title: Concomitant binding of Afadin to LGN and F-actin directs planar spindle orientation.
abstract: Polarized epithelia form by oriented cell divisions in which the mitotic spindle aligns parallel to the epithelial plane. To orient the mitotic spindle, cortical cues trigger the recruitment of NuMA-dynein-based motors, which pull on astral microtubules via the protein LGN. We demonstrate that the junctional protein Afadin is required for spindle orientation and correct epithelial morphogenesis of Caco-2 cysts. Molecularly, Afadin binds directly and concomitantly to F-actin and to LGN. We determined the crystallographic structure of human Afadin in complex with LGN and show that it resembles the LGN-NuMA complex. In mitosis, Afadin is necessary for cortical accumulation of LGN and NuMA above the spindle poles, in an F-actin-dependent manner. Collectively, our results depict Afadin as a molecular hub governing the enrichment of LGN and NuMA at the cortex. To our knowledge, Afadin is the first-described mechanical anchor between dynein and cortical F-actin.
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Manuel
authors.initials: M
authors.lastname: Carminati
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Gallini
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Pirovano
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Alfieri
authors.affiliation: Fondazione Italiana Ricerca sul Cancro Institute of Molecular Oncology, Milan, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Bisi
authors.affiliation: Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.
authors.forename: Marina
authors.initials: M
authors.lastname: Mapelli
mesh_terms: D000199:Actins / Q000032:analysis / Q000378:metabolism*; D000595:Amino Acid Sequence; D018938:Caco-2 Cells; D018360:Crystallography, X-Ray; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000378:metabolism*; D008840:Microfilament Proteins / Q000032:analysis / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D060066:Protein Interaction Maps; D008941:Spindle Apparatus / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*
chemical_list: D000199:Actins; C102754:GPSM2 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008840:Microfilament Proteins; C108835:afadin