title: NuMA Phosphorylation by Aurora-A Orchestrates Spindle Orientation.
abstract: Spindle positioning is essential for tissue morphogenesis and homeostasis. The signaling network synchronizing spindle placement with mitotic progression relies on timely recruitment at the cell cortex of NuMA:LGN:Gαi complexes, in which NuMA acts as a receptor for the microtubule motor Dynein. To study the implication of Aurora-A in spindle orientation, we developed protocols for the partial inhibition of its activity. Under these conditions, in metaphase NuMA and Dynein accumulate abnormally at the spindle poles and do not reach the cortex, while the cortical distribution of LGN remains unperturbed. FRAP experiments revealed that Aurora-A governs the dynamic exchange between the cytoplasmic and the spindle pole-localized pools of NuMA. We show that Aurora-A phosphorylates directly the C terminus of NuMA on three Ser residues, of which Ser1969 determines the dynamic behavior and the spindle orientation functions of NuMA. Most interestingly, we identify a new microtubule-binding domain of NuMA, which does not overlap with the LGN-binding motif. Our study demonstrates that in metaphase the direct phosphorylation of NuMA by Aurora-A controls its cortical enrichment, and that this is the major event underlying the spindle orientation functions of Aurora-A in transformed and non-transformed cells in culture. Phosphorylation of NuMA by Aurora-A does not affect its affinity for microtubules or for LGN but rather determines the mobility of the protein at the spindle poles. The finding that NuMA can associate concomitantly with LGN and microtubules suggests that its microtubule-binding activity contributes to anchor Dynein-loaded microtubule +TIPs at cortical sites with LGN.
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Gallini
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Manuel
authors.initials: M
authors.lastname: Carminati
authors.affiliation: Institute of Molecular Biology and Pathology, CNR National Research Council, c/o Department of Biology and Biotechnology, Sapienza University of Rome, Via degli Apuli 4, 00185 Rome, Italy.
authors.forename: Fabiola
authors.initials: F
authors.lastname: De Mattia
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Pirovano
authors.affiliation: Cogentech S.c.a.r.l., Via Adamello 16, 20139 Milan, Italy; IFOM, the FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Emanuele
authors.initials: E
authors.lastname: Martini
authors.affiliation: Cogentech S.c.a.r.l., Via Adamello 16, 20139 Milan, Italy; IFOM, the FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Amanda
authors.initials: A
authors.lastname: Oldani
authors.affiliation: Institute of Molecular Biology and Pathology, CNR National Research Council, c/o Department of Biology and Biotechnology, Sapienza University of Rome, Via degli Apuli 4, 00185 Rome, Italy.
authors.forename: Italia Anna
authors.initials: IA
authors.lastname: Asteriti
authors.affiliation: Institute of Molecular Biology and Pathology, CNR National Research Council, c/o Department of Biology and Biotechnology, Sapienza University of Rome, Via degli Apuli 4, 00185 Rome, Italy. Electronic address: giulia.guarguaglini@uniroma1.it.
authors.forename: Giulia
authors.initials: G
authors.lastname: Guarguaglini
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Via Adamello 16, 20139 Milan, Italy. Electronic address: marina.mapelli@ieo.eu.
authors.forename: Marina
authors.initials: M
authors.lastname: Mapelli
mesh_terms: D034961:Antigens, Nuclear / Q000235:genetics* / Q000378:metabolism; D064096:Aurora Kinase A / Q000235:genetics* / Q000378:metabolism; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D004398:Dyneins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008677:Metaphase; D034521:Nuclear Matrix-Associated Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D008941:Spindle Apparatus / Q000378:metabolism
chemical_list: D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; C576917:AURKA protein, human; D064096:Aurora Kinase A; D004398:Dyneins