Tracheal motile cilia in mice require CAMSAP3 for formation of central microtubule pair and coordinated beating

Motile cilia of multiciliated epithelial cells undergo synchronized beating to produce fluid flow along the luminal surface of various organs. Each motile cilium consists of an axoneme and a basal body, which are linked by a ‘transition zone’. The axoneme exhibits a characteristic 9+2 microtubule arrangement important for ciliary motion, but how this microtubule system is generated is not yet fully understood. Here we show that CAMSAP3, a protein that can stabilize the minus end of a microtubule, concentrates at multiple sites of the cilium–basal body complex, including the upper region of the transition zone or the axonemal basal plate where the central pair of microtubules (CP) initiates. CAMSAP3 dysfunction resulted in loss of the CP and partial distortion of the basal plate, as well as the failure of multicilia to undergo synchronized beating. These findings suggest that CAMSAP3 plays pivotal roles in the formation or stabilization of the CP by localizing at the basal region of the axoneme, and thereby supports the coordinated motion of multicilia in airway epithelial cells. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text]

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Tracheal motile cilia require CAMSAP3 for formation of central microtubule pair and coordinated beating

10.1101/2021.04.21.440849 ◽

2021 ◽

Author(s):

Hiroko Saito ◽

Fumiko Matsukawa-Usami ◽

Toshihiko Fujimori ◽

Toshiya Kimura ◽

Takahiro Ide ◽

...

Keyword(s):

Epithelial Cells ◽

Transition Zone ◽

Basal Body ◽

Airway Epithelial Cells ◽

Basal Plate ◽

Airway Epithelial ◽

Superresolution Microscopy ◽

Ciliary Motion ◽

Motile Cilia ◽

Partial Distortion

AbstractMotile cilia of multiciliated epithelial cells undergo synchronized beating to produce fluid flow along the luminal surface of various organs. Each motile cilium consists of an axoneme and a basal body, which are linked by a ‘transition zone’. The axoneme exhibits a characteristic 9+2 microtubule arrangement important for ciliary motion, but how this microtubule system is generated is not yet fully understood. Here, using superresolution microscopy, we show that CAMSAP3, a protein that can stabilize the minus end of a microtubule, concentrates at multiple sites of the cilium-basal body complex, including the upper region of the transition zone or the axonemal basal plate where the central pair of microtubules (CP) terminates. CAMSAP3 dysfunction resulted in loss of the CP and partial distortion of the basal plate, as well as the failure of multicilia to undergo synchronized beating. These findings indicate that CAMSAP3 plays pivotal roles in the formation or stabilization of the CP, and thereby supports the coordinated motion of multicilia in airway epithelial cells.

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Role of f-box factor foxj1 in differentiation of ciliated airway epithelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00170.2003 ◽

2004 ◽

Vol 286 (4) ◽

pp. L650-L657 ◽

Cited By ~ 110

Author(s):

Yingjian You ◽

Tao Huang ◽

Edward J. Richer ◽

Jens-Erik Harboe Schmidt ◽

Joseph Zabner ◽

...

Keyword(s):

Epithelial Cell ◽

Epithelial Cells ◽

Basal Body ◽

Ciliated Cell ◽

Airway Epithelial Cells ◽

Cell Phenotype ◽

Basal Bodies ◽

Airway Epithelial ◽

Ciliated Cells ◽

Wild Type

Factors required for commitment of an undifferentiated airway epithelial cell to a ciliated cell are unknown. Cell ultrastructure analysis indicates ciliated cell commitment activates a multistage program involving synthesis of cilia precursor proteins and assembly of macromolecular complexes. Foxj1 is an f-box transcription factor expressed in ciliated cells and shown to be required for cilia formation by gene deletion in a mouse model. To identify a specific role for foxj1 in directing the ciliated cell phenotype, we evaluated the capacity of foxj1 to induce ciliogenesis and direct cilia assembly. In a primary culture model of wild-type mouse airway epithelial cells, foxj1 expression preceded the appearance of cilia and in cultured foxj1 null cells cilia did not develop. Delivery of foxj1 to polarized epithelial cell lines and primary cultured alveolar epithelial cells failed to promote ciliogenesis. Similarly, delivery of foxj1 to wild-type airway epithelial cells did not enhance the total number of ciliated cells. In contrast, delivery of foxj1 to null cells resulted in the appearance of cilia. Analysis revealed that, in the absence of foxj1, null cells contained cilia precursor basal bodies, indicating prior commitment to ciliogenesis. However, the basal bodies were disorganized within the apical compartment and failed to dock with the apical membrane. Reconstitution of foxj1 in null cells restored normal basal body organization, resulting in axoneme growth. Thus foxj1 functions in late-stage ciliogenesis to regulate programs promoting basal body docking and axoneme formation in cells previously committed to the ciliated cell phenotype.

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AC6 regulates the microtubule-depolymerizing kinesin KIF19A to control ciliary length in mammals

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.013703 ◽

2020 ◽

Vol 295 (42) ◽

pp. 14250-14259 ◽

Cited By ~ 1

Author(s):

Kavisha Arora ◽

John R. Lund ◽

Nevin A. Naren ◽

Basilia Zingarelli ◽

Anjaparavanda P. Naren

Keyword(s):

Epithelial Cells ◽

Airway Epithelial Cells ◽

Therapeutic Interventions ◽

Airway Epithelial ◽

Microtubule Depolymerization ◽

Signaling Mechanism ◽

Protein Levels ◽

Energy Conserving ◽

Motile Cilia

Motile cilia are hairlike structures that line the respiratory and reproductive tracts and the middle ear and generate fluid flow in these organs via synchronized beating. Cilium growth is a highly regulated process that is assumed to be important for flow generation. Recently, Kif19a, a kinesin residing at the cilia tip, was identified to be essential for ciliary length control through its microtubule depolymerization function. However, there is a lack of information on the nature of proteins and the integrated signaling mechanism regulating growth of motile cilia. Here, we report that adenylate cyclase 6 (AC6), a highly abundant AC isoform in airway epithelial cells, inhibits degradation of Kif19a by inhibiting autophagy, a cellular recycling mechanism for damaged proteins and organelles. Using epithelium-specific knockout mice of AC6, we demonstrated that AC6 knockout airway epithelial cells have longer cilia compared with the WT cells because of decreased Kif19a protein levels in the cilia. We demonstrated in vitro that AC6 inhibits AMP-activated kinase (AMPK), an important modulator of cellular energy-conserving mechanisms, and uncouples its binding with ciliary kinesin Kif19a. In the absence of AC6, activation of AMPK mobilizes Kif19a into autophagosomes for degradation in airway epithelial cells. Lower Kif19a levels upon pharmacological activation of AMPK in airway epithelial cells correlated with elongated cilia and vice versa. In all, the AC6–AMPK pathway, which is tunable to cellular cues, could potentially serve as one of the crucial ciliary growth checkpoints and could be channeled to develop therapeutic interventions for cilia-associated disorders.

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