APC/C Ubiquitin Ligase: Coupling Cellular Differentiation to G1/G0 Phase in Multicellular Systems

The anaphase-promoting complex/cyclosome (APC/C) is an evolutionarily conserved ubiquitin ligase that controls cell cycle progression through spatiotemporally regulated proteolysis. Although recent studies revealed its postmitotic function, our knowledge of the role of APC/C beyond cell cycle regulation in the biology of multicellular organisms is far from complete. Here, I review recent advances in the function of APC/C in animal development, specifically focusing on its emerging role in regulating cell differentiation. I describe how APC/C regulates distinct processes during the course of differentiation by deploying diverse molecular machineries in a variety of developmental contexts. Also, I discuss the significance and clinical relevance of the unique capacity of APC/C and other cell cycle regulators to couple distinct cellular processes with cell proliferation control.This work is supported by the start-up funding from ShanghaiTech University (2018F0202-000-06) and Cancer Research UK (CRUK-A12874) to Y.K

Targeting of Fzr/Cdh1 for timely activation of the APC/C at the centrosome during mitotic exit.

A multi-subunit ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), regulates critical cellular processes including the cell cycle. To accomplish its diverse functions, APC/C activity must be precisely regulated in time and space. The interphase APC/C activator Fizzy-related (Fzr or Cdh1) is localized at centrosomes in animal cells. However, neither the mechanism of its localization nor its importance is clear. Here we identify the centrosome component Spd2 as a major partner of Fzr in Drosophila. The localization of Fzr to the centriole during interphase depends on direct interaction with Spd2. By generating Spd2 mutants unable to bind Fzr, we show that centrosomal localization of Fzr is essential for optimal APC/C activation towards its centrosomal substrate Aurora A. Finally, we show that Spd2 is also a novel APC/C(Fzr) substrate. Our study is the first to demonstrate the critical importance of distinct subcellular pools of APC/C activators in the spatiotemporal control of APC/C activity.Cancer Research UK (Career Development Fellowship), Biotechnology and Biological Sciences Research Council (project grant), Medical Research Council (project grant), Japan Society for the Promotion of Science (Postdoctoral Fellowship for Research Abroad), European Commission (Marie Skłodowska-Curie actions individual fellowship)This is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/ncomms1260

DAPPER: a data-mining resource for protein-protein interactions.

BACKGROUND: The identification of interaction networks between proteins and complexes holds the promise of offering novel insights into the molecular mechanisms that regulate many biological processes. With increasing volumes of such datasets, especially in model organisms such as Drosophila melanogaster, there exists a pressing need for specialised tools, which can seamlessly collect, integrate and analyse these data. Here we describe a database coupled with a mining tool for protein-protein interactions (DAPPER), developed as a rich resource for studying multi-protein complexes in Drosophila melanogaster. RESULTS: This proteomics database is compiled through mass spectrometric analyses of many protein complexes affinity purified from Drosophila tissues and cultured cells. The web access to DAPPER is provided via an accelerated version of BioMart software enabling data-mining through customised querying and output formats. The protein-protein interaction dataset is annotated with FlyBase identifiers, and further linked to the Ensembl database using BioMart's data-federation model, thereby enabling complex multi-dataset queries. DAPPER is open source, with all its contents and source code are freely available. CONCLUSIONS: DAPPER offers an easy-to-navigate and extensible platform for real-time integration of diverse resources containing new and existing protein-protein interaction datasets of Drosophila melanogaster.This work was supported financially by grants from the Cancer Research UK (CRUK), the Biotechnology and Biological Sciences Research Council and the Medical Research Council to DMG (C3/A11431, BB/I013938/1, G1001696), by a Cancer Research UK Career Development Fellowship to YK (C40697/A12874), and by Cancer Research UK grants to PPD (C12296/A8039 and C12296/A12541). ZL is on leave from the Biological Research Centre of the Hungarian Academy of Sciences (Institute of Biochemistry, Szeged, Hungary) and was supported by a Long-Term Fellowship of the Federation of European Biochemical Societies (FEBS)

Plk4 Regulates Centriole Asymmetry and Spindle Orientation in Neural Stem Cells

Defects in mitotic spindle orientation (MSO) disrupt the organization of stem cell niches impacting tissue morphogenesis and homeostasis. Mutations in centrosome genes reduce MSO fidelity, leading to tissue dysplasia and causing several diseases such as microcephaly, dwarfism, and cancer. Whether these mutations perturb spindle orientation solely by affecting astral microtubule nucleation or whether centrosome proteins have more direct functions in regulatingMSO is unknown. To investigate this question, we analyzed the consequences of deregulating Plk4 (the master centriole duplication kinase) activity in Drosophila asymmetrically dividing neural stem cells. We found that Plk4 functions upstream of MSO control, orchestrating centriole symmetry breaking and consequently centrosome positioning. Mechanistically, we show that Plk4 acts through Spd2 phosphorylation, which induces centriole release from the apical cortex. Overall, this work not only reveals a role for Plk4 in regulating centrosome function but also links the centrosome biogenesis machinery with the MSO apparatus.ERC starting grant CentroStemCancer [242598]; Institut Curie; CNRS; NCI [P30CA23074]; NIGMS [R01 GM110166, GM126035]; FRM; IC; FRM installation grant; ATIP grantOpen access articleThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

CCDC61/VFL3 Is a Paralog of SAS6 and Promotes Ciliary Functions.

Centrioles are cylindrical assemblies whose peripheral microtubule array displays a 9-fold rotational symmetry that is established by the scaffolding protein SAS6. Centriole symmetry can be broken by centriole-associated structures, such as the striated fibers in Chlamydomonas that are important for ciliary function. The conserved protein CCDC61/VFL3 is involved in this process, but its exact role is unclear. Here, we show that CCDC61 is a paralog of SAS6. Crystal structures of CCDC61 demonstrate that it contains two homodimerization interfaces that are similar to those found in SAS6, but result in the formation of linear filaments rather than rings. Furthermore, we show that CCDC61 binds microtubules and that residues involved in CCDC61 microtubule binding are important for ciliary function in Chlamydomonas. Together, our findings suggest that CCDC61 and SAS6 functionally diverged from a common ancestor while retaining the ability to scaffold the assembly of basal body-associated structures or centrioles, respectively

Targeting of Fzr/Cdh1 for timely activation of the APC/C at the centrosome during mitotic exit

A multi-subunit ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), regulates critical cellular processes including the cell cycle. To accomplish its diverse functions, APC/C activity must be precisely regulated in time and space. The interphase APC/C activator Fizzy-related (Fzr or Cdh1) is localized at centrosomes in animal cells. However, neither the mechanism of its localization nor its importance is clear. Here we identify the centrosome component Spd2 as a major partner of Fzr in Drosophila. The localization of Fzr to the centriole during interphase depends on direct interaction with Spd2. By generating Spd2 mutants unable to bind Fzr, we show that centrosomal localization of Fzr is essential for optimal APC/C activation towards its centrosomal substrate Aurora A. Finally, we show that Spd2 is also a novel APC/C^(Fzr) substrate. Our study is the first to demonstrate the critical importance of distinct subcellular pools of APC/C activators in the spatiotemporal control of APC/C activity

APC/C-dependent degradation of Spd2 regulates centrosome asymmetry in Drosophila neural stem cells.

Funder: Marie Curie (Marie Curie Cancer Care); Id: http://dx.doi.org/10.13039/501100000654A functional centrosome is vital for the development and physiology of animals. Among numerous regulatory mechanisms of the centrosome, ubiquitin-mediated proteolysis is known to be critical for the precise regulation of centriole duplication. However, its significance beyond centrosome copy number control remains unclear. Using an in vitro screen for centrosomal substrates of the APC/C ubiquitin ligase in Drosophila, we identify several conserved pericentriolar material (PCM) components, including the inner PCM protein Spd2. We show that Spd2 levels are controlled by the interphase-specific form of APC/C, APC/CFzr , in cultured cells and developing brains. Increased Spd2 levels compromise neural stem cell-specific asymmetric PCM recruitment and microtubule nucleation at interphase centrosomes, resulting in partial randomisation of the division axis and segregation patterns of the daughter centrosome in the following mitosis. We further provide evidence that APC/CFzr -dependent Spd2 degradation restricts the amount and mobility of Spd2 at the daughter centrosome, thereby facilitating the accumulation of Polo-dependent Spd2 phosphorylation for PCM recruitment. Our study underpins the critical role of cell cycle-dependent proteolytic regulation of the PCM in stem cells