Superresolution characterization of core centriole architecture

Abstract

The centrosome is the main microtubule-organizing center in animal cells. It comprises of two centrioles and the surrounding pericentriolar material. Protein organization at the outer layer of the centriole and outward has been studied extensively; however, an overall picture of the protein architecture at the centriole core has been missing. Here we report a direct view of Drosophila centriolar proteins at ∼50-nm resolution. This reveals a Sas6 ring at the C-terminus, where it overlaps with the C-terminus of Cep135. The ninefold symmetrical pattern of Cep135 is further conveyed through Ana1–Asterless axes that extend past the microtubule wall from between the blades. Ana3 and Rcd4, whose termini are close to Cep135, are arranged in ninefold symmetry that does not match the above axes. During centriole biogenesis, Ana3 and Rcd4 are sequentially loaded on the newly formed centriole and are required for centriole-to-centrosome conversion through recruiting the Cep135–Ana1–Asterless complex. Together, our results provide a spatiotemporal map of the centriole core and implications of how the structure might be built.

Ana1 helps recruit Polo to centrioles to promote mitotic PCM assembly and centriole elongation

“…To identify proteins involved in recruiting Polo to the mother centriole we examined a small number of candidates that are important for centriole assembly and/or function in flies and that, like Polo, localise in a ring around the mother centriole: Sas-4 (known as CPAP in vertebrates), Asl, Cep135, Ana1 and PLP (the Drosophila homologue of pericentrin) ( Mennella et al, 2012 ; Fu and Glover, 2012 ; Fu et al, 2016 ; Saurya et al, 2016 ; Tian et al, 2021 ). The PBD is required to efficiently target PLK1 to centrioles and centrosomes ( Elia et al, 2003a , b ; Hanisch et al, 2006 ; Jang et al, 2002 ; Lee et al, 1998 ; Seong et al, 2002 ; Song et al, 2000 ; Reynolds and Ohkura, 2003 ), and we previously uncovered the role of Spd-2 in recruiting Polo to the mitotic PCM by mutating all 34 of the potential PBD-binding S-S/T motifs in Spd-2 to T-S/T.…”

Nine‐fold symmetry of centriole: The joint efforts of its core proteins

“…[ 3,13–15,17,23,24 ] The length of the cartwheel and the microtubule wall vary vastly among organisms. The total length of the basal body is ≈400 nm in Chlamydomonas , and the cartwheel‐containing proximal region is ≈100 nm [ 25 ] ; the cartwheel is ≈75 nm long in Paramecium , ≈300 nm in Naegleria , ≈190 nm in the procentriole of human cells [ 23 ] ; Drosophila embryonic centriole is ≈150 nm long, and the cartwheel extends its entire length [ 3,26,27 ] ; the centriole is extremely long in Trichonympha , about 4000 nm, and hundreds of cartwheel layers form nearly 90% of its centriole length. [ 15,28 ]…”

“…Recently, Drosophila centriolar core proteins, mainly located in zone I and II, were revealed with better resolution by stimulated emission depletion (STED) microscopy. [ 27 ] Sas6, Ana2, Cep135, as well as two novel centriolar components, Ana3 and Rcd4, were revealed into ring structure whose diameters were able to be measured accurately (Figure 4). Moreover, U‐ExM has been applied to physically expand the centriole and successfully revealed the nine‐fold distribution of centriolar proteins, [ 27,37 ] which we will discuss in the following section.…”

“…In Drosophila , Cep135, Ana1 and Asl have been identified to form a protein complex that connects Sas6 to the inner PCM, with four proteins overlapping sequentially via their extended termini. [ 9,27,120 ] Combination of U‐ExM and 3D‐SIM revealed that the C terminus of Cep135, both termini of Ana1 and Asl are each distributed as nine discrete dots forming rings of various diameters (Figure 4). Interestingly, their signals are well aligned along the nine radial axes of the cartwheel, indicating they constitute the spokes and the pinheads.…”

The structure and function of centriolar rootlets

“…Electron microscopy suggests that rootlets attach centrally to the centriole in ciliated osteocytes ( Uzbekov et al, 2012 ). Cep135 is found in the centriolar lumen, is required for centrosome cohesion and binds to cNap1, so it is possible that cNap1 attaches to it there ( Hardy et al, 2014 ; Kim et al, 2008 ; Sonnen et al, 2012 ; Tian et al, 2021 ). However, it is notable that, based on electron microscopy observations, rootlets appear to attach to multiple different locations on centrioles, either in the centriole lumen or at the side of the barrel ( Hagiwara et al, 2008 ).…”