Hierarchical assembly of centriole subdistal appendages via centrosome binding proteins CCDC120 and CCDC68

Abstract

In animal cells, the centrosome is the main microtubule-organizing centre where microtubules are nucleated and anchored. The centriole subdistal appendages (SDAs) are the key structures that anchor microtubules in interphase cells, but the composition and assembly mechanisms of SDAs are not well understood. Here, we reveal that centrosome-binding proteins, coiled-coil domain containing (CCDC) 120 and CCDC68 are two novel SDA components required for hierarchical SDA assembly in human cells. CCDC120 is anchored to SDAs by ODF2 and recruits CEP170 and Ninein to the centrosome through different coiled-coil domains at its N terminus. CCDC68 is a CEP170-interacting protein that competes with CCDC120 in recruiting CEP170 to SDAs. Furthermore, CCDC120 and CCDC68 are required for centrosome microtubule anchoring. Our findings elucidate the molecular basis for centriole SDA hierarchical assembly and microtubule anchoring in human interphase cells.

Super-resolution microscopy reveals coupling between mammalian centriole subdistal appendages and distal appendages

“…The basal foot is oriented to align with the beating direction of motile cilia, suggesting its role in mechanical coupling or anchoring (Gibbons, 1961, Clare et al, 2014. We and others have identified a series of sDAP proteins, including ODF2, CEP128, centriolin, CCDC68, CCDC120, ninein and CEP170 (Mazo et al, 2016, Huang et al, 2017, Guarguaglini et al, 2005, Nakagawa et al, 2001, Ou et al, 2002, Mogensen et al, 2000, Schrøder et al, 2012. Despite its structural presence routinely observed in EM images, detailed geometrical information of these sDAP proteins relative to each other remains largely missing.…”

“…To systematically characterize sDAP architecture, we imaged multiple sDAP proteins reported previously (Mazo et al, 2016, Huang et al, 2017, Tateishi et al, 2013, Mönnich et al, 2018 by means of dSTORM super-resolution microscopy. ODF2, CEP128, centriolin, CCDC68, ninein and CEP170 of sDAPs were imaged.…”

The regulation of cilium assembly and disassembly in development and disease

“…A number of proteins, including six DA proteins and approximately ten SDA proteins, which are hierarchically recruited to the distal end of the MC have been discovered thus far (Graser et al, 2007;Huang et al, 2017;Joo et al, 2013;Kodani et al, 2013;Kurtulmus et al, 2017 preprint;Mazo et al, 2016;Schmidt et al, 2012;Tanos et al, 2013;Wei et al, 2013). Using super-resolution microscopy, the DA proteins CEP83, CEP89, SCLT1 and CEP164 were shown to form the pinwheel-like spokes of DAs, whereas the interspoke matrix is populated by FBF1 (Yang et al, 2018).…”

Super-resolution Molecular Map of Basal Foot Reveals Novel Cilium in Airway Multiciliated Cells

“…Most basal foot proteins are clustered with ODF2, a component critical for basal foot 133 assembly 4,5 region that was termed the basal body anchoring region or region I. Among the proteins previously 138 assigned to the basal foot/subdistal appendage, e-tubulin, CCDC120, CCDC68 could not be 139 reliably detected at the basal foot with available commercial antibodies, while TCHP, a protein 140 thought to be associated only with subdistal appendages 31,35 , was located to the basal foot in 141 primary cilia. To correctly assign proteins to the basal foot, we then measured the position of basal 142 foot proteins along the axoneme relative to the proximal end of the basal body ( Fig.…”

“…satellite arms 21 , basal feet 25 or subdistal appendages 17 ). 84 Several proteins have been assigned to the basal foot and subdistal appendages in 85 mammalian cells through conventional fluorescence microscopy and immuno-EM (Ninein 86 (NIN) 26 , ODF2/Cenexin 4,27,28 , CC2D2A 29 , CEP170 30 , Galactin-3 3 , e-Tubulin 31 , Centriolin 87 (CNTRL) 32 , Trichoplein (TCHP) 33 , CEP128 17 , CEP19 34 , CCDC120 and CCDC68 35 ). 88 ODF2/Cenexin is a fibrillar protein related to the intermediate filament (IF) superfamily that plays 89 a critical role in basal foot assembly since lack of basal foot-specific ODF2/Cenexin isoform 90 results in loss of the entire structure 4,5,12,36 .…”