CEP192 localises mitotic Aurora-A activity by priming its interaction with TPX2.

Aurora-A is an essential cell-cycle kinase with critical roles in mitotic entry and spindle dynamics. These functions require binding partners such as CEP192 and TPX2, which modulate both kinase activity and localisation of Aurora-A. Here we investigate the structure and role of the centrosomal Aurora-A:CEP192 complex in the wider molecular network. We find that CEP192 wraps around Aurora-A, occupies the binding sites for mitotic spindle-associated partners, and thus competes with them. Comparison of two different Aurora-A conformations reveals how CEP192 modifies kinase activity through the site used for TPX2-mediated activation. Deleting the Aurora-A-binding interface in CEP192 prevents centrosomal accumulation of Aurora-A, curtails its activation-loop phosphorylation, and reduces spindle-bound TPX2:Aurora-A complexes, resulting in error-prone mitosis. Thus, by supplying the pool of phosphorylated Aurora-A necessary for TPX2 binding, CEP192:Aurora-A complexes regulate spindle function. We propose an evolutionarily conserved spatial hierarchy, which protects genome integrity through fine-tuning and correctly localising Aurora-A activity.

Temporally distinct roles of Aurora A in polarization of the C. elegans zygote.

During asymmetric cell division, cell polarity is coordinated with the cell cycle to allow proper inheritance of cell fate determinants and the generation of cellular diversity. In the Caenorhabditis elegans zygote, polarity is governed by evolutionarily conserved Partitioning-defective (PAR) proteins that segregate to opposing cortical domains to specify asymmetric cell fates. Timely establishment of PAR domains requires a cell cycle kinase, Aurora A (AIR-1 in C. elegans). Aurora A depletion by RNAi causes a spectrum of phenotypes including reversed polarity, excess posterior domains and no posterior domain. How depletion of a single kinase can cause seemingly opposite phenotypes remains obscure. Using an auxin-inducible degradation system and drug treatments, we found that AIR-1 regulates polarity differently at different times of the cell cycle. During meiosis I, AIR-1 acts to prevent later formation of bipolar domains, whereas in meiosis II, AIR-1 is necessary to recruit PAR-2 onto the membrane. Together, these data clarify the origin of multiple polarization phenotypes in RNAi experiments and reveal multiple roles of AIR-1 in coordinating PAR protein localization with cell cycle progression.

Patterns of Ciliation and Ciliary Signaling in Cancer.

Among the factors that have been strongly implicated in regulating cancerous transformation, the primary monocilium (cilium) has gained increasing attention. The cilium is a small organelle extending from the plasma membrane, which provides a localized hub for concentration of transmembrane receptors. These receptors transmit signals from soluble factors (including Sonic hedgehog (SHH), platelet-derived growth factor (PDGF-AA), WNT, TGFß, NOTCH, and others) that regulate cell growth, as well as mechanosensory cues provided by flow or extracellular matrix. Ciliation is regulated by cell cycle, with most cells that are in G0 (quiescent) or early G1 ciliation and cilia typically absent in G2/M cells. Notably, while most cells organized in solid tissues are ciliated, cancerous transformation induces significant changes in ciliation. Most cancer cells lose cilia; medulloblastomas and basal cell carcinomas, dependent on an active SHH pathway, rely on ciliary maintenance. Changes in cancer cell ciliation are driven by core oncogenic pathways (EGFR, KRAS, AURKA, PI3K), and importantly ciliation status regulates functionality of those pathways. Ciliation is both influenced by targeted cancer therapies and linked to therapeutic resistance; recent studies suggest ciliation may also influence cancer cell metabolism and stem cell identity. We review recent studies defining the relationship between cilia and cancer.

The GCN2/eIF2αK stress kinase regulates PP1 to ensure mitotic fidelity.

GCN2/eIF2αK4 is exclusively seen as an eIF2α kinase, which regulates reprogramming of protein translation in response to stress. Here, we show that GCN2 has an unexpected role in unstressed cells as a regulator of mitosis. This function is not through its canonical role in translation reprogramming, but through the regulation of two previously unidentified substrates, PP1α and γ. In the absence of GCN2 function, timing and levels of phosphorylation of key mitotic players are altered, leading to aberrant chromosome alignment, missegregating chromosomes, elevated number of tripolar spindles, and a delay in progression through mitosis. Pharmacological inhibition of GCN2 results in similar effects and is synergistic with Aurora A inhibition in causing more severe mitotic errors and cell death. We suggest that GCN2-dependent phosphorylation of PP1α and γ restrains their activity and this is important to ensure the timely regulation of phosphorylation of several PP1 substrates during early mitosis. These findings highlight a druggable PP1 inhibitor and open new avenues of research on the therapeutic potential of GCN2 inhibitors.

Aurora-A inhibitor synergistically enhances the inhibitory effect of anlotinib on hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a highly malignant tumor with a global prevalence. In addition to the existing clinical guidelines, the effectiveness of anlotinib and Aurora-A inhibitors in treating HCC has also been demonstrated. However, Anlotinib, as an anti-angiogenesis therapy, has shown significant benefits in clinical trials but is limited by its single-agent treatment and the development of drug resistance. Aurora-A inhibitors are currently being tested in clinical trials but have limited efficacy. Combination therapy may offer clear advantages over monotherapy in this context. METHODS: In this study, we used HCC cell lines to investigate whether the combination of the two drugs could enhance their individual strengths and mitigate their weaknesses, thereby providing greater clinical benefits both in vitro and in vivo. RESULTS: Our findings confirmed that the Aurora-A inhibitor alisertib and anlotinib exhibited a time-dose-dependent inhibitory effect on HCC cells. In vitro cytological experiments demonstrated that the combination of the two drugs synergistically inhibited cell proliferation, invasion, and metastasis, while promoting cell apoptosis. Furthermore, we identified the underlying molecular mechanism by which the combination of the Aurora-A inhibitor alisertib and anlotinib inhibited HCC through the inhibition of the NF-ĸB signaling pathway. CONCLUSIONS: In summary, we have demonstrated the effectiveness of combining anlotinib with an Aurora-A inhibitor, which expands the potential applications of anlotinib in the clinical treatment of HCC in the future.

Inhibition of Aurora-A/N-Myc Protein-Protein Interaction Using Peptidomimetics: Understanding the Role of Peptide Cyclization.

Using N-Myc61-89 as a starting template we showcase the systematic use of truncation and maleimide constraining to develop peptidomimetic inhibitors of the N-Myc/Aurora-A protein-protein interaction (PPI); a potential anticancer drug discovery target. The most promising of these - N-Myc73-94-N85C/G89C-mal - is shown to favour a more Aurora-A compliant binding ensemble in comparison to the linear wild-type sequence as observed through fluorescence anisotropy competition assays, circular dichroism (CD) and nuclear magnetic resonance (NMR) experiments. Further in silico investigation of this peptide in its Aurora-A bound state, by molecular dynamics (MD) simulations, imply (i) the bound conformation is more stable as a consequence of the constraint, which likely suppresses dissociation and (ii) the constraint may make further stabilizing interactions with the Aurora-A surface. Taken together this work unveils the first orthosteric N-Myc/Aurora-A inhibitor and provides useful insights on the biophysical properties and thus design of constrained peptides, an attractive therapeutic modality.

Design, synthesis, and biochemical and computational screening of novel oxindole derivatives as inhibitors of Aurora A kinase and SARS-CoV-2 spike/host ACE2 interaction.

Isatin (indol-2,3-dione), a secondary metabolite of tryptophan, has been used as the core structure to design several compounds that have been tested and identified as potent inhibitors of apoptosis, potential antitumor agents, anticonvulsants, and antiviral agents. In this work, several analogs of isatin hybrids have been synthesized and characterized, and their activities were established as inhibitors of both Aurora A kinase and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike/host angiotensin-converting enzyme II (ACE2) interactions. Amongst the synthesized isatin hybrids, compounds 6a, 6f, 6g, and 6m exhibited Aurora A kinase inhibitory activities (with IC50 values < 5 µM), with GScore values of -7.9, -7.6, -8.2 and -7.7 kcal/mol, respectively. Compounds 6g and 6i showed activities in blocking SARS-CoV-2 spike/ACE2 binding (with IC50 values in the range < 30 µM), with GScore values of -6.4 and -6.6 kcal/mol, respectively. Compounds 6f, 6g, and 6i were both capable of inhibiting spike/ACE2 binding and blocking Aurora A kinase. Pharmacophore profiling indicated that compound 6g tightly fits Aurora A kinase and SARS-CoV-2 pharmacophores, while 6d fits SARS-CoV-2 and 6l fits Aurora A kinase pharmacophore. This work is a proof of concept that some existing cancer drugs may possess antiviral properties. Molecular modeling showed that the active compound for each protein adopted different binding modes, hence interacting with a different set of amino acid residues in the binding site. The weaker activities against spike/ACE2 could be explained by the small sizes of the ligands that fail to address the important interactions for binding to the ACE2 receptor site.

Aurora A-mediated pyruvate kinase M2 phosphorylation promotes biosynthesis with glycolytic metabolites and tumor cell cycle progression.

Cancer cells have distinctive demands for intermediates from glucose metabolism for biosynthesis and energy in different cell cycle phases. However, how cell cycle regulators and glycolytic enzymes coordinate to orchestrate the essential metabolic processes are still poorly characterized. Here, we report a novel interaction between the mitotic kinase, Aurora A, and the glycolytic enzyme, pyruvate kinase M2 (PKM2), in the interphase of the cell cycle. We found Aurora A-mediated phosphorylation of PKM2 at threonine 45. This phosphorylation significantly attenuated PKM2 enzymatic activity by reducing its tetramerization and also promoted glycolytic flux and the branching anabolic pathways. Replacing the endogenous PKM2 with a nonphosphorylated PKM2 T45A mutant inhibited glycolysis, glycolytic branching pathways, and tumor growth in both in vitro and in vivo models. Together, our study revealed a new protumor function of Aurora A through modulating a rate-limiting glycolytic enzyme, PKM2, mainly during the S phase of the cell cycle. Our findings also showed that although both Aurora A and Aurora B kinase phosphorylate PKM2 at the same residue, the spatial and temporal regulations of the specific kinase and PKM2 interaction are context dependent, indicating intricate interconnectivity between cell cycle and glycolytic regulators.

The involvement of Aurora-A and p53 in oxaliplatin-resistant colon cancer cells.

Resistance to chemotherapy is the deadlock in cancer treatment. In this study, we used wild-type LOVO (LOVOWT ), a human colon cancer cell line, and the oxaliplatin-resistant sub-clone LOVOOR cells to investigate the molecular mechanisms of the development of drug resistance in colon cancer. Compared with LOVOWT cells, LOVOOR cells had a high proliferation capacity and a high percentage on the G2/M phase. The expression and activation of Aurora-A, a critical kinase in G2/M phase, were higher in LOVOOR cells than in LOVOWT cells. The results from immunofluorescence indicated an irregular distribution of Aurora-A in LOVOOR cells. To evaluate the importance of Aurora-A in oxaliplatin-resistant property of LOVOOR cells, overexpression of Aurora-A in LOVOWT cells and otherwise knockdown of Aurora-A in LOVOOR cells were performed and followed by administration of oxaliplatin. The results indicated that Aurora-A might contribute to the resistance of LOVOOR cells to oxaliplatin treatment by depressing p53 signaling. The specific findings in this study provide a possibility that targeting Aurora-A might be a solution for patients who have failed oxaliplatin treatment.

SUMO proteases SENP3 and SENP5 spatiotemporally regulate the kinase activity of Aurora A.

Precise chromosome segregation is mediated by a well-assembled mitotic spindle, which requires balance of the kinase activity of Aurora A (AurA, also known as AURKA). However, how this kinase activity is regulated remains largely unclear. Here, using in vivo and in vitro assays, we report that conjugation of SUMO2 with AurA at K258 in early mitosis promotes the kinase activity of AurA and facilitates the binding with its activator Bora. Knockdown of the SUMO proteases SENP3 and SENP5 disrupts the deSUMOylation of AurA, leading to increased kinase activity and abnormalities in spindle assembly and chromosome segregation, which could be rescued by suppressing the kinase activity of AurA. Collectively, these results demonstrate that SENP3 and SENP5 deSUMOylate AurA to render spatiotemporal control on its kinase activity in mitosis. This article has an associated First Person interview with the first author of the paper.

Endosulfine alpha maintains spindle pole integrity by recruiting Aurora A during mitosis.

BACKGROUND: The maintenance of spindle pole integrity is essential for spindle assembly and chromosome segregation during mitosis. However, the underlying mechanisms governing spindle pole integrity remain unclear. METHODS: ENSA was inhibited by siRNA or MKI-2 treatment and its effect on cell cycle progression, chromosome alignment and microtubule alignment was observed by immunohistochemical staining and western blotting. PP2A-B55α knockdown by siRNA was performed to rescue the phenotype caused by ENSA inhibition. The interaction between ENSA and Aurora A was detected by in situ PLA. Furthermore, orthotopic implantation of 4Tl-luc cancer cells was conducted to confirm the consistency between the in vitro and in vivo relationship of the ENSA-Aurora A interaction. RESULTS: During mitosis, p-ENSA is localized at the spindle poles, and the inhibition of ENSA results in mitotic defects, such as misaligned chromosomes, multipolar spindles, asymmetric bipolar spindles, and centrosome defects, with a delay in mitotic progression. Although the mitotic delay caused by ENSA inhibition was rescued by PP2A-B55α depletion, spindle pole defects persisted. Notably, we observed a interaction between ENSA and Aurora A during mitosis, and inhibition of ENSA reduced Aurora A expression at the mitotic spindle poles. Injecting MKI-2-sensitized tumors led to increased chromosomal instability and downregulation of the MASTL-ENSA-Aurora A pathway in an orthotopic breast cancer mouse model. CONCLUSIONS: These findings provide novel insights into the regulation of spindle pole integrity by the MASTL-ENSA-Aurora A pathway during mitosis, highlighting the significance of ENSA in recruiting Aurora A to the spindle pole, independent of PP2A-B55α.

The crescent-like Golgi ribbon is shaped by the Ajuba/PRMT5/Aurora-A complex-modified HURP.

BACKGROUND: Golgi apparatus (GA) is assembled as a crescent-like ribbon in mammalian cells under immunofluorescence microscope without knowing the shaping mechanisms. It is estimated that roughly 1/5 of the genes encoding kinases or phosphatases in human genome participate in the assembly of Golgi ribbon, reflecting protein modifications play major roles in building Golgi ribbon. METHODS: To explore how Golgi ribbon is shaped as a crescent-like structure under the guidance of protein modifications, we identified a protein complex containing the scaffold proteins Ajuba, two known GA regulators including the protein kinase Aurora-A and the protein arginine methyltransferase PRMT5, and the common substrate of Aurora-A and PRMT5, HURP. Mutual modifications and activation of PRMT5 and Aurora-A in the complex leads to methylation and in turn phosphorylation of HURP, thereby producing HURP p725. The HURP p725 localizes to GA vicinity and its distribution pattern looks like GA morphology. Correlation study of the HURP p725 statuses and GA structure, site-directed mutagenesis and knockdown-rescue experiments were employed to identify the modified HURP as a key regulator assembling GA as a crescent ribbon. RESULTS: The cells containing no or extended distribution of HURP p725 have dispersed GA membranes or longer GA. Knockdown of HURP fragmentized GA and HURP wild type could, while its phosphorylation deficiency mutant 725A could not, restore crescent Golgi ribbon in HURP depleted cells, collectively indicating a crescent GA-constructing activity of HURP p725. HURP p725 is transported, by GA membrane-associated ARF1, Dynein and its cargo adaptor Golgin-160, to cell center where HURP p725 forms crescent fibers, binds and stabilizes Golgi assembly factors (GAFs) including TRIP11, GRASP65 and GM130, thereby dictating the formation of crescent Golgi ribbon at nuclear periphery. CONCLUSIONS: The Ajuba/PRMT5/Aurora-A complex integrates the signals of protein methylation and phosphorylation to HURP, and the HURP p725 organizes GA by stabilizing and recruiting GAFs to its crescent-like structure, therefore shaping GA as a crescent ribbon. Therefore, the HURP p725 fiber serves a template to construct GA according to its shape. Video Abstract.

A novel class of pyrazole analogues as aurora kinase A inhibitor: design, synthesis, and anticancer evaluation.

Pyrazole, as a small molecule, was discovered for higher cytotoxicity and affinity towards Aurora-A kinase. Based on these facts, a novel pyrazole substituted at the 4th position was designed, synthesized, and evaluated against MCF-7, MDA-MB-23, and Vero (non-cancerous kidney cell) cell lines. Compounds5hand5eexhibited greater cytotoxicity in the series against MCF-7 and MDA-MB-231, with GI50 values of 0.12 µM and 0.63 µM, respectively, as compared to Imatinib (GI50 values of 16.08 µM and 10.36 µM). All of the compounds displayed selective cytotoxicity against cancer cells but not on normal Vero cells, supporting the design strategy to be a selective anticancer agent. Furthermore, compounds 5h and 5e inhibited Aurora-A kinase with IC50 values of 0.78 µM (4.70-fold) and 1.12 µM (2.84-fold), respectively, as compared to alisertib (IC50 = 3.36 µM). In addition, compound 5h significantly arrested the cell cycle at G2/M (34.89 %, 5.56-fold) and the apoptotic phase (25.04 %, 11.81-fold) compared to the control. It also triggered an increase in early (7.43 %) and late (14.89 %) phase apoptosis compared to vehicle (0.235 and 0.36 %, respectively), causing 37.89-fold higher total apoptosis in the annexin-V assay. These data imply that Aurora-A kinase inhibition may be linked to apoptosis induction and cell cycle arrest. Furthermore, their higher docking score in the study confirmed evidence of Aurora-kinase suppression. It was observed that fluorine and imidazole increased the H-bond and lipophilic interactions with the binding residue. Also, the substitution of electron-rich and lipophilic groups increased hydrophobic interactions. Moreover, the three-atom linkage (CH2NHCH2) expanded compound 5h to fill the cavity. Based on current findings, it is concluded that compounds 5h and 5e with strong Aurora-A kinase suppression may be promising anticancer agents.

Structure optimization, synthesis, and biological evaluation of 6-(2-amino-1H-benzo[d]imidazole-6-yl)-quinazolin-4(3H)-one derivatives as potential multi-targeted anticancer agents via Aurora A/ PI3K/BRD4 inhibition.

Aurora A (Aurora kinase A), a critical regulator of cell mitosis, is frequently overexpressed in many malignant cancers, and has been considered as a promising drug target for cancer therapy. Likewise, Phosphatidylinositol 3-kinase alpha (PI3Kα) is also regarded as one of the most important targets in cancer therapy by mediating the cell growth and angiogenesis of various human cancers. In addition, Bromodomain-containing protein 4 (BRD4) modulates oncogene expressions of Myc, Aurora kinase and various RTKs. Recently, accumulating evidences indicated that hyperactivated or abnormally expressed Aurora A, PI3Kα or BRD4 are closely associated with drug resistance and poor prognosis of non-small cell lung cancer (NSCLC). Hence, simultaneous inhibition of Aurora A, PI3Kα, and BRD4 is expected to be a new strategy for NSCLC therapy. In this study, we performed further structure optimization of 6-(2-amino-1H-benzo[d]imidazole-6-yl)-quinazolin-4(3H) -one based on previous study to obtain a series of derivatives for discovering potential Aurora A, PI3Kα and BRD4 multi-targeted inhibitors. MTT assay showed that most of the newly synthesized compounds exhibited an evident anticancer activity against the NSCLC cells. Among them, the IC50 values of the most potent compound 9a were 0.83, 0.26 and 1.02 µM against A549, HCC827 and H1975 cells, respectively. In addition, 9a markedly inhibited the Aurora A and PI3Kα kinase activities with IC50 values of 10.19 nM and 13.12 nM. Compound 9a induced G2/M phase arrests and apoptosis of HCC827 cells by simultaneous inhibition of Aurora A/PI3K/ BRD4 signaling pathways. Collectively, our studies suggested that 9a might be a potential multi-targeted inhibitor for NSCLC therapy.

Analysis of Predicted Amino Acid Sequences of Diatom Microtubule Center Components.

Diatoms synthesize species-specific exoskeletons inside cells under the control of the cytoskeleton and microtubule center. Previous studies have been conducted with the visualization of the microtubule center; however, its composition has not been studied and reliably established. In the present study, several components of MTOC in diatoms, GCP (gamma complex proteins), Aurora A, and centrins have been identified. Analysis of the predicted amino acid sequences of these proteins revealed structural features typical for diatoms. We analyzed the conserved amino acids and the motives necessary for the functioning of proteins. Phylogenetic analysis of GCP showed that all major groups of diatoms are distributed over phylogenetic trees according to their systematic position. This work is a theoretical study; however, it allows drawing some conclusions about the functioning of the studied components and possible ways to regulate them.

When Just One Phosphate Is One Too Many: The Multifaceted Interplay between Myc and Kinases.

Myc transcription factors are key regulators of many cellular processes, with Myc target genes crucially implicated in the management of cell proliferation and stem pluripotency, energy metabolism, protein synthesis, angiogenesis, DNA damage response, and apoptosis. Given the wide involvement of Myc in cellular dynamics, it is not surprising that its overexpression is frequently associated with cancer. Noteworthy, in cancer cells where high Myc levels are maintained, the overexpression of Myc-associated kinases is often observed and required to foster tumour cells' proliferation. A mutual interplay exists between Myc and kinases: the latter, which are Myc transcriptional targets, phosphorylate Myc, allowing its transcriptional activity, highlighting a clear regulatory loop. At the protein level, Myc activity and turnover is also tightly regulated by kinases, with a finely tuned balance between translation and rapid protein degradation. In this perspective, we focus on the cross-regulation of Myc and its associated protein kinases underlying similar and redundant mechanisms of regulation at different levels, from transcriptional to post-translational events. Furthermore, a review of the indirect effects of known kinase inhibitors on Myc provides an opportunity to identify alternative and combined therapeutic approaches for cancer treatment.

BRCA1 transports the DNA damage signal for CDDP-induced centrosome amplification through the centrosomal Aurora A.

Breast cancer gene 1 (BRCA1) plays roles in DNA repair and centrosome regulation and is involved in DNA damage-induced centrosome amplification (DDICA). Here, the centrosomal localization of BRCA1 and the kinases involved in centrosome duplication were analyzed in each cell cycle phase after treatment with DNA crosslinker cisplatin (CDDP). CDDP treatment increased the centrosomal localization of BRCA1 in early S-G2 phase. BRCA1 contributed to the increased centrosomal localization of Aurora A in S phase and that of phosphorylated Polo-like kinase 1 (PLK1) in late S phase after CDDP treatment, resulting in centriole disengagement and overduplication. The increased centrosomal localization of BRCA1 and Aurora A induced by CDDP treatment involved the nuclear export of BRCA1 and BRCA1 phosphorylation by ataxia telangiectasia mutated (ATM). Patient-derived variants and mutations at phosphorylated residues of BRCA1 suppressed the interaction between BRCA1 and Aurora A, as well as the CDDP-induced increase in the centrosomal localization of BRCA1 and Aurora A. These results suggest that CDDP induces the phosphorylation of BRCA1 by ATM in the nucleus and its transport to the cytoplasm, thereby promoting the centrosomal localization Aurora A, which phosphorylates PLK1. The function of BRCA1 in the translocation of the DNA damage signal from the nucleus to the centrosome to induce centrosome amplification after CDDP treatment might support its role as a tumor suppressor.

RACK1 regulates centriole duplication through promoting the activation of polo-like kinase 1 by Aurora A.

Breast cancer gene 1 (BRCA1) contributes to the regulation of centrosome number. We previously identified receptor for activated C kinase 1 (RACK1) as a BRCA1-interacting partner. RACK1, a scaffold protein that interacts with multiple proteins through its seven WD40 domains, directly binds to BRCA1 and localizes to centrosomes. RACK1 knockdown suppresses centriole duplication, whereas RACK1 overexpression causes centriole overduplication in a subset of mammary gland-derived cells. In this study, we showed that RACK1 binds directly to polo-like kinase 1 (PLK1) and Aurora A, and promotes the Aurora A-PLK1 interaction. RACK1 knockdown decreased phosphorylated PLK1 (p-PLK1) levels and the centrosomal localization of Aurora A and p-PLK1 in S phase, whereas RACK1 overexpression increased p-PLK1 level and the centrosomal localization of Aurora A and p-PLK1 in interphase, resulting in an increase of cells with abnormal centriole disengagement. Overexpression of cancer-derived RACK1 variants failed to enhance the Aurora A-PLK1 interaction, PLK1 phosphorylation and the centrosomal localization of p-PLK1. These results suggest that RACK1 functions as a scaffold protein that promotes the activation of PLK1 by Aurora A in order to promote centriole duplication.This article has an associated First Person interview with the first author of the paper.

A novel mitosis-specific Cep215 domain interacts with Cep192 and phosphorylated Aurora A for organization of spindle poles.

The centrosome, which consists of centrioles and pericentriolar material (PCM), becomes mature and assembles mitotic spindles by increasing the number of microtubules (MTs) emanating from the PCM. Among the molecules involved in centrosome maturation, Cep192 and Aurora A (AurA, also known as AURKA) are primarily responsible for recruitment of γ-tubulin and MT nucleators, whereas pericentrin (PCNT) is required for PCM organization. However, the role of Cep215 (also known as CDK5RAP2) in centrosome maturation remains elusive. Cep215 possesses binding domains for γ-tubulin, PCNT and MT motors that transport acentrosomal MTs towards the centrosome. We identify a mitosis-specific centrosome-targeting domain of Cep215 (215N) that interacts with Cep192 and phosphorylated AurA (pAurA). Cep192 is essential for targeting 215N to centrosomes, and centrosomal localization of 215N and pAurA is mutually dependent. Cep215 has a relatively minor role in γ-tubulin recruitment to the mitotic centrosome. However, it has been shown previously that this protein is important for connecting mitotic centrosomes to spindle poles. Based on the results of rescue experiments using versions of Cep215 with different domain deletions, we conclude that Cep215 plays a role in maintaining the structural integrity of the spindle pole by providing a platform for the molecules involved in centrosome maturation.

AURKA destruction is decoupled from its activity at mitotic exit but is essential to suppress interphase activity.

Activity of AURKA is controlled through multiple mechanisms including phosphorylation, ubiquitin-mediated degradation and allosteric interaction with TPX2. Activity peaks at mitosis, before AURKA is degraded during and after mitotic exit in a process strictly dependent on the APC/C coactivator FZR1. We used FZR1 knockout cells (FZR1KO) and a novel FRET-based AURKA biosensor to investigate how AURKA activity is regulated in the absence of destruction. We found that AURKA activity in FZR1KO cells dropped at mitotic exit as rapidly as in parental cells, despite absence of AURKA destruction. Unexpectedly, TPX2 was degraded normally in FZR1KO cells. Overexpression of an N-terminal TPX2 fragment sufficient for AURKA binding, but that is not degraded at mitotic exit, caused delay in AURKA inactivation. We conclude that inactivation of AURKA at mitotic exit is determined not by AURKA degradation but by degradation of TPX2 and therefore is dependent on CDC20 rather than FZR1. The biosensor revealed that FZR1 instead suppresses AURKA activity in interphase and is critically required for assembly of the interphase mitochondrial network after mitosis.This article has an associated First Person interview with the first authors of the paper.