Abstract
Centrioles are vital cellular structures that form centrosomes and cilia. The formation and function of cilia depends on a set of centriole’s distal appendages. In this study, we use correlative super resolution and electron microscopy to precisely determine where distal appendage proteins localize in relation to the centriole microtubules and appendage electron densities. Here we characterize a novel distal appendage protein ANKRD26 and detail, in high resolution, the initial steps of distal appendage assembly. We further show that distal appendages undergo a dramatic ultra-structural reorganization before mitosis, during which they temporarily lose outer components, while inner components maintain a nine-fold organization. Finally, using electron tomography we reveal that mammalian distal appendages associate with two centriole microtubule triplets via an elaborate filamentous base and that they appear as almost radial finger-like protrusions. Our findings challenge the traditional portrayal of mammalian distal appendage as a pinwheel-like structure that is maintained throughout mitosis.
Highlights
Centrioles are vital cellular structures that form centrosomes and cilia
This study demonstrates the power of correlative imaging approaches in revealing architectural and dynamic properties of complex cellular structures with a nanoscale resolution
In nanoscale resolution, the early phases of DA assembly, which demonstrated that DA formation occurs gradually through accumulation of the inner components CCDC41 and SCLT1 in G2
Summary
Centrioles are vital cellular structures that form centrosomes and cilia. The formation and function of cilia depends on a set of centriole’s distal appendages. Human centrioles are ~500 nm long and exhibit proximal-distal polarity[1,2] On their proximal ends, a centriole’s wall is ~230 nm wide and built of nine MT triplets, which consist of one full MT and two partial MTs (Fig. 1). The distal end of centrioles is the assembly site of two types of electron-dense projections called distal and subdistal appendages (DAs and SDAs, respectively). The assembly of SDAs is initiated by the recruitment of Odf[2] around the centriole’s MTs, followed by the recruitment of CCDC68, CCDC120, Cep[170], Cep[128], and Ninein[17,18,19]. Odf[2] remains associated with centrioles throughout the cell cycle, SDA electron microscopy (EM) densities become undetectable in mitosis[2]. The levels of centriole-associated Cep[164] decrease before mitosis[16,20], suggesting that DAs might remodel in mitosis similar to SDAs
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