CCDC15 localizes to the centriole inner scaffold and regulates centriole integrity and ciliogenesis

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Abstract

Centrioles are evolutionarily conserved microtubule-based organelles critical to form centrosomes and cilia, which act as microtubule-organizing, signaling and motility centers. Biogenesis and maintenance of centrioles with proper number, size and architecture are crucial for their functions during development and physiology. Consequently, their deregulation causes developmental disorders and cancer. Although centriole number control has been extensively studied, less is known about how centrioles are maintained as stable structures with conserved size and architecture over successive cell divisions and upon ciliary and flagellar motility. Here, we addressed this question by identifying and characterizing new components of the centriole inner scaffold, a recently discovered centriolar sub-compartment critical for centriole size control and integrity. To this end, we generated proximity interactomes of Centrin-2 and POC5 and used them to define CCDC15 as a new centriolar protein that co-localizes and interacts with known inner scaffold proteins. Ultrastructure expansion microscopy analysis of CCDC15-depleted cells revealed its functions in centriole length control and integrity, resulting in defective ciliogenesis and Hedgehog signaling. Loss-of-function experiments also defined CCDC15 as a dual regulator for the recruitment of the inner scaffold protein POC1B and the distal SFI1/Centrin complex to the centrioles. Together, our findings uncovered new players and mechanisms of centriole architectural integrity and thereby, provide insights into diseases linked to centriolar defects.

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  1. Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.

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    Reply to the reviewers

    We thank the reviewers for their comments and constructive suggestions to improve the manuscript. We are encouraged to see that both reviewers acknowledge how the results from our manuscript uses state-of-art technologies to advance molecular underpinnings of centriole length, integrity and function regulation. Both reviewers also highlighted that the manuscript is well laid out and presents clear, rigorous, and convincing data. Reviewer#1 described our manuscript of highest experimental quality and broad interest to the field of centrosome and cell biology form a basic research and genetics/clinical point of view. Here, we explain the revisions, additional …

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    Referee #2

    Evidence, reproducibility and clarity

    Summary:

    In this study, Arslanhan et al. propose CCDC15 as a novel component of the centriole inner scaffold structure with potential roles in centriole length control, stability and the primary cilium formation in cultured epithelial cells. Using proximity labelling they explore the common interactors of Poc5 and Centrin-2, two resident molecules of the centriole inner scaffold, to hunt for novel regulators of this structure. The authors leverage expansion microscopy-based localization and siRNA-dependent loss-of-function experiments to follow up on one such protein they identify, CCDC15, with the aforementioned roles in centriole …

  3. Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.

    Learn more at Review Commons


    Referee #1

    Evidence, reproducibility and clarity

    Summary:

    In this manuscript, Arslanhan and colleagues use proximity proteomics to identify CCDC15 as a new centriolar protein that co-localizes and interacts with known inner scaffold proteins in cell culture-based systems. Functional characterization using state-of-the-art expansion microscopy techniques reveals defects in centriole length and integrity. The authors further reveal intriguing aberrations in the recruitment of other centriole inner scaffold proteins, such as POC1B and the SFI1/centrin complex, in CCDC15-deficient cells, and observe defects in primary cilia.

    Major comments:

    • The authors present a high-quality …