Insights into centriole geometry revealed by cryotomography of doublet and triplet centrioles

Abstract

Centrioles are cylindrical assemblies comprised of 9 singlet, doublet, or triplet microtubules, essential for the formation of motile and sensory cilia. While the structure of the cilium is being defined at increasing resolution, centriolar structure remains poorly understood. Here, we used electron cryo-tomography to determine the structure of mammalian (triplet) and Drosophila (doublet) centrioles. Mammalian centrioles have two distinct domains: a 200 nm proximal core region connected by A-C linkers, and a distal domain where the C-tubule is incomplete and a pair of novel linkages stabilize the assembly producing a geometry more closely resembling the ciliary axoneme. Drosophila centrioles resemble the mammalian core, but with their doublet microtubules linked through the A tubules. The commonality of core-region length, and the abrupt transition in mammalian centrioles, suggests a conserved length-setting mechanism. The unexpected linker diversity suggests how unique centriolar architectures arise in different tissues and organisms.

Electron cryotomography of intact motile cilia defines the basal body to axoneme transition

“…5, A–C). We had previously observed a similar behavior in centrioles, where the disappearance of the distal C-tubule goes through stages of disappearance (Greenan et al, 2018). The end result is a region at the distal BB that is composed of DMTs but still contains the A09/A10 MIP, which is characteristic of the BB.…”

“…Overall, the ciliary base has an hourglass shape, with the BB tapering inward to a restriction point at the proximal TZ, and then spreading outward along the TZ. We previously observed a tapering along the proximal–distal axis of the mammalian centriole (Greenan et al, 2018). A reexamination of our centriole data allowed us to determine the geometry at the extreme distal end of a centriole (Fig.…”

“…In this study, we have gained a comprehensive understanding of the architecture of the motile cilium. Centrioles/BBs and axonemes are relatively straightforward to purify and have been the subject of previous cryoET studies (Greenan et al, 2018; Guichard et al, 2012; Ichikawa et al, 2017; Li et al, 2012; Paintrand et al, 1992). However, purifying the entire structure of the BB through the axoneme, allowing an analysis of the TZ, has been challenging.…”

“…Furthermore, the angled geometry of the BB TMTs is markedly different from the inline geometry of the axonemal DMTs (Paintrand et al, 1992; Sui and Downing, 2006; Vaughan and Gull, 2016). Finally, while the BB TMTs are linked and stabilized via structures such as the A-C linker (Greenan et al, 2018; Guichard et al, 2012; Li et al, 2012), the axoneme DMTs are linked via axonemal dyneins and nexin-DRC necessary for ciliary motility (Heuser et al, 2009; Ishikawa, 2017). Although many of these components have been studied at different resolutions using a combination of light and electron microscopy, the basic process by which the BB transitions to the axoneme is unknown.…”

Electron Cryo-Tomography Provides Insight into Procentriole Architecture and Assembly Mechanism

“…Interestingly, many MIPs identified in this work and in mammalian centrioles (Greenan et al, 2018) are localized at the junction of A-B or B-C tubules in the triplet. For example, MIPs 5~6 are found at the inner A-B junction, while MIPs 9~11 are at the inner B-C tubules junction.…”