title: Mechanisms of HsSAS-6 assembly promoting centriole formation in human cells.
abstract: SAS-6 proteins are thought to impart the ninefold symmetry of centrioles, but the mechanisms by which their assembly occurs within cells remain elusive. In this paper, we provide evidence that the N-terminal, coiled-coil, and C-terminal domains of HsSAS-6 are each required for procentriole formation in human cells. Moreover, the coiled coil is necessary and sufficient to mediate HsSAS-6 centrosomal targeting. High-resolution imaging reveals that GFP-tagged HsSAS-6 variants localize in a torus around the base of the parental centriole before S phase, perhaps indicative of an initial loading platform. Moreover, fluorescence recovery after photobleaching analysis demonstrates that HsSAS-6 is immobilized progressively at centrosomes during cell cycle progression. Using fluorescence correlation spectroscopy and three-dimensional stochastic optical reconstruction microscopy, we uncover that HsSAS-6 is present in the cytoplasm primarily as a homodimer and that its oligomerization into a ninefold symmetrical ring occurs at centrioles. Together, our findings lead us to propose a mechanism whereby HsSAS-6 homodimers are targeted to centrosomes where the local environment and high concentration of HsSAS-6 promote oligomerization, thus initiating procentriole formation.
authors.affiliation: Swiss Institute for Experimental Cancer Research, School of Life Sciences, 2 Laboratory for Experimental Biophysics, and 3 Institute for Science and Chemical Engineering, Swiss Federal Institute of Technology, 1015 Lausanne, Switzerland.
authors.forename: Debora
authors.initials: D
authors.lastname: Keller
authors.forename: Meritxell
authors.initials: M
authors.lastname: Orpinell
authors.forename: Nicolas
authors.initials: N
authors.lastname: Olivier
authors.forename: Malte
authors.initials: M
authors.lastname: Wachsmuth
authors.forename: Robert
authors.initials: R
authors.lastname: Mahen
authors.forename: Romain
authors.initials: R
authors.lastname: Wyss
authors.forename: Virginie
authors.initials: V
authors.lastname: Hachet
authors.forename: Jan
authors.initials: J
authors.lastname: Ellenberg
authors.forename: Suliana
authors.initials: S
authors.lastname: Manley
authors.forename: Pierre
authors.initials: P
authors.lastname: Gönczy
mesh_terms: D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000032:analysis / Q000378:metabolism* / Q000502:physiology; D045744:Cell Line, Tumor; D002502:Centrioles / Q000378:metabolism* / Q000648:ultrastructure; D019281:Dimerization; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D008954:Models, Biological; D021381:Protein Transport
chemical_list: D018797:Cell Cycle Proteins; C497202:SASS6 protein, human