scholarly journals Mcidas mutant mice reveal a two-step process for the specification and differentiation of multiciliated cells in mammals

2018 ◽  
Author(s):  
Hao Lu ◽  
Priyanka Anujan ◽  
Feng Zhou ◽  
Yiliu Zhang ◽  
Yan Ling Chong ◽  
...  
Keyword(s):  
Basal Body ◽  
Target Genes ◽  
Initial Step ◽  
Mutant Mice ◽  
Respiratory Disorder ◽  
Basal Bodies ◽  
Cell Cycle Regulators ◽  
Multiciliated Cells ◽  
Motile Cilia ◽  
Body Production

ABSTRACTMotile cilia on multiciliated cells (MCCs) function in fluid clearance over epithelia. Studies with Xenopus embryos and patients with the congenital respiratory disorder reduced generation of multiple motile cilia, have implicated the nuclear protein MCIDAS (MCI), in the transcriptional regulation of MCC specification and differentiation. Recently, a paralogous protein, GMNC, was also shown to be required for MCC formation. Surprisingly, and in contrast to the presently held view, we find that Mci mutant mice can specify MCC precursors. However, these precursors cannot produce multiple basal bodies, and mature into single ciliated cells. We show that MCI is required specifically to induce deuterosome pathway components for the production of multiple basal bodies. Moreover, GMNC and MCI associate differentially with the cell-cycle regulators E2F4 and E2F5, which enables them to activate distinct sets of target genes (ciliary transcription factor genes versus genes for basal body generation). Our data establish a previously unrecognized two-step model for MCC development: GMNC functions in the initial step for MCC precursor specification. GMNC induces Mci expression, which then drives the second step of basal body production for multiciliation.SUMMARY STATEMENTWe show how two GEMININ family proteins function in mammalian multiciliated cell development: GMNC regulates precursor specification and MCIDAS induces multiple basal body formation for multiciliation.

scholarly journals Asymmetrically localized proteins stabilize basal bodies against ciliary beating forces

2016 ◽  
Vol 215 (4) ◽  
pp. 457-466 ◽  
Author(s):  
Brian A. Bayless ◽  
Domenico F. Galati ◽  
Anthony D. Junker ◽  
Chelsea B. Backer ◽  
Jacek Gaertig ◽  
...  
Keyword(s):  
Basal Body ◽  
Mechanical Force ◽  
Basal Bodies ◽  
Mechanical Forces ◽  
Ciliary Beating ◽  
Motile Cilia ◽  
Molecular Components

Basal bodies are radially symmetric, microtubule-rich structures that nucleate and anchor motile cilia. Ciliary beating produces asymmetric mechanical forces that are resisted by basal bodies. To resist these forces, distinct regions within the basal body ultrastructure and the microtubules themselves must be stable. However, the molecular components that stabilize basal bodies remain poorly defined. Here, we determine that Fop1 functionally interacts with the established basal body stability components Bld10 and Poc1. We find that Fop1 and microtubule glutamylation incorporate into basal bodies at distinct stages of assembly, culminating in their asymmetric enrichment at specific triplet microtubule regions that are predicted to experience the greatest mechanical force from ciliary beating. Both Fop1 and microtubule glutamylation are required to stabilize basal bodies against ciliary beating forces. Our studies reveal that microtubule glutamylation and Bld10, Poc1, and Fop1 stabilize basal bodies against the forces produced by ciliary beating via distinct yet interdependent mechanisms.

scholarly journals Sfr1, a Tetrahymena thermophila Sfi1 Repeat Protein, Modulates the Production of Cortical Row Basal Bodies

mSphere ◽  
2016 ◽  
Vol 1 (6) ◽  
Author(s):  
Westley Heydeck ◽  
Alexander J. Stemm-Wolf ◽  
Janin Knop ◽  
Christina C. Poh ◽  
Mark Winey
Keyword(s):  
Basal Body ◽  
Binding Proteins ◽  
Tetrahymena Thermophila ◽  
Binding Protein ◽  
Extensive Study ◽  
Basal Bodies ◽  
Content Type ◽  
Inhibitory Role ◽  
Body Production

ABSTRACT Basal bodies and centrioles are structurally similar and, when rendered dysfunctional as a result of improper assembly or maintenance, are associated with human diseases. Centrins are conserved and abundant components of both structures whose basal body and centriolar functions remain incompletely understood. Despite the extensive study of centrins in Tetrahymena thermophila, little is known about how centrin-binding proteins contribute to centrin’s roles in basal body assembly, stability, and orientation. The sole previous study of the large centrin-binding protein family in Tetrahymena revealed a role for Sfr13 in the stabilization and separation of basal bodies. In this study, we found that Sfr1 localizes to all Tetrahymena basal bodies and complete genetic deletion of SFR1 leads to overproduction of basal bodies. The uncovered inhibitory role of Sfr1 in basal body production suggests that centrin-binding proteins, as well as centrins, may influence basal body number both positively and negatively. Basal bodies are essential microtubule-based structures that template, anchor, and orient cilia at the cell surface. Cilia act primarily in the generation of directional fluid flow and sensory reception, both of which are utilized for a broad spectrum of cellular processes. Although basal bodies contribute to vital cell functions, the molecular contributors of their assembly and maintenance are poorly understood. Previous studies of the ciliate Tetrahymena thermophila revealed important roles for two centrin family members in basal body assembly, separation of new basal bodies, and stability. Here, we characterize the basal body function of a centrin-binding protein, Sfr1, in Tetrahymena. Sfr1 is part of a large family of 13 proteins in Tetrahymena that contain Sfi1 repeats (SFRs), a motif originally identified in Saccharomyces cerevisiae Sfi1 that binds centrin. Sfr1 is the only SFR protein in Tetrahymena that localizes to all cortical row and oral apparatus basal bodies. In addition, Sfr1 resides predominantly at the microtubule scaffold from the proximal cartwheel to the distal transition zone. Complete genomic knockout of SFR1 (sfr1Δ) causes a significant increase in both cortical row basal body density and the number of cortical rows, contributing to an overall overproduction of basal bodies. Reintroduction of Sfr1 into sfr1Δ mutant cells leads to a marked reduction of cortical row basal body density and the total number of cortical row basal bodies. Therefore, Sfr1 directly modulates cortical row basal body production. This study reveals an inhibitory role for Sfr1, and potentially centrins, in Tetrahymena basal body production. IMPORTANCE Basal bodies and centrioles are structurally similar and, when rendered dysfunctional as a result of improper assembly or maintenance, are associated with human diseases. Centrins are conserved and abundant components of both structures whose basal body and centriolar functions remain incompletely understood. Despite the extensive study of centrins in Tetrahymena thermophila, little is known about how centrin-binding proteins contribute to centrin’s roles in basal body assembly, stability, and orientation. The sole previous study of the large centrin-binding protein family in Tetrahymena revealed a role for Sfr13 in the stabilization and separation of basal bodies. In this study, we found that Sfr1 localizes to all Tetrahymena basal bodies and complete genetic deletion of SFR1 leads to overproduction of basal bodies. The uncovered inhibitory role of Sfr1 in basal body production suggests that centrin-binding proteins, as well as centrins, may influence basal body number both positively and negatively.

scholarly journals THE FORMATION OF BASAL BODIES (CENTRIOLES) IN THE RHESUS MONKEY OVIDUCT

1971 ◽  
Vol 50 (1) ◽  
pp. 10-34 ◽  
Author(s):  
Richard G. W. Anderson ◽  
Robert M. Brenner
Keyword(s):  
Rhesus Monkey ◽  
Basal Body ◽  
Wall Material ◽  
Basal Bodies ◽  
Major Pathway ◽  
Tubule Formation ◽  
Body Formation ◽  
Body Production ◽  
Basal Foot ◽  
Section Analysis

Basal body replication during estrogen-driven ciliogenesis in the rhesus monkey (Macaca mulatta) oviduct has been studied by stereomicroscopy, rotation photography, and serial section analysis. Two pathways for basal body production are described: acentriolar basal body formation (major pathway) where procentrioles are generated from a spherical aggregate of fibers; and centriolar basal body formation, where procentrioles are generated by the diplosomal centrioles. In both pathways, the first step in procentriole formation is the arrangement of a fibrous granule precursor into an annulus. A cartwheel structure, present within the lumen of the annulus, is composed of a central cylinder with a core, spoke components, and anchor filaments. Tubule formation consists of an initiation and a growth phase. The A tubule of each triplet set first forms within the wall material of the annulus in juxtaposition to a spoke of the cartwheel. After all nine A tubules are initiated, B and C tubules begin to form. The initiation of all three tubules occurs sequentially around the procentriole. Simultaneous with tubule initiation is a nonsequential growth of each tubule. The tubules lengthen and the procentriole is complete when it is about 200 mµ long. The procentriole increases in length and diameter during its maturation into a basal body. The addition of a basal foot, nine alar sheets, and a rootlet completes the maturation process. Fibrous granules are also closely associated with the formation of these basal body accessory structures.

scholarly journals Protein turnover dynamics suggest a diffusion-to-capture mechanism for peri-basal body recruitment and retention of intraflagellar transport proteins

2021 ◽  
pp. mbc.E20-11-0717
Author(s):  
Jaime V.K. Hibbard ◽  
Neftali Vazquez ◽  
Rohit Satija ◽  
John B. Wallingford
Keyword(s):  
Protein Dynamics ◽  
Basal Body ◽  
Fluorescence Recovery After Photobleaching ◽  
Protein Complexes ◽  
Intraflagellar Transport ◽  
Basal Bodies ◽  
Multiciliated Cells ◽  
Capture Mechanism ◽  
Body Components ◽  
Ciliary Signaling

Intraflagellar transport (IFT) is essential for construction and maintenance of cilia. IFT proteins concentrate at the basal body, where they are thought to assemble into trains and bind cargoes for transport. To study the mechanisms of IFT recruitment to this peri-basal body pool, we quantified protein dynamics of eight IFT proteins, as well as five other basal body localizing proteins, using fluorescence recovery after photobleaching in vertebrate multiciliated cells. We found that members of the IFT-A and IFT-B protein complexes show distinct turnover kinetics from other basal body components. Additionally, known IFT sub-complexes displayed shared dynamics, suggesting shared basal body recruitment and/or retention mechanisms. Finally, we evaluated the mechanisms of basal body recruitment by depolymerizing cytosolic MTs, which suggested that IFT proteins are recruited to basal bodies through a diffusion-to-capture mechanism. Our survey of IFT protein dynamics provides new insights into IFT recruitment to basal bodies, a crucial step in ciliogenesis and ciliary signaling.

scholarly journals Bld10/Cep135 stabilizes basal bodies to resist cilia-generated forces

2012 ◽  
Vol 23 (24) ◽  
pp. 4820-4832 ◽  
Author(s):  
Brian A. Bayless ◽  
Thomas H. Giddings ◽  
Mark Winey ◽  
Chad G. Pearson
Keyword(s):  
Basal Body ◽  
Structural Integrity ◽  
Basal Bodies ◽  
Ciliary Beating ◽  
Full Complement ◽  
Domain Protein ◽  
Motile Cilia ◽  
The Stability

Basal bodies nucleate, anchor, and organize cilia. As the anchor for motile cilia, basal bodies must be resistant to the forces directed toward the cell as a consequence of ciliary beating. The molecules and generalized mechanisms that contribute to the maintenance of basal bodies remain to be discovered. Bld10/Cep135 is a basal body outer cartwheel domain protein that has established roles in the assembly of nascent basal bodies. We find that Bld10 protein first incorporates stably at basal bodies early during new assembly. Bld10 protein continues to accumulate at basal bodies after assembly, and we hypothesize that the full complement of Bld10 is required to stabilize basal bodies. We identify a novel mechanism for Bld10/Cep135 in basal body maintenance so that basal bodies can withstand the forces produced by motile cilia. Bld10 stabilizes basal bodies by promoting the stability of the A- and C-tubules of the basal body triplet microtubules and by properly positioning the triplet microtubule blades. The forces generated by ciliary beating promote basal body disassembly in bld10Δ cells. Thus Bld10/Cep135 acts to maintain the structural integrity of basal bodies against the forces of ciliary beating in addition to its separable role in basal body assembly.

scholarly journals The Distal Appendage Protein CEP164 Is Essential for Efferent Duct Multiciliogenesis and Male Fertility

2020 ◽  
Author(s):  
Mohammed Hoque ◽  
Danny Chen ◽  
Rex A. Hess ◽  
Feng-Qian Li ◽  
Ken-Ichi Takemaru
Keyword(s):  
Basal Body ◽  
Male Fertility ◽  
Apical Cell ◽  
Proper Function ◽  
Seminiferous Tubules ◽  
Basal Bodies ◽  
Epithelial Surface ◽  
Efferent Ducts ◽  
Multiciliated Cells

AbstractCilia are evolutionarily conserved microtubule-based structures that perform diverse biological functions. Cilia are assembled on basal bodies and anchored to the plasma membrane via distal appendages. Multiciliated cells (MCCs) are a specialized cell type with hundreds of motile multicilia, lining the brain ventricles, airways, and reproductive tracts to propel fluids/substances across the epithelial surface. In the male reproductive tract, MCCs in efferent ducts (EDs) move in a whip-like motion to stir the luminal contents and prevent sperm agglutination. Previously, we demonstrated that the essential distal appendage protein CEP164 recruits Chibby1 (Cby1), a small coiled-coil-containing protein, to basal bodies to facilitate basal body docking and ciliogenesis. Mice lacking CEP164 in MCCs (FoxJ1-Cre;CEP164fl/fl) show a significant loss of multicilia in the trachea, oviduct, and ependyma. In addition, we observed male sterility, however, the precise role of CEP164 in male fertility remained unknown. Here, we report that the seminiferous tubules and rete testis of FoxJ1-Cre;CEP164fl/fl mice exhibit substantial dilation, indicative of dysfunctional multicilia in the EDs. Consistent with these findings, multicilia were hardly detectable in the EDs of FoxJ1-Cre;CEP164fl/fl mice although FoxJ1-positive immature cells were present. Sperm aggregation and agglutination were commonly noticeable in the lumen of the seminiferous tubules and EDs of FoxJ1-Cre;CEP164fl/fl mice. In FoxJ1-Cre;CEP164fl/fl mice, the apical localization of Cby1 and the transition zone marker NPHP1 was severely diminished, suggesting basal body docking defects. TEM analysis of EDs further confirmed basal body accumulation in the cytoplasm of MCCs. Collectively, we conclude that deletion of CEP164 in the MCCs of EDs causes basal body docking defects and loss of multicilia, leading to sperm agglutination, obstruction of EDs, and male infertility. Our study therefore unravels an essential role of the distal appendage protein CEP164 in male fertility.Author SummaryMulticilia are tinny hair-like microtubule-based structures that beat in a whip-like pattern to generate a fluid flow on the apical cell surface. Multiciliated cells are essential for the proper function of major organs such as brain, airway, and reproductive tracts. In the male reproductive system, multiciliated cells are present in the efferent ducts, which are small tubules that connect the testis to the epididymis. However, the importance of multiciliated cells in male fertility remains poorly understood. Here, we investigated the role of the critical ciliary protein CEP164 in male fertility using a mouse model lacking CEP164 in multiciliated cells. Male mice are infertile with reduced sperm counts. We demonstrate that, in the absence of CEP164, multiciliated cells are present in the efferent ducts but fail to extend multicilia due to basal body docking defects. Consistent with this, the recruitment of key ciliary proteins is perturbed. As a result, these mice show sperm agglutination, obstruction of sperm transport, and degeneration of germ cells in the testis, leading to infertility. Our study therefore reveals essential roles of CEP164 in the formation of multicilia in the efferent ducts and male fertility.

scholarly journals CAMSAP3 facilitates basal body polarity and the formation of the central pair of microtubules in motile cilia

2020 ◽  
Vol 117 (24) ◽  
pp. 13571-13579 ◽  
Author(s):  
Alan M. Robinson ◽  
Satoe Takahashi ◽  
Eva J. Brotslaw ◽  
Aisha Ahmad ◽  
Emma Ferrer ◽  
...  
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Basal Bodies ◽  
Nasal Airway ◽  
Directional Flow ◽  
Multiple Phenotypes ◽  
Multiciliated Cells ◽  
Motile Cilia ◽  
Ciliary Dyskinesia ◽  
Different Tissues ◽  
Proper Orientation

Synchronized beating of cilia on multiciliated cells (MCCs) generates a directional flow of mucus across epithelia. This motility requires a “9 + 2” microtubule (MT) configuration in axonemes and the unidirectional array of basal bodies of cilia on the MCCs. However, it is not fully understood what components are needed for central MT-pair assembly as they are not continuous with basal bodies in contrast to the nine outer MT doublets. In this study, we discovered that a homozygous knockdown mouse model for MT minus-end regulator calmodulin-regulated spectrin-associated protein 3 (CAMSAP3),Camsap3tm1a/tm1a, exhibited multiple phenotypes, some of which are typical of primary ciliary dyskinesia (PCD), a condition caused by motile cilia defects. Anatomical examination ofCamsap3tm1a/tm1amice revealed severe nasal airway blockage and abnormal ciliary morphologies in nasal MCCs. MCCs from different tissues exhibited defective synchronized beating and ineffective generation of directional flow likely underlying the PCD-like phenotypes. In normal mice, CAMSAP3 localized to the base of axonemes and at the basal bodies in MCCs. However, inCamsap3tm1a/tm1a, MCCs lacked CAMSAP3 at the ciliary base. Importantly, the central MT pairs were missing in the majority of cilia, and the polarity of the basal bodies was disorganized. These phenotypes were further confirmed in MCCs ofXenopusembryos when CAMSAP3 expression was knocked down by morpholino injection. Taken together, we identified CAMSAP3 as being important for the formation of central MT pairs, proper orientation of basal bodies, and synchronized beating of motile cilia.

scholarly journals Emerging Picture of Deuterosome-Dependent Centriole Amplification in MCCs

Cells ◽  
2018 ◽  
Vol 7 (10) ◽  
pp. 152 ◽  
Author(s):  
Umama Shahid ◽  
Priyanka Singh
Keyword(s):  
Basal Body ◽  
De Novo ◽  
Super Resolution ◽  
Basal Bodies ◽  
Parallel Pathways ◽  
Multiciliated Cells ◽  
Dependent Pathway ◽  
Microtubule Structure

Multiciliated cells (MCCs) have several hair-like structures called cilia, which are required to propel substances on their surface. A cilium is organized from a basal body which resembles a hollow microtubule structure called a centriole. In terminally differentiated MCCs, hundreds of new basal bodies/centrioles are formed via two parallel pathways: the centriole- and deuterosome-dependent pathways. The deuterosome-dependent pathway is also referred to as “de novo” because unlike the centriole-dependent pathway which requires pre-existing centrioles, in the de novo pathway multiple new centrioles are organized around non-microtubule structures called deuterosomes. In the last five years, some deuterosome-specific markers have been identified and concurrent advancements in the super-resolution techniques have significantly contributed to gaining insights about the major stages of centriole amplification during ciliogenesis. Altogether, a new picture is emerging which also challenges the previous notion that deuterosome pathway is de novo. This review is primarily focused on studies that have contributed towards the better understanding of deuterosome-dependent centriole amplification and presents a developing model about the major stages identified during this process.

scholarly journals Evidence that the Synchronized Production of New Basal Bodies is not Associated with Dna Synthesis in Stentor Coeruleus

1972 ◽  
Vol 11 (2) ◽  
pp. 621-637
Author(s):  
K. B. YOUNGER ◽  
S. BANERJEE ◽  
J. K. KELLEHER ◽  
M. WINSTON ◽  
LYNN MARGULIS
Keyword(s):  
Dna Synthesis ◽  
Ethidium Bromide ◽  
Basal Body ◽  
Actinomycin D ◽  
Thymidine Incorporation ◽  
Basal Bodies ◽  
Rna And Protein Synthesis ◽  
Stentor Coeruleus ◽  
Body Production ◽  
Dna Synthesis Inhibitors

Stentors were induced to produce synchronously thousands of new ciliated oral membranellar band basal bodies in less than 3 h. DNA synthesis does not accompany this process, as determined by [3H]thymidine incorporation into isolated bands and by sensitivity to DNA synthesis inhibitors (mitomycin C, ethidium bromide, cytosine arabinoside and hydroxyurea). Yet DNA could be detected in the cortex and the band at basal body sites by autoradiography. Since [3H]thymidine incorporation into membranellar band was eliminated in concentrations of ethidium bromide that had no effect on basal body formation, the previous reports of ciliate kinetosomal (basal body) DNA are interpreted as due to mitochondrial contamination. Specific cortical patterns of DNA that could have been easily misinterpreted as basal body-related were especially apparent in autoradiographs using [3H]actinomycin D as a ‘stain’. In no experiment involving induced basal body regeneration could evidence be found for a correlation between new basal body production and DNA synthesis; RNA and protein synthesis correlated with basal body and cilia regeneration were, however, easily detected by the same techniques. We concluded that there is no evidence that basal body DNA synthesis is required for new basal body production.

scholarly journals WDR5 stabilizes actin architecture to promote multiciliated cell formation

2017 ◽  
Author(s):  
Saurabh S. Kulkarni ◽  
John N. Griffin ◽  
Karel F. Liem ◽  
Mustafa K. Khokha
Keyword(s):  
Cell Surface ◽  
Actin Cytoskeleton ◽  
Basal Body ◽  
Chromatin Modification ◽  
Cell Formation ◽  
Basal Bodies ◽  
Actin Network ◽  
Tissue Morphogenesis ◽  
Multiciliated Cells ◽  
Intracellular Organelles

The actin cytoskeleton is critical to shape cells and pattern intracellular organelles to drive tissue morphogenesis. In multiciliated cells (MCCs), apical actin forms a lattice that drives expansion of the cell surface necessary to host hundreds of cilia. The actin lattice also uniformly distributes basal bodies across this surface. This apical actin network is dynamically remodeled, but the molecules that regulate its architecture remain poorly understood. We identify the chromatin modifier, WDR5, as a regulator of apical F-actin in multiciliated cells. Unexpectedly, WDR5 functions independently of chromatin modification in MCCs. Instead, we discover a scaffolding role for WDR5 between the basal body and F-actin. Specifically, WDR5 binds to basal bodies and migrates apically, where F-actin organizes around WDR5. Using a monomer trap for G-actin, we show that WDR5 stabilizes F-actin to maintain apical lattice architecture. In summary, we identify a novel, non-chromatin role for WDR5 in stabilizing F-actin in multiciliated cells.

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