of the ectopic centriolar protein complexes in turn assemble into structures more akin to centrioles, presumably by incorporating HsSAS-6, a cartwheel component, and cause multipolar spindle formation. We further demonstrate that such structures assemble in the cytoplasm even in the presence of pre-existing centrioles. This study sheds light on the possibility that ectopic formation of aberrant structures related to centrioles may contribute to genome instability and tumorigenesis.
    The EMBO Journal 11/2014; · 10.75 Impact Factor

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