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Sample records for motile mammalian cilia

  1. Structure and function of mammalian cilia.

    PubMed

    Satir, Peter; Christensen, Søren T

    2008-06-01

    In the past half century, beginning with electron microscopic studies of 9 + 2 motile and 9 + 0 primary cilia, novel insights have been obtained regarding the structure and function of mammalian cilia. All cilia can now be viewed as sensory cellular antennae that coordinate a large number of cellular signaling pathways, sometimes coupling the signaling to ciliary motility or alternatively to cell division and differentiation. This view has had unanticipated consequences for our understanding of developmental processes and human disease.

  2. Structure and function of mammalian cilia

    PubMed Central

    Satir, Peter

    2008-01-01

    In the past half century, beginning with electron microscopic studies of 9 + 2 motile and 9 + 0 primary cilia, novel insights have been obtained regarding the structure and function of mammalian cilia. All cilia can now be viewed as sensory cellular antennae that coordinate a large number of cellular signaling pathways, sometimes coupling the signaling to ciliary motility or alternatively to cell division and differentiation. This view has had unanticipated consequences for our understanding of developmental processes and human disease. PMID:18365235

  3. Axoneme Structure from Motile Cilia.

    PubMed

    Ishikawa, Takashi

    2017-01-03

    The axoneme is the main extracellular part of cilia and flagella in eukaryotes. It consists of a microtubule cytoskeleton, which normally comprises nine doublets. In motile cilia, dynein ATPase motor proteins generate sliding motions between adjacent microtubules, which are integrated into a well-orchestrated beating or rotational motion. In primary cilia, there are a number of sensory proteins functioning on membranes surrounding the axoneme. In both cases, as the study of proteomics has elucidated, hundreds of proteins exist in this compartmentalized biomolecular system. In this article, we review the recent progress of structural studies of the axoneme and its components using electron microscopy and X-ray crystallography, mainly focusing on motile cilia. Structural biology presents snapshots (but not live imaging) of dynamic structural change and gives insights into the force generation mechanism of dynein, ciliary bending mechanism, ciliogenesis, and evolution of the axoneme.

  4. Ion channels and calcium signaling in motile cilia

    PubMed Central

    Doerner, Julia F; Delling, Markus; Clapham, David E

    2015-01-01

    The beating of motile cilia generates fluid flow over epithelia in brain ventricles, airways, and Fallopian tubes. Here, we patch clamp single motile cilia of mammalian ependymal cells and examine their potential function as a calcium signaling compartment. Resting motile cilia calcium concentration ([Ca2+] ~170 nM) is only slightly elevated over cytoplasmic [Ca2+] (~100 nM) at steady state. Ca2+ changes that arise in the cytoplasm rapidly equilibrate in motile cilia. We measured CaV1 voltage-gated calcium channels in ependymal cells, but these channels are not specifically enriched in motile cilia. Membrane depolarization increases ciliary [Ca2+], but only marginally alters cilia beating and cilia-driven fluid velocity within short (~1 min) time frames. We conclude that beating of ependymal motile cilia is not tightly regulated by voltage-gated calcium channels, unlike that of well-studied motile cilia and flagella in protists, such as Paramecia and Chlamydomonas. DOI: http://dx.doi.org/10.7554/eLife.11066.001 PMID:26650848

  5. Electrical Signaling in Motile and Primary Cilia

    PubMed Central

    Kleene, Steven J.; Van Houten, Judith L.

    2014-01-01

    Cilia are highly conserved for their structure and also for their sensory functions. They serve as antennae for extracellular information. Whether the cilia are motile or not, they respond to environmental mechanical and chemical stimuli and send signals to the cell body. The information from extracellular stimuli is commonly converted to electrical signals through the repertoire of ion-conducting channels in the ciliary membrane, which results in changes in concentrations of ions, especially calcium ions, in the cilia. These changes, in turn, affect motility and the ability of the signaling pathways in the cilia and cell body to carry on the signal transduction. We review here the activities of ion channels in cilia in animals from protists to vertebrates. PMID:25892740

  6. Overview of structure and function of mammalian cilia.

    PubMed

    Satir, Peter; Christensen, Søren Tvorup

    2007-01-01

    Cilia are membrane-bounded, centriole-derived projections from the cell surface that contain a microtubule cytoskeleton, the ciliary axoneme, surrounded by a ciliary membrane. Axonemes in multiciliated cells of mammalian epithelia are 9 + 2, possess dynein arms, and are motile. In contrast, single nonmotile 9 + 0 primary cilia are found on epithelial cells, such as those of the kidney tubule, but also on nonepithelial cells, such as chondrocytes, fibroblasts, and neurons. The ciliary membranes of all cilia contain specific receptors and ion channel proteins that initiate signaling pathways controlling motility and/or linking mechanical or chemical stimuli, including sonic hedgehog and growth factors, to intracellular transduction cascades regulating differentiation, migration, and cell growth during development and in adulthood. Unique motile 9 + 0 cilia, found during development at the embryonic node, determine left-right asymmetry of the body.

  7. Realizing the Physics of Motile Cilia Synchronization with Driven Colloids

    NASA Astrophysics Data System (ADS)

    Bruot, Nicolas; Cicuta, Pietro

    2016-03-01

    Cilia and flagella in biological systems often show large scale cooperative behaviors such as the synchronization of their beats in "metachronal waves." These are beautiful examples of emergent dynamics in biology, and are essential for life, allowing diverse processes from the motility of eukaryotic microorganisms, to nutrient transport and clearance of pathogens from mammalian airways. How these collective states arise is not fully understood, but it is clear that individual cilia interact mechanically, and that a strong and long-ranged component of the coupling is mediated by the viscous fluid. We review here the work by ourselves and others aimed at understanding the behavior of hydrodynamically coupled systems, and particularly a set of results that have been obtained both experimentally and theoretically by studying actively driven colloidal systems. In these controlled scenarios, it is possible to selectively test aspects of living motile cilia, such as the geometrical arrangement, the effects of the driving profile and the distance to no-slip boundaries. We outline and give examples of how it is possible to link model systems to observations on living systems, which can be made on microorganisms, on cell cultures or on tissue sections. This area of research has clear clinical application in the long term, as severe pathologies are associated with compromised cilia function in humans.

  8. Mutation of Growth Arrest Specific 8 Reveals a Role in Motile Cilia Function and Human Disease

    PubMed Central

    Lewis, Wesley R.; Malarkey, Erik B.; Tritschler, Douglas; Bower, Raqual; Pasek, Raymond C.; Porath, Jonathan D.; Birket, Susan E.; Saunier, Sophie; Antignac, Corinne; Leigh, Margaret W.; Zariwala, Maimoona A.; Drummond, Iain A.; Parant, John M.; Hildebrandt, Friedhelm; Yoder, Bradley K.

    2016-01-01

    Ciliopathies are genetic disorders arising from dysfunction of microtubule-based cellular appendages called cilia. Different cilia types possess distinct stereotypic microtubule doublet arrangements with non-motile or ‘primary’ cilia having a 9+0 and motile cilia have a 9+2 array of microtubule doublets. Primary cilia are critical sensory and signaling centers needed for normal mammalian development. Defects in their structure/function result in a spectrum of clinical and developmental pathologies including abnormal neural tube and limb patterning. Altered patterning phenotypes in the limb and neural tube are due to perturbations in the hedgehog (Hh) signaling pathway. Motile cilia are important in fluid movement and defects in motility result in chronic respiratory infections, altered left-right asymmetry, and infertility. These features are the hallmarks of Primary Ciliary Dyskinesia (PCD, OMIM 244400). While mutations in several genes are associated with PCD in patients and animal models, the genetic lesion in many cases is unknown. We assessed the in vivo functions of Growth Arrest Specific 8 (GAS8). GAS8 shares strong sequence similarity with the Chlamydomonas Nexin-Dynein Regulatory Complex (NDRC) protein 4 (DRC4) where it is needed for proper flagella motility. In mammalian cells, the GAS8 protein localizes not only to the microtubule axoneme of motile cilia, but also to the base of non-motile cilia. Gas8 was recently implicated in the Hh signaling pathway as a regulator of Smoothened trafficking into the cilium. Here, we generate the first mouse with a Gas8 mutation and show that it causes severe PCD phenotypes; however, there were no overt Hh pathway phenotypes. In addition, we identified two human patients with missense variants in Gas8. Rescue experiments in Chlamydomonas revealed a subtle defect in swim velocity compared to controls. Further experiments using CRISPR/Cas9 homology driven repair (HDR) to generate one of these human missense variants

  9. Adenylate cyclase regulates elongation of mammalian primary cilia

    SciTech Connect

    Ou, Young; Ruan, Yibing; Cheng, Min; Moser, Joanna J.; Rattner, Jerome B.; Hoorn, Frans A. van der

    2009-10-01

    The primary cilium is a non-motile microtubule-based structure that shares many similarities with the structures of flagella and motile cilia. It is well known that the length of flagella is under stringent control, but it is not known whether this is true for primary cilia. In this study, we found that the length of primary cilia in fibroblast-like synoviocytes, either in log phase culture or in quiescent state, was confined within a range. However, when lithium was added to the culture to a final concentration of 100 mM, primary cilia of synoviocytes grew beyond this range, elongating to a length that was on average approximately 3 times the length of untreated cilia. Lithium is a drug approved for treating bipolar disorder. We dissected the molecular targets of this drug, and observed that inhibition of adenylate cyclase III (ACIII) by specific inhibitors mimicked the effects of lithium on primary cilium elongation. Inhibition of GSK-3{beta} by four different inhibitors did not induce primary cilia elongation. ACIII was found in primary cilia of a variety of cell types, and lithium treatment of these cell types led to their cilium elongation. Further, we demonstrate that different cell types displayed distinct sensitivities to the lithium treatment. However, in all cases examined primary cilia elongated as a result of lithium treatment. In particular, two neuronal cell types, rat PC-12 adrenal medulla cells and human astrocytes, developed long primary cilia when lithium was used at or close to the therapeutic relevant concentration (1-2 mM). These results suggest that the length of primary cilia is controlled, at least in part, by the ACIII-cAMP signaling pathway.

  10. Inactivation of Chibby affects function of motile airway cilia

    PubMed Central

    Voronina, Vera A.; Treuting, Piper; Love, Damon; Grubb, Barbara R.; Hajjar, Adeline M.; Adams, Allison; Li, Feng-Qian; Moon, Randall T.

    2009-01-01

    Chibby (Cby) is a conserved component of the Wnt–β-catenin pathway. Cby physically interacts with β-catenin to repress its activation of transcription. To elucidate the function of Cby in vertebrates, we generated Cby−/− mice and found that after 2–3 d of weight loss, the majority of mice die before or around weaning. All Cby−/− mice develop rhinitis and sinusitis. When challenged with Pseudomonas aeruginosa isolates, Cby−/− mice are unable to clear the bacteria from the nasal cavity. Notably, Cby−/− mice exhibit a complete absence of mucociliary transport caused by a marked paucity of motile cilia in the nasal epithelium. Moreover, ultrastructural experiments reveal impaired basal body docking to the apical surface of multiciliated cells. In support of these phenotypes, endogenous Cby protein is localized at the base of cilia. As the phenotypes of Cby−/− mice bear striking similarities to primary ciliary dyskinesia, Cby−/− mice may prove to be a useful model for this condition. PMID:19364920

  11. The coiled-coil domain containing protein CCDC151 is required for the function of IFT-dependent motile cilia in animals.

    PubMed

    Jerber, Julie; Baas, Dominique; Soulavie, Fabien; Chhin, Brigitte; Cortier, Elisabeth; Vesque, Christine; Thomas, Joëlle; Durand, Bénédicte

    2014-02-01

    Cilia are evolutionarily conserved organelles endowed with essential physiological and developmental functions. In humans, disruption of cilia motility or signaling leads to complex pleiotropic genetic disorders called ciliopathies. Cilia motility requires the assembly of multi-subunit motile components such as dynein arms, but mechanisms underlying their assembly pathway and transport into the axoneme are still largely unknown. We identified a previously uncharacterized coiled-coil domain containing protein CCDC151, which is evolutionarily conserved in motile ciliated species and shares ancient features with the outer dynein arm-docking complex 2 of Chlamydomonas. In Drosophila, we show that CG14127/CCDC151 is associated with motile intraflagellar transport (IFT)-dependent cilia and required for geotaxis behavior of adult flies. In zebrafish, Ccdc151 is expressed in tissues with motile cilia, and morpholino-induced depletion of Ccdc151 leads to left-right asymmetry defects and kidney cysts. We demonstrate that Ccdc151 is required for proper motile function of cilia in the Kupffer's vesicle and in the pronephros by controlling dynein arm assembly, showing that Ccdc151 is a novel player in the control of IFT-dependent dynein arm assembly in animals. However, we observed that CCDC151 is also implicated in other cellular functions in vertebrates. In zebrafish, ccdc151 is involved in proper orientation of cell divisions in the pronephros and genetically interacts with prickle1 in this process. Furthermore, knockdown experiments in mammalian cells demonstrate that CCDC151 is implicated in the regulation of primary cilium length. Hence, CCDC151 is required for motile cilia function in animals but has acquired additional non-motile functions in vertebrates.

  12. Motile and non-motile cilia in human pathology: from function to phenotypes.

    PubMed

    Mitchison, Hannah M; Valente, Enza Maria

    2017-01-01

    Ciliopathies are inherited human disorders caused by both motile and non-motile cilia dysfunction that form an important and rapidly expanding disease category. Ciliopathies are complex conditions to diagnose, being multisystem disorders characterized by extensive genetic heterogeneity and clinical variability with high levels of lethality. There is marked phenotypic overlap among distinct ciliopathy syndromes that presents a major challenge for their recognition, diagnosis, and clinical management, in addition to posing an on-going task to develop the most appropriate family counselling. The impact of next-generation sequencing and high-throughput technologies in the last decade has significantly improved our understanding of the biological basis of ciliopathy disorders, enhancing our ability to determine the possible reasons for the extensive overlap in their symptoms and genetic aetiologies. Here, we review the diverse functions of cilia in human health and disease and discuss a growing shift away from the classical clinical definitions of ciliopathy syndromes to a more functional categorization. This approach arises from our improved understanding of this unique organelle, revealed through new genetic and cell biological insights into the discrete functioning of subcompartments of the cilium (basal body, transition zone, intraflagellar transport, motility). Mutations affecting these distinct ciliary protein modules can confer different genetic diseases and new clinical classifications are possible to define, according to the nature and extent of organ involvement. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  13. Gene therapy rescues cilia defects and restores olfactory function in a mammalian ciliopathy model.

    PubMed

    McIntyre, Jeremy C; Davis, Erica E; Joiner, Ariell; Williams, Corey L; Tsai, I-Chun; Jenkins, Paul M; McEwen, Dyke P; Zhang, Lian; Escobado, John; Thomas, Sophie; Szymanska, Katarzyna; Johnson, Colin A; Beales, Philip L; Green, Eric D; Mullikin, James C; Sabo, Aniko; Muzny, Donna M; Gibbs, Richard A; Attié-Bitach, Tania; Yoder, Bradley K; Reed, Randall R; Katsanis, Nicholas; Martens, Jeffrey R

    2012-09-01

    Cilia are evolutionarily conserved microtubule-based organelles that are crucial for diverse biological functions, including motility, cell signaling and sensory perception. In humans, alterations in the formation and function of cilia manifest clinically as ciliopathies, a growing class of pleiotropic genetic disorders. Despite the substantial progress that has been made in identifying genes that cause ciliopathies, therapies for these disorders are not yet available to patients. Although mice with a hypomorphic mutation in the intraflagellar transport protein IFT88 (Ift88Tg737Rpw mice, also known as ORPK mice)5 have been well studied, the relevance of IFT88 mutations to human pathology is unknown. We show that a mutation in IFT88 causes a hitherto unknown human ciliopathy. In vivo complementation assays in zebrafish and mIMCD3 cells show the pathogenicity of this newly discovered allele. We further show that ORPK mice are functionally anosmic as a result of the loss of cilia on their olfactory sensory neurons (OSNs). Notably, adenoviral-mediated expression of IFT88 in mature, fully differentiated OSNs of ORPK mice is sufficient to restore ciliary structures and rescue olfactory function. These studies are the first to use in vivo therapeutic treatment to reestablish cilia in a mammalian ciliopathy. More broadly, our studies indicate that gene therapy is a viable option for cellular and functional rescue of the complex ciliary organelle in established differentiated cells.

  14. Primary cilia and signaling pathways in mammalian development, health and disease

    PubMed Central

    VELAND, IBEN R.; AWAN, AASHIR; PEDERSEN, LOTTE B.; YODER, BRADLEY K.; CHRISTENSEN, SØREN T.

    2010-01-01

    SUMMARY Although first described 1898 and long considered a vestigial organelle of little functional importance, the primary cilium has become one of the hottest research topics in modern cell biology and physiology. Primary cilia are non-motile sensory organelles present in a single copy on the surface of most growth-arrested or differentiated mammalian cells, and defects in their assembly or function are tightly coupled to many developmental defects, diseases and disorders. In normal tissues the primary cilium coordinates a series of signal transduction pathways, including Hedgehog, Wnt, PDGFRα and integrin signaling. In the kidney the primary cilium may function as a mechano-, chemo- and osmosensing unit that probes the extracellular environment and transmits signals to the cell via e.g. polycystins, which depend on ciliary localization for appropriate function. Indeed, hypomorphic mutations in the mouse ift88 (previously called Tg737) gene, which encodes a ciliogenic intraflagellar transport (IFT) protein, result in malformation of primary cilia, and in the collecting ducts of kidney tubules this is accompanied by development of autosomal recessive polycystic kidney disease (PKD; (1)). While PKD was one of the first diseases to be linked to dysfunctional primary cilia, defects in this organelle have subsequently been associated with many other phenotypes, including cancer, obesity, diabetes as well as a number of developmental defects. Collectively, these disorders of the cilium are now referred to as the ciliopathies. In this review we provide a brief overview of the structure and function of primary cilia and some of their roles in coordinating signal transduction pathways in mammalian development, health and disease. This review was written in conjunction with the Takis Anagnostopoulos Symposium on Renal and Epithelial Physiology and Physiopathology at Faculté de Médecine Necker in Paris, June 26-27, 2008. PMID:19276629

  15. A bio-inspired inner-motile photocatalyst film: a magnetically actuated artificial cilia photocatalyst

    NASA Astrophysics Data System (ADS)

    Zhang, Dunpu; Wang, Wei; Peng, Fengping; Kou, Jiahui; Ni, Yaru; Lu, Chunhua; Xu, Zhongzi

    2014-04-01

    A new type of inner-motile photocatalyst film is explored to enhance photocatalytic performance using magnetically actuated artificial cilia. The inner-motile photocatalyst film is capable of generating flow and mixing on the microscale because it produces a motion similar to that of natural cilia when it is subjected to a rotational magnetic field. Compared with traditional photocatalyst films, the inner-motile photocatalyst film exhibits the unique ability of microfluidic manipulation. It uses an impactful and self-contained design to accelerate interior mass transfer and desorption of degradation species. Moreover, the special cilia-like structures increase the surface area and light absorption. Consequently, the photocatalytic activity of the inner-motile photocatalyst film is dramatically improved to approximately 3.0 times that of the traditional planar film. The inner-motile photocatalyst film also exhibits high photocatalytic durability and can be reused several times with ease. Furthermore, this feasible yet versatile platform can be extended to other photocatalyst systems, such as TiO2, P25, ZnO, and Co3O4 systems, to improve their photocatalytic performance.A new type of inner-motile photocatalyst film is explored to enhance photocatalytic performance using magnetically actuated artificial cilia. The inner-motile photocatalyst film is capable of generating flow and mixing on the microscale because it produces a motion similar to that of natural cilia when it is subjected to a rotational magnetic field. Compared with traditional photocatalyst films, the inner-motile photocatalyst film exhibits the unique ability of microfluidic manipulation. It uses an impactful and self-contained design to accelerate interior mass transfer and desorption of degradation species. Moreover, the special cilia-like structures increase the surface area and light absorption. Consequently, the photocatalytic activity of the inner-motile photocatalyst film is dramatically

  16. The Kinesin-4 Protein KIF7 Regulates Mammalian Hedgehog Signaling by Organizing the Cilia Tip Compartment

    PubMed Central

    He, Mu; Subramanian, Radhika; Bangs, Fiona; Omelchenko, Tatiana; Liem, Karel F.; Kapoor, Tarun M.; Anderson, Kathryn V.

    2014-01-01

    Mammalian Hedgehog (Hh) signal transduction requires the primary cilium, a microtubule-based organelle, and the Gli/Sufu complexes that mediate Hh signaling are enriched at cilia tips. KIF7, a kinesin-4 family protein, is a conserved regulator of the Hh signaling pathway and a human ciliopathy protein. Here we show that KIF7 localizes to cilia tips, the site of microtubule plus-ends, where it limits cilia length and controls cilia structure. Purified recombinant KIF7 binds the plus-ends of growing microtubules in vitro, where it reduces the rate of microtubule growth and increases the frequency of microtubule catastrophe. KIF7 is not required for normal intraflagellar transport or for trafficking of Hh pathway proteins into cilia. Instead, a central function of KIF7 in the mammalian Hh pathway is to control cilia architecture and to create a single cilia tip compartment where Gli/Sufu activation can be correctly regulated. PMID:24952464

  17. Mammalian Sperm Motility: Observation and Theory

    NASA Astrophysics Data System (ADS)

    Gaffney, E. A.; Gadêlha, H.; Smith, D. J.; Blake, J. R.; Kirkman-Brown, J. C.

    2011-01-01

    Mammalian spermatozoa motility is a subject of growing importance because of rising human infertility and the possibility of improving animal breeding. We highlight opportunities for fluid and continuum dynamics to provide novel insights concerning the mechanics of these specialized cells, especially during their remarkable journey to the egg. The biological structure of the motile sperm appendage, the flagellum, is described and placed in the context of the mechanics underlying the migration of mammalian sperm through the numerous environments of the female reproductive tract. This process demands certain specific changes to flagellar movement and motility for which further mechanical insight would be valuable, although this requires improved modeling capabilities, particularly to increase our understanding of sperm progression in vivo. We summarize current theoretical studies, highlighting the synergistic combination of imaging and theory in exploring sperm motility, and discuss the challenges for future observational and theoretical studies in understanding the underlying mechanics.

  18. Cryo-electron tomography of motile cilia and flagella.

    PubMed

    Ishikawa, Takashi

    2015-01-01

    Cryo-electron tomography has been a valuable tool in the analysis of 3D structures of cilia at molecular and cellular levels. It opened a way to reconstruct 3D conformations of proteins in cilia at 3-nm resolution, revealed networks of a number of component proteins in cilia, and has even allowed the study of component dynamics. In particular, we have identified the locations and conformations of all the regular inner and outer dyneins, as well as various regulators such as radial spokes. Since the mid 2000s, cryo-electron tomography has provided us with new knowledge, concepts, and questions in the area of cilia research. Now, after nearly 10 years of application of this technique, we are turning a corner and are at the stage to discuss the next steps. We expect further development of this technique for specimen preparation, data acquisition, and analysis. While combining this tool with other methodologies has already made cryo-electron tomography more biologically significant, we need to continue this cooperation using recently developed biotechnology and cell biology approaches. In this review, we will provide an up-to-date overview of the biological insights obtained by cryo-electron tomography and will discuss future possibilities of this technique in the context of cilia research.

  19. Sperm-Associated Antigen–17 Gene Is Essential for Motile Cilia Function and Neonatal Survival

    PubMed Central

    Teves, Maria Eugenia; Zhang, Zhibing; Costanzo, Richard M.; Henderson, Scott C.; Corwin, Frank D.; Zweit, Jamal; Sundaresan, Gobalakrishnan; Subler, Mark; Salloum, Fadi N.; Rubin, Bruce K.

    2013-01-01

    Primary ciliary dyskinesia (PCD), resulting from defects in cilia assembly or motility, is caused by mutations in a number of genes encoding axonemal proteins. PCD phenotypes are variable, and include recurrent respiratory tract infections, bronchiectasis, hydrocephaly, situs inversus, and male infertility. We generated knockout mice for the sperm-associated antigen–17 (Spag17) gene, which encodes a central pair (CP) protein present in the axonemes of cells with “9 + 2” motile cilia or flagella. The targeting of Spag17 resulted in a severe phenotype characterized by immotile nasal and tracheal cilia, reduced clearance of nasal mucus, profound respiratory distress associated with lung fluid accumulation and disruption of the alveolar epithelium, cerebral ventricular expansion consistent with emerging hydrocephalus, failure to suckle, and neonatal demise within 12 hours of birth. Ultrastructural analysis revealed the loss of one CP microtubule in approximately one quarter of tracheal cilia axonemes, an absence of a C1 microtubule projection, and other less frequent CP structural abnormalities. SPAG6 and SPAG16 (CP proteins that interact with SPAG17) were increased in tracheal tissue from SPAG17-deficient mice. We conclude that Spag17 plays a critical role in the function and structure of motile cilia, and that neonatal lethality is likely explained by impaired airway mucociliary clearance. PMID:23418344

  20. Loss of Dishevelleds disrupts planar polarity in ependymal motile cilia and results in hydrocephalus.

    PubMed

    Ohata, Shinya; Nakatani, Jin; Herranz-Pérez, Vicente; Cheng, JrGang; Belinson, Haim; Inubushi, Toshiro; Snider, William D; García-Verdugo, Jose Manuel; Wynshaw-Boris, Anthony; Alvarez-Buylla, Arturo

    2014-08-06

    Defects in ependymal (E) cells, which line the ventricle and generate cerebrospinal fluid flow through ciliary beating, can cause hydrocephalus. Dishevelled genes (Dvls) are essential for Wnt signaling, and Dvl2 has been shown to localize to the rootlet of motile cilia. Using the hGFAP-Cre;Dvl1(-/-);2(flox/flox);3(+/-) mouse, we show that compound genetic ablation of Dvls causes hydrocephalus. In hGFAP-Cre;Dvl1(-/-);2(flox/flox);3(+/-) mutants, E cells differentiated normally, but the intracellular and intercellular rotational alignments of ependymal motile cilia were disrupted. As a consequence, the fluid flow generated by the hGFAP-Cre;Dvl1(-/-);2(flox/flox);3(+/-) E cells was significantly slower than that observed in control mice. Dvls were also required for the proper positioning of motile cilia on the apical surface. Tamoxifen-induced conditional removal of Dvls in adult mice also resulted in defects in intracellular rotational alignment and positioning of ependymal motile cilia. These results suggest that Dvls are continuously required for E cell planar polarity and may prevent hydrocephalus.

  1. Loss of Dishevelleds disrupts planar polarity in ependymal motile cilia and results in hydrocephalus

    PubMed Central

    Ohata, Shinya; Nakatani, Jin; Herranz-Pérez, Vicente; Cheng, JrGang; Belinson, Haim; Inubushi, Toshiro; Snider, William D.; García-Verdugo, Jose Manuel; Wynshaw-Boris, Anthony; Álvarez-Buylla, Arturo

    2014-01-01

    SUMMARY Defects in ependymal (E) cells, which line the ventricle and generate cerebrospinal fluid flow through ciliary beating, can cause hydrocephalus. Dishevelled genes (Dvls) are essential for Wnt signaling and Dvl2 has been shown to localize to the rootlet of motile cilia. Using the hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− mouse, we show that compound genetic ablation of Dvls causes hydrocephalus. In hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− mutants, E cells differentiated normally, but the intracellular and intercellular rotational alignments of ependymal motile cilia were disrupted. As a consequence, the fluid flow generated by the hGFAP-Cre;Dvl1−/−;2flox/flox;3+/− E cells was significantly slower than that observed in control mice. Dvls were also required for the proper positioning of motile cilia on the apical surface. Tamoxifen-induced conditional removal of Dvls in adult mice also resulted in defects in intracellular rotational alignment and positioning of ependymal motile cilia. These results suggest that Dvls are continuously required for E cell planar polarity and may prevent hydrocephalus. PMID:25043421

  2. Motile cilia harbor serum response factor as a mechanism of environment sensing and injury response in the airway.

    PubMed

    Nordgren, Tara M; Wyatt, Todd A; Sweeter, Jenea; Bailey, Kristina L; Poole, Jill A; Heires, Art J; Sisson, Joseph H; Romberger, Debra J

    2014-05-01

    Nonmotile primary cilia are recognized as important sensory organelles during development and normal biological functioning. For example, recent work demonstrates that transcriptional regulators of the sonic hedgehog signaling pathway localize to primary cilia and participate in sensing and transducing signals regarding the cellular environment. In contrast, motile cilia are traditionally viewed as mechanical machinery, vital for the movement of solutes and clearance of bacteria and debris, but not participants in cellular sensing and signaling mechanisms. Recently, motile cilia were found to harbor receptors responsible for sensing and responding to environmental stimuli. However, no transcription factors are known to be regulated by cilia localization as a sensing mechanism in vertebrates. Using a mouse model of organic dust-induced airway inflammation, we found that the transcription factor serum response factor (SRF) localizes to motile cilia of airway epithelial cells and alters its localization in response to inflammatory stimuli. Furthermore, inhibition of SRF signaling using the small molecule CCG-1423 reduces organic dust-induced IL-8 release from bronchial epithelial cells and stimulates cilia beat frequency in ciliated mouse tracheal epithelial cells. Immunohistochemical analyses reveal that SRF localizes to the cilia of mouse brain ependymal and ovarian epithelial cells as well. These data reveal a novel mechanism by which a transcription factor localizes to motile cilia and modulates cell activities including cilia motility and inflammation response. These data challenge current dogma regarding motile cilia functioning and may lead to significant contributions in understanding motile ciliary signaling dynamics, as well as mechanisms involving SRF-mediated responses to inflammation and injury.

  3. Reptin/Ruvbl2 is a Lrrc6/Seahorse interactor essential for cilia motility.

    PubMed

    Zhao, Lu; Yuan, Shiaulou; Cao, Ying; Kallakuri, Sowjanya; Li, Yuanyuan; Kishimoto, Norihito; DiBella, Linda; Sun, Zhaoxia

    2013-07-30

    Primary ciliary dyskinesia (PCD) is an autosomal recessive disease caused by defective cilia motility. The identified PCD genes account for about half of PCD incidences and the underlying mechanisms remain poorly understood. We demonstrate that Reptin/Ruvbl2, a protein known to be involved in epigenetic and transcriptional regulation, is essential for cilia motility in zebrafish. We further show that Reptin directly interacts with the PCD protein Lrrc6/Seahorse and this interaction is critical for the in vivo function of Lrrc6/Seahorse in zebrafish. Moreover, whereas the expression levels of multiple dynein arm components remain unchanged or become elevated, the density of axonemal dynein arms is reduced in reptin(hi2394) mutants. Furthermore, Reptin is highly enriched in the cytosol and colocalizes with Lrrc6/Seahorse. Combined, these results suggest that the Reptin-Lrrc6/Seahorse complex is involved in dynein arm formation. We also show that although the DNA damage response is induced in reptin(hi2394) mutants, it remains unchanged in cilia biogenesis mutants and lrrc6/seahorse mutants, suggesting that increased DNA damage response is not intrinsic to ciliary defects and that in vertebrate development, Reptin functions in multiple processes, both cilia specific and cilia independent.

  4. Reptin/Ruvbl2 is a Lrrc6/Seahorse interactor essential for cilia motility

    PubMed Central

    Zhao, Lu; Yuan, Shiaulou; Cao, Ying; Kallakuri, Sowjanya; Li, Yuanyuan; Kishimoto, Norihito; DiBella, Linda; Sun, Zhaoxia

    2013-01-01

    Primary ciliary dyskinesia (PCD) is an autosomal recessive disease caused by defective cilia motility. The identified PCD genes account for about half of PCD incidences and the underlying mechanisms remain poorly understood. We demonstrate that Reptin/Ruvbl2, a protein known to be involved in epigenetic and transcriptional regulation, is essential for cilia motility in zebrafish. We further show that Reptin directly interacts with the PCD protein Lrrc6/Seahorse and this interaction is critical for the in vivo function of Lrrc6/Seahorse in zebrafish. Moreover, whereas the expression levels of multiple dynein arm components remain unchanged or become elevated, the density of axonemal dynein arms is reduced in reptinhi2394 mutants. Furthermore, Reptin is highly enriched in the cytosol and colocalizes with Lrrc6/Seahorse. Combined, these results suggest that the Reptin-Lrrc6/Seahorse complex is involved in dynein arm formation. We also show that although the DNA damage response is induced in reptinhi2394 mutants, it remains unchanged in cilia biogenesis mutants and lrrc6/seahorse mutants, suggesting that increased DNA damage response is not intrinsic to ciliary defects and that in vertebrate development, Reptin functions in multiple processes, both cilia specific and cilia independent. PMID:23858445

  5. A bio-inspired inner-motile photocatalyst film: a magnetically actuated artificial cilia photocatalyst.

    PubMed

    Zhang, Dunpu; Wang, Wei; Peng, Fengping; Kou, Jiahui; Ni, Yaru; Lu, Chunhua; Xu, Zhongzi

    2014-05-21

    A new type of inner-motile photocatalyst film is explored to enhance photocatalytic performance using magnetically actuated artificial cilia. The inner-motile photocatalyst film is capable of generating flow and mixing on the microscale because it produces a motion similar to that of natural cilia when it is subjected to a rotational magnetic field. Compared with traditional photocatalyst films, the inner-motile photocatalyst film exhibits the unique ability of microfluidic manipulation. It uses an impactful and self-contained design to accelerate interior mass transfer and desorption of degradation species. Moreover, the special cilia-like structures increase the surface area and light absorption. Consequently, the photocatalytic activity of the inner-motile photocatalyst film is dramatically improved to approximately 3.0 times that of the traditional planar film. The inner-motile photocatalyst film also exhibits high photocatalytic durability and can be reused several times with ease. Furthermore, this feasible yet versatile platform can be extended to other photocatalyst systems, such as TiO2, P25, ZnO, and Co3O4 systems, to improve their photocatalytic performance.

  6. An outer arm Dynein conformational switch is required for metachronal synchrony of motile cilia in planaria.

    PubMed

    Rompolas, Panteleimon; Patel-King, Ramila S; King, Stephen M

    2010-11-01

    Motile cilia mediate the flow of mucus and other fluids across the surface of specialized epithelia in metazoans. Efficient clearance of peri-ciliary fluids depends on the precise coordination of ciliary beating to produce metachronal waves. The role of individual dynein motors and the mechanical feedback mechanisms required for this process are not well understood. Here we used the ciliated epithelium of the planarian Schmidtea mediterranea to dissect the role of outer arm dynein motors in the metachronal synchrony of motile cilia. We demonstrate that animals that completely lack outer dynein arms display a significant decline in beat frequency and an inability of cilia to coordinate their oscillations and form metachronal waves. Furthermore, lack of a key mechanosensitive regulatory component (LC1) yields a similar phenotype even though outer arms still assemble in the axoneme. The lack of metachrony was not due simply to a decrease in ciliary beat frequency, as reducing this parameter by altering medium viscosity did not affect ciliary coordination. In addition, we did not observe a significant temporal variability in the beat cycle of impaired cilia. We propose that this conformational switch provides a mechanical feedback system within outer arm dynein that is necessary to entrain metachronal synchrony.

  7. An Outer Arm Dynein Conformational Switch Is Required for Metachronal Synchrony of Motile Cilia in Planaria

    PubMed Central

    Rompolas, Panteleimon; Patel-King, Ramila S.

    2010-01-01

    Motile cilia mediate the flow of mucus and other fluids across the surface of specialized epithelia in metazoans. Efficient clearance of peri-ciliary fluids depends on the precise coordination of ciliary beating to produce metachronal waves. The role of individual dynein motors and the mechanical feedback mechanisms required for this process are not well understood. Here we used the ciliated epithelium of the planarian Schmidtea mediterranea to dissect the role of outer arm dynein motors in the metachronal synchrony of motile cilia. We demonstrate that animals that completely lack outer dynein arms display a significant decline in beat frequency and an inability of cilia to coordinate their oscillations and form metachronal waves. Furthermore, lack of a key mechanosensitive regulatory component (LC1) yields a similar phenotype even though outer arms still assemble in the axoneme. The lack of metachrony was not due simply to a decrease in ciliary beat frequency, as reducing this parameter by altering medium viscosity did not affect ciliary coordination. In addition, we did not observe a significant temporal variability in the beat cycle of impaired cilia. We propose that this conformational switch provides a mechanical feedback system within outer arm dynein that is necessary to entrain metachronal synchrony. PMID:20844081

  8. A Structural Basis for How Motile Cilia Beat

    PubMed Central

    Satir, Peter; Heuser, Thomas; Sale, Winfield S.

    2014-01-01

    The motile cilium is a mechanical wonder, a cellular nanomachine that produces a high-speed beat based on a cycle of bends that move along an axoneme made of 9+2 microtubules. The molecular motors, dyneins, power the ciliary beat. The dyneins are compacted into inner and outer dynein arms, whose activity is highly regulated to produce microtubule sliding and axonemal bending. The switch point hypothesis was developed long ago to account for how sliding in the presence of axonemal radial spoke–central pair interactions causes the ciliary beat. Since then, a new genetic, biochemical, and structural complexity has been discovered, in part, with Chlamydomonas mutants, with high-speed, high-resolution analysis of movement and with cryoelectron tomography. We stand poised on the brink of new discoveries relating to the molecular control of motility that extend and refine our understanding of the basic events underlying the switching of arm activity and of bend formation and propagation. PMID:26955066

  9. A Structural Basis for How Motile Cilia Beat.

    PubMed

    Satir, Peter; Heuser, Thomas; Sale, Winfield S

    2014-12-01

    The motile cilium is a mechanical wonder, a cellular nanomachine that produces a high-speed beat based on a cycle of bends that move along an axoneme made of 9+2 microtubules. The molecular motors, dyneins, power the ciliary beat. The dyneins are compacted into inner and outer dynein arms, whose activity is highly regulated to produce microtubule sliding and axonemal bending. The switch point hypothesis was developed long ago to account for how sliding in the presence of axonemal radial spoke-central pair interactions causes the ciliary beat. Since then, a new genetic, biochemical, and structural complexity has been discovered, in part, with Chlamydomonas mutants, with high-speed, high-resolution analysis of movement and with cryoelectron tomography. We stand poised on the brink of new discoveries relating to the molecular control of motility that extend and refine our understanding of the basic events underlying the switching of arm activity and of bend formation and propagation.

  10. Collective motion of motile cilia: from human airways to model systems

    NASA Astrophysics Data System (ADS)

    Cicuta, Pietro; Feriani, Luigi; Chioccioli, Maurizio; Kotar, Jurij

    Mammalian airways are a fantastic playground of nonlinear phenomena, from the function of individual active filaments, to the emerging collective behaviour, to the rheology of the mucus solution surrounding cilia. We have been investigating the fundamental physics of this system through a variety of model system approaches, both experimental and computational. In the last year we have started measurements on living human cells, observing cilia shape during beating, and measuring speed and coherence of the collective dynamics. We report on significant differences in the collective motion in ciliated cell carpets from a variety of diseases, and we attempt to reconcile the collective dynamical phenotypes to the properties of individual filaments and the mechanics of the environment.

  11. Lipopolysaccharide (LPS) disrupts particle transport, cilia function and sperm motility in an ex vivo oviduct model

    PubMed Central

    O’Doherty, A. M.; Di Fenza, M.; Kölle, S.

    2016-01-01

    The oviduct functions in the transportation of gametes to the site of fertilization (the ampulla) and is the site of early embryonic development. Alterations of this early developmental environment, such as the presence of sexually transmitted pathogens, may affect oviduct function leading to reduced fertilization rates and contribute to compromised embryonic development. In this study, sperm interactions, particle transport speed (PTS) and cilia beat frequency (CBF) in the ampulla following exposure to lipopolysaccharide (LPS), a constituent of the sexually transmitted pathogens Chlamydia trachomatis and Chlamydia abortus, was investigated. Three complementary experiments were performed to analyse; (1) bound sperm motility and cilia function (2) transport velocity in the oviduct and (3) the expression of genes related to immune function and inflammatory response (CASP3, CD14, MYD88, TLR4 and TRAF6). The motility of bound sperm was significantly lower in ampullae that were exposed to LPS. CBF and PTS significantly increased after treatment with LPS for 2 hours. Finally, gene expression analysis revealed that CASP3 and CD14 were significantly upregulated and TLR4 trended towards increased expression following treatment with LPS. These findings provide an insight on the impact of LPS on the oviduct sperm interaction, and have implications for both male and female fertility. PMID:27079521

  12. Loss of ASP but not ROPN1 reduces mammalian ciliary motility.

    PubMed

    Fiedler, Sarah E; Sisson, Joseph H; Wyatt, Todd A; Pavlik, Jacqueline A; Gambling, Todd M; Carson, Johnny L; Carr, Daniel W

    2012-01-01

    Protein kinase A (PKA) signaling is targeted by interactions with A-kinase anchoring proteins (AKAPs) via a dimerization/docking domain on the regulatory (R) subunit of PKA. Four other mammalian proteins [AKAP-associated sperm protein (ASP), ropporin (ROPN1), sperm protein 17 (SP17) and calcium binding tyrosine-(Y)-phosphorylation regulated protein (CABYR)] share this highly conserved RII dimerization/docking (R2D2) domain. ASP and ROPN1 are 41% identical in sequence, interact with a variety of AKAPs in a manner similar to PKA, and are expressed in ciliated and flagellated human cells. To test the hypothesis that these proteins regulate motility, we developed mutant mouse lines lacking ASP or ROPN1. Both mutant lines produced normal numbers of cilia with intact ciliary ultrastructure. Lack of ROPN1 had no effect on ciliary motility. However, the beat frequency of cilia from mice lacking ASP is significantly slower than wild type, indicating that ASP signaling may regulate ciliary motility. This is the first demonstration of in vivo function for ASP. Similar localization of ASP in mice and humans indicates that these findings may translate to human physiology, and that these mice will be an excellent model for future studies related to the pathogenesis of human disease.

  13. Loss of ASP but not ROPN1 reduces mammalian ciliary motility

    PubMed Central

    Fiedler, Sarah E.; Sisson, Joseph H.; Wyatt, Todd A.; Pavlik, Jacqueline A.; Gambling, Todd M.; Carson, Johnny L.; Carr, Daniel W.

    2011-01-01

    Protein kinase A (PKA) signaling is targeted by interactions with A-kinase anchoring proteins (AKAPs) via a dimerization/docking domain on the regulatory (R) subunit of PKA. Four other mammalian proteins (ASP, ROPN1, SP17, and CABYR) share this highly conserved RII dimerization/docking (R2D2) domain. ASP and ROPN1 are 41% identical in sequence, interact with a variety of AKAPs in a manner similar to PKA, and are expressed in ciliated and flagellated human cells. To test the hypothesis that these proteins regulate motility, we developed mutant mouse lines lacking ASP or ROPN1. Both mutant lines produced normal numbers of cilia with intact ciliary ultrastructure. Lack of ROPN1 had no effect on ciliary motility. However, the beat frequency of cilia from mice lacking ASP is significantly slower than wild type, indicating that ASP signaling may regulate ciliary motility. This is the first demonstration of in vivo function for ASP. Similar localization of ASP in mice and humans indicates that these findings may translate to human physiology, and that these mice will be an excellent model for future studies related to the pathogenesis of human disease. PMID:22021175

  14. Modulation of mammalian sperm motility by quercetin.

    PubMed

    Nass-Arden, L; Breitbart, H

    1990-04-01

    The flavonoid quercetin inhibits collective motility of ejaculated ram spermatozoa in the first 2 hr of incubation; during the next 3-4 hr motility is stimulated. To explain this interesting effect, we followed the influence of quercetin on sperm glycolysis, extracellular pH, ATP content, mitochondrial respiration, and lipid peroxidation. The collective motility of untreated cells is decreased to about 40% of the original motility during two hours of incubation. During this time, the rate of glycolysis is constant, respiration rate is increasing, there is no change in ATP content, the rate of lipid peroxidation is very slow, and the extracellular pH became very acidic (pH 5.5). It is concluded that motility is decreased due to this acidification. This acidification is prevented to some extent by quercetin, which indirectly inhibits glycolysis. Quercetin inhibits motility due to the inhibition of the plasma membrane calcium pump, as we showed previously (Breitbart et al., J Biol Chem 260:11548-11553, 1985). The motility of untreated cells is arrested after 3.5 hr of incubation, whereas quercetin-treated cells show high motility, which continues for additional 2-3 hr. After 3.5 hr, the control cells show no glycolytic activity, ATP content and respiration rates are decreased, and rate of lipid peroxidation is highly increased. At this time, quercetin-treated cells show no glycolytic activity, only a small decrease in ATP content and respiratory rate, and a very low rate of lipid peroxidation. Based on these data it is concluded that sperm motility after 3.5 hr of incubation is dependent mainly on mitochondrial respiration.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Regulation of Chlamydomonas flagella and ependymal cell motile cilia by ceramide-mediated translocation of GSK3.

    PubMed

    Kong, Ji Na; Hardin, Kara; Dinkins, Michael; Wang, Guanghu; He, Qian; Mujadzic, Tarik; Zhu, Gu; Bielawski, Jacek; Spassieva, Stefka; Bieberich, Erhard

    2015-12-01

    Cilia are important organelles formed by cell membrane protrusions; however, little is known about their regulation by membrane lipids. We characterize a novel activation mechanism for glycogen synthase kinase-3 (GSK3) by the sphingolipids phytoceramide and ceramide that is critical for ciliogenesis in Chlamydomonas and murine ependymal cells, respectively. We show for the first time that Chlamydomonas expresses serine palmitoyl transferase (SPT), the first enzyme in (phyto)ceramide biosynthesis. Inhibition of SPT in Chlamydomonas by myriocin led to loss of flagella and reduced tubulin acetylation, which was prevented by supplementation with the precursor dihydrosphingosine. Immunocytochemistry showed that (phyto)ceramide was colocalized with phospho-Tyr-216-GSK3 (pYGSK3) at the base and tip of Chlamydomonas flagella and motile cilia in ependymal cells. The (phyto)ceramide distribution was consistent with that of a bifunctional ceramide analogue UV cross-linked and visualized by click-chemistry-mediated fluorescent labeling. Ceramide depletion, by myriocin or neutral sphingomyelinase deficiency (fro/fro mouse), led to GSK3 dephosphorylation and defective flagella and cilia. Motile cilia were rescued and pYGSK3 localization restored by incubation of fro/fro ependymal cells with exogenous C24:1 ceramide, which directly bound to pYGSK3. Our findings suggest that (phyto)ceramide-mediated translocation of pYGSK into flagella and cilia is an evolutionarily conserved mechanism fundamental to the regulation of ciliogenesis.

  16. The PDZ Protein Na+/H+ Exchanger Regulatory Factor-1 (NHERF1) Regulates Planar Cell Polarity and Motile Cilia Organization

    PubMed Central

    Stolz, Donna B.; Tsang, Michael; Friedman, Peter A.; Romero, Guillermo

    2016-01-01

    Directional flow of the cerebrospinal fluid requires coordinated movement of the motile cilia of the ependymal epithelium that lines the cerebral ventricles. Here we report that mice lacking the Na+/H+ Exchanger Regulatory Factor 1 (NHERF1/Slc9a3r1, also known as EBP50) develop profound communicating hydrocephalus associated with fewer and disorganized ependymal cilia. Knockdown of NHERF1/slc9a3r1 in zebrafish embryos also causes severe hydrocephalus of the hindbrain and impaired ciliogenesis in the otic vesicle. Ultrastructural analysis did not reveal defects in the shape or organization of individual cilia. Similar phenotypes have been described in animals with deficiencies in Wnt signaling and the Planar Cell Polarity (PCP) pathway. We show that NHERF1 binds the PCP core genes Frizzled (Fzd) and Vangl. We further show that NHERF1 assembles a ternary complex with Fzd4 and Vangl2 and promotes translocation of Vangl2 to the plasma membrane, in particular to the apical surface of ependymal cells. Taken together, these results strongly support an important role for NHERF1 in the regulation of PCP signaling and the development of functional motile cilia. PMID:27055101

  17. A prefoldin-associated WD-repeat protein (WDR92) is required for the correct architectural assembly of motile cilia

    PubMed Central

    Patel-King, Ramila S.; King, Stephen M.

    2016-01-01

    WDR92 is a highly conserved WD-repeat protein that has been proposed to be involved in apoptosis and also to be part of a prefoldin-like cochaperone complex. We found that WDR92 has a phylogenetic signature that is generally compatible with it playing a role in the assembly or function of specifically motile cilia. To test this hypothesis, we performed an RNAi-based knockdown of WDR92 gene expression in the planarian Schmidtea mediterranea and were able to achieve a robust reduction in mRNA expression to levels undetectable under our standard RT-PCR conditions. We found that this treatment resulted in a dramatic reduction in the rate of organismal movement that was caused by a switch in the mode of locomotion from smooth, cilia-driven gliding to muscle-based, peristaltic contractions. Although the knockdown animals still assembled cilia of normal length and in similar numbers to controls, these structures had reduced beat frequency and did not maintain hydrodynamic coupling. By transmission electron microscopy we observed that many cilia had pleiomorphic defects in their architecture, including partial loss of dynein arms, incomplete closure of the B-tubule, and occlusion or replacement of the central pair complex by accumulated electron-dense material. These observations suggest that WDR92 is part of a previously unrecognized cytoplasmic chaperone system that is specifically required to fold key components necessary to build motile ciliary axonemes. PMID:26912790

  18. Notch/Her12 signalling modulates, motile/immotile cilia ratio downstream of Foxj1a in zebrafish left-right organizer.

    PubMed

    Tavares, Barbara; Jacinto, Raquel; Sampaio, Pedro; Pestana, Sara; Pinto, Andreia; Vaz, Andreia; Roxo-Rosa, Mónica; Gardner, Rui; Lopes, Telma; Schilling, Britta; Henry, Ian; Saúde, Leonor; Lopes, Susana Santos

    2017-09-06

    Foxj1a is necessary and sufficient to specify motile cilia. Using transcriptional studies and slow-scan two-photon live imaging capable of identifying the number of motile and immotile cilia, we now established that the final number of motile cilia depends on Notch signalling (NS). We found that despite all left-right organizer (LRO) cells express foxj1a and the ciliary axonemes of these cells have dynein arms, some cilia remain immotile. We identified that this decision is taken early in development in the Kupffer's Vesicle (KV) precursors the readout being her12 transcription. We demonstrate that overexpression of either her12 or Notch intracellular domain (NICD) increases the number of immotile cilia at the expense of motile cilia, and leads to an accumulation of immotile cilia at the anterior half of the KV. This disrupts the normal fluid flow intensity and pattern, with consequent impact on dand5 expression pattern and left-right (L-R) axis establishment.

  19. Notch/Her12 signalling modulates, motile/immotile cilia ratio downstream of Foxj1a in zebrafish left-right organizer

    PubMed Central

    Sampaio, Pedro; Pestana, Sara; Pinto, Andreia; Vaz, Andreia; Roxo-Rosa, Mónica; Gardner, Rui; Lopes, Telma; Schilling, Britta; Henry, Ian; Saúde, Leonor

    2017-01-01

    Foxj1a is necessary and sufficient to specify motile cilia. Using transcriptional studies and slow-scan two-photon live imaging capable of identifying the number of motile and immotile cilia, we now established that the final number of motile cilia depends on Notch signalling (NS). We found that despite all left-right organizer (LRO) cells express foxj1a and the ciliary axonemes of these cells have dynein arms, some cilia remain immotile. We identified that this decision is taken early in development in the Kupffer’s Vesicle (KV) precursors the readout being her12 transcription. We demonstrate that overexpression of either her12 or Notch intracellular domain (NICD) increases the number of immotile cilia at the expense of motile cilia, and leads to an accumulation of immotile cilia at the anterior half of the KV. This disrupts the normal fluid flow intensity and pattern, with consequent impact on dand5 expression pattern and left-right (L-R) axis establishment. PMID:28875937

  20. Cilia-mediated Hedgehog signaling controls form and function in the mammalian larynx.

    PubMed

    Tabler, Jacqueline M; Rigney, Maggie M; Berman, Gordon J; Gopalakrishnan, Swetha; Heude, Eglantine; Al-Lami, Hadeel Adel; Yannakoudiadkis, Basil Z; Fitch, Rebecca D; Carter, Christopher; Vokes, Steven; Liu, Karen J; Tajbakhsh, Shahragim; Egnor, Se Roian; Wallingford, John B

    2017-02-13

    Acoustic communication is fundamental to social interactions among animals, including humans. In fact, deficits in voice impair the quality of life for a large and diverse population of patients. Understanding the molecular genetic mechanisms of development and function in the vocal apparatus is thus an important challenge with relevance both to the basic biology of animal communication and to biomedicine. However, surprisingly little is known about the developmental biology of the mammalian larynx. Here, we used genetic fate mapping to chart the embryological origins of the tissues in the mouse larynx, and we describe the developmental etiology of laryngeal defects in mice with disruptions in cilia-mediated Hedgehog signaling. In addition, we show that mild laryngeal defects correlate with changes in the acoustic structure of vocalizations. Together, these data provide key new insights into the molecular genetics of form and function in the mammalian vocal apparatus.

  1. The role of Ca2+ in deflection-induced excitation of motile, mechanoresponsive balancer cilia in the ctenophore statocyst.

    PubMed

    Lowe, B

    1997-06-01

    Motile, mechanoresponsive cilia (balancers) in ctenophore statocysts, like vertebrate hair cells, are excited or inhibited depending upon the direction in which they are deflected. Balancers, however, may become either excited (beat rapidly) or inhibited (beat slowly) by deflection in the same direction, depending on the sign of ctenophore geotaxis (positive or negative). The beat frequency of many cilia is controlled by concentrations of Ca2+, membrane potential and neural input. How these factors affect deflection-induced ciliary beating in balancers was investigated. Deflection-induced excitation of balancers in whole Mnemiopsis leidyi larvae and dissected adult (Mnemiopsis leidyi, Pleurobrachia pileus) statocysts was reversibly inhibited by the Ca2+ channel inhibitors Co2+, Mg2+, Ni2+, and Mn2+. Deflection-induced excitation in balancers of isolated adult M. leidyi balancer groups was also inhibited by Co2+ or by Ca(2+)-free medium. Isolated balancer group cilia, like balancer cilia of intact ctenophores, exhibited responses to either sign of geotaxis and graded responses to deflection. Isolated balancers that were chemically depolarized in high-[K+], Ca(2+)-free medium were excited by local application of Ca2+ onto the ciliary bases, but not onto the cell bases or the ciliary tips. It is proposed that deflection-induced excitation of balancers is due to influx of Ca2+ through stretch- and voltage-activated channel activity. The sign of geotaxis of whole larvae and dissected adult statocysts was switched by electrical stimulation. Thus, neural input may participate in reversing the directional sensitivity of balancer cells.

  2. Cilia-mediated Hedgehog signaling controls form and function in the mammalian larynx

    PubMed Central

    Tabler, Jacqueline M; Rigney, Maggie M; Berman, Gordon J; Gopalakrishnan, Swetha; Heude, Eglantine; Al-lami, Hadeel Adel; Yannakoudiadkis, Basil Z; Fitch, Rebecca D; Carter, Christopher; Vokes, Steven; Liu, Karen J; Tajbakhsh, Shahragim; Egnor, SE Roian; Wallingford, John B

    2017-01-01

    Acoustic communication is fundamental to social interactions among animals, including humans. In fact, deficits in voice impair the quality of life for a large and diverse population of patients. Understanding the molecular genetic mechanisms of development and function in the vocal apparatus is thus an important challenge with relevance both to the basic biology of animal communication and to biomedicine. However, surprisingly little is known about the developmental biology of the mammalian larynx. Here, we used genetic fate mapping to chart the embryological origins of the tissues in the mouse larynx, and we describe the developmental etiology of laryngeal defects in mice with disruptions in cilia-mediated Hedgehog signaling. In addition, we show that mild laryngeal defects correlate with changes in the acoustic structure of vocalizations. Together, these data provide key new insights into the molecular genetics of form and function in the mammalian vocal apparatus. DOI: http://dx.doi.org/10.7554/eLife.19153.001 PMID:28177282

  3. Motile cilia create fluid-mechanical microhabitats for the active recruitment of the host microbiome.

    PubMed

    Nawroth, Janna C; Guo, Hanliang; Koch, Eric; Heath-Heckman, Elizabeth A C; Hermanson, John C; Ruby, Edward G; Dabiri, John O; Kanso, Eva; McFall-Ngai, Margaret

    2017-09-05

    We show that mucociliary membranes of animal epithelia can create fluid-mechanical microenvironments for the active recruitment of the specific microbiome of the host. In terrestrial vertebrates, these tissues are typically colonized by complex consortia and are inaccessible to observation. Such tissues can be directly examined in aquatic animals, providing valuable opportunities for the analysis of mucociliary activity in relation to bacteria recruitment. Using the squid-vibrio model system, we provide a characterization of the initial engagement of microbial symbionts along ciliated tissues. Specifically, we developed an empirical and theoretical framework to conduct a census of ciliated cell types, create structural maps, and resolve the spatiotemporal flow dynamics. Our multiscale analyses revealed two distinct, highly organized populations of cilia on the host tissues. An array of long cilia ([Formula: see text]25 [Formula: see text]m) with metachronal beat creates a flow that focuses bacteria-sized particles, at the exclusion of larger particles, into sheltered zones; there, a field of randomly beating short cilia ([Formula: see text]10 [Formula: see text]m) mixes the local fluid environment, which contains host biochemical signals known to prime symbionts for colonization. This cilia-mediated process represents a previously unrecognized mechanism for symbiont recruitment. Each mucociliary surface that recruits a microbiome such as the case described here is likely to have system-specific features. However, all mucociliary surfaces are subject to the same physical and biological constraints that are imposed by the fluid environment and the evolutionary conserved structure of cilia. As such, our study promises to provide insight into universal mechanisms that drive the recruitment of symbiotic partners.

  4. Studies on cilia. 3. Further studies on the cilium tip and a "sliding filament" model of ciliary motility.

    PubMed

    Satir, P

    1968-10-01

    This study confirms and extends previous work on the lateral cilia of the fresh-water mussel, Elliptio complanatus, in support of a "sliding filament" mechanism of ciliary motility wherein peripheral filaments (microtubules) do not change length during beat (see Satir, 1967). Short sequences of serial sections of tips are examined in control (nonbeating) and activated (metachronal wave) preparations. Several different tip types, functional rather than morphogenetic variants, are demonstrated, but similarly bent cilia have similar tips. The peripheral filaments are composed of two subfibers: a and b. The bent regions of cilia are in the form of circular arcs, and apparent differences in subfiber-b length at the tip are those predicted solely by geometry of the stroke without the necessity of assuming filament contraction. Various subfibers b apparently move with respect to one another during beat, since small systematic variations in relative position can be detected from cilium to cilium. While subfiber-b lengths are uniform throughout, subfiber-a lengths are morphologically different for each filament: 8 and 3 are about 0.8 micro longer than 1, 4 and 5, but each unique length is independent of stroke position or tip type. Subfiber-a does not contract, nor does it move, e.g. slide, with respect to subfiber-b of the same doublet. The central pair of filaments extends to the tip of the cilium where its members fuse. Subunit assembly in ciliary microtubules is evidently precise. This may be of importance in establishing the relationships needed for mechanochemical interactions that produce sliding and beat.

  5. CILIA: before and after.

    PubMed

    Satir, Peter

    2017-01-01

    This is a history of cilia research before and after the discovery of intraflagellar transport (IFT) and the link between primary cilia ciliogenesis and polycystic kidney disease (PKD). Before IFT, ca. the beginning of the new millennium, although sensory and primary cilia were well described, research was largely focused on motile cilia, their structure, movement, and biogenesis. After IFT and the link to PKD, although work on motile cilia has continued to progress, research on primary cilia has exploded, leading to new insights into the role of cilia in cell signaling and development. Genomics, proteomics, and new imaging techniques have unified the field and pointed out the critical role of cilia as a restricted cell organellar compartment, functionally integrated with other cell organelles including the autophagosome and the nucleus.

  6. Mutations in the Motile Cilia Gene DNAAF1 Are Associated with Neural Tube Defects in Humans.

    PubMed

    Miao, Chunyue; Jiang, Qian; Li, Huili; Zhang, Qin; Bai, Baoling; Bao, Yihua; Zhang, Ting

    2016-10-13

    Neural tube defects (NTDs) are severe malformations of the central nervous system caused by complex genetic and environmental factors. Among genes involved in NTD, cilia-related genes have been well defined and found to be essential for the completion of neural tube closure (NTC). We have carried out next-generation sequencing on target genes in 373 NTDs and 222 healthy controls, and discovered eight disease-specific rare mutations in cilia-related gene DNAAF1 DNAAF1 plays a central role in cytoplasmic preassembly of distinct dynein-arm complexes, and is expressed in some key tissues involved in neural system development, such as neural tube, floor plate, embryonic node, and brain ependyma epithelial cells in zebrafish and mouse. Therefore, we evaluated the expression and functions of mutations in DNAAF1 in transfected cells to analyze the potential correlation of these mutants to NTDs in humans. One rare frameshift mutation (p.Gln341Argfs*10) resulted in significantly diminished DNAAF1 protein expression, compared to the wild type. Another mutation, p.Lys231Gln, disrupted cytoplasmic preassembly of the dynein-arm complexes in cellular assay. Furthermore, results from NanoString assay on mRNA from NTD samples indicated that DNAAF1 mutants altered the expression level of NTC-related genes. Altogether, these findings suggest that the rare mutations in DNAAF1 may contribute to the susceptibility for NTDs in humans.

  7. Mutations in the Motile Cilia Gene DNAAF1 Are Associated with Neural Tube Defects in Humans

    PubMed Central

    Miao, Chunyue; Jiang, Qian; Li, Huili; Zhang, Qin; Bai, Baoling; Bao, Yihua; Zhang, Ting

    2016-01-01

    Neural tube defects (NTDs) are severe malformations of the central nervous system caused by complex genetic and environmental factors. Among genes involved in NTD, cilia-related genes have been well defined and found to be essential for the completion of neural tube closure (NTC). We have carried out next-generation sequencing on target genes in 373 NTDs and 222 healthy controls, and discovered eight disease-specific rare mutations in cilia-related gene DNAAF1. DNAAF1 plays a central role in cytoplasmic preassembly of distinct dynein-arm complexes, and is expressed in some key tissues involved in neural system development, such as neural tube, floor plate, embryonic node, and brain ependyma epithelial cells in zebrafish and mouse. Therefore, we evaluated the expression and functions of mutations in DNAAF1 in transfected cells to analyze the potential correlation of these mutants to NTDs in humans. One rare frameshift mutation (p.Gln341Argfs*10) resulted in significantly diminished DNAAF1 protein expression, compared to the wild type. Another mutation, p.Lys231Gln, disrupted cytoplasmic preassembly of the dynein-arm complexes in cellular assay. Furthermore, results from NanoString assay on mRNA from NTD samples indicated that DNAAF1 mutants altered the expression level of NTC-related genes. Altogether, these findings suggest that the rare mutations in DNAAF1 may contribute to the susceptibility for NTDs in humans. PMID:27543293

  8. Quantifying hyperoxia-mediated damage to mammalian respiratory cilia-driven fluid flow using particle tracking velocimetry optical coherence tomography.

    PubMed

    Gamm, Ute A; Huang, Brendan K; Syed, Mansoor; Zhang, Xuchen; Bhandari, Vineet; Choma, Michael A

    2015-08-01

    Oxygen supplementation [hyperoxia, increased fraction of inspired oxygen (FiO 2 )] is an indispensable treatment in the intensive care unit for patients in respiratory failure. Like other treatments or drugs, hyperoxia has a risk-benefit profile that guides its clinical use. While hyperoxia is known to damage respiratory epithelium, it is unknown if damage can result in impaired capacity to generate cilia-driven fluid flow. Here, we demonstrate that quantifying cilia-driven fluid flow velocities in the sub-100 μm/s regime (sub-0.25 in./min regime) reveals hyperoxia-mediated damage to the capacity of ciliated respiratory mucosa to generate directional flow. Flow quantification was performed using particle tracking velocimetry optical coherence tomography (PTV-OCT) in ex vivo mouse trachea. The ability of PTV-OCT to detect biomedically relevant flow perturbations in the sub-100 μm/s regime was validated by quantifying temperature- and drug-mediated modulation of flow performance in ex vivo mouse trachea. Overall, PTV-OCT imaging of cilia-driven fluid flow in ex vivo mouse trachea is a powerful and straightforward approach for studying factors that modulate and damage mammalian respiratory ciliary physiology.

  9. Cilia Dysfunction in Lung Disease

    PubMed Central

    Tilley, Ann E.; Walters, Matthew S.; Shaykhiev, Renat; Crystal, Ronald G.

    2015-01-01

    A characteristic feature of the human airway epithelium is the presence of ciliated cells bearing motile cilia, specialized cell surface projections containing axonemes comprised of microtubules and dynein arms, which provide ATP-driven motility. In the airways, cilia function in concert with airway mucus to mediate the critical function of mucociliary clearance, cleansing the airways of inhaled particles and pathogens. The prototypical disorder of respiratory cilia is primary ciliary dyskinesia, an inherited disorder that leads to impaired mucociliary clearance, repeated chest infections, and progressive destruction of lung architecture. Numerous acquired lung diseases are also marked by abnormalities in both cilia structure and function. In this review we summarize current knowledge regarding airway ciliated cells and cilia, how they function to maintain a healthy epithelium, and how disorders of cilia structure and function contribute to inherited and acquired lung disease. PMID:25386990

  10. Evaluating efficiency and robustness in cilia design

    NASA Astrophysics Data System (ADS)

    Guo, Hanliang; Kanso, Eva

    2016-03-01

    Motile cilia are used by many eukaryotic cells to transport flow. Cilia-driven flows are important to many physiological functions, yet a deep understanding of the interplay between the mechanical structure of cilia and their physiological functions in healthy and diseased conditions remains elusive. To develop such an understanding, one needs a quantitative framework to assess cilia performance and robustness when subject to perturbations in the cilia apparatus. Here we link cilia design (beating patterns) to function (flow transport) in the context of experimentally and theoretically derived cilia models. We particularly examine the optimality and robustness of cilia design. Optimality refers to efficiency of flow transport, while robustness is defined as low sensitivity to variations in the design parameters. We find that suboptimal designs can be more robust than optimal ones. That is, designing for the most efficient cilium does not guarantee robustness. These findings have significant implications on the understanding of cilia design in artificial and biological systems.

  11. PTEN regulates cilia through Dishevelled

    PubMed Central

    Shnitsar, Iryna; Bashkurov, Mikhail; Masson, Glenn R.; Ogunjimi, Abiodun A.; Mosessian, Sherly; Cabeza, Eduardo Aguiar; Hirsch, Calley L.; Trcka, Daniel; Gish, Gerald; Jiao, Jing; Wu, Hong; Winklbauer, Rudolf; Williams, Roger L.; Pelletier, Laurence; Wrana, Jeffrey L.; Barrios-Rodiles, Miriam

    2015-01-01

    Cilia are hair-like cellular protrusions important in many aspects of eukaryotic biology. For instance, motile cilia enable fluid movement over epithelial surfaces, while primary (sensory) cilia play roles in cellular signalling. The molecular events underlying cilia dynamics, and particularly their disassembly, are not well understood. Phosphatase and tensin homologue (PTEN) is an extensively studied tumour suppressor, thought to primarily act by antagonizing PI3-kinase signalling. Here we demonstrate that PTEN plays an important role in multicilia formation and cilia disassembly by controlling the phosphorylation of Dishevelled (DVL), another ciliogenesis regulator. DVL is a central component of WNT signalling that plays a role during convergent extension movements, which we show here are also regulated by PTEN. Our studies identify a novel protein substrate for PTEN that couples PTEN to regulation of cilia dynamics and WNT signalling, thus advancing our understanding of potential underlying molecular etiologies of PTEN-related pathologies. PMID:26399523

  12. PACRG, a protein linked to ciliary motility, mediates cellular signaling.

    PubMed

    Loucks, Catrina M; Bialas, Nathan J; Dekkers, Martijn P J; Walker, Denise S; Grundy, Laura J; Li, Chunmei; Inglis, P Nick; Kida, Katarzyna; Schafer, William R; Blacque, Oliver E; Jansen, Gert; Leroux, Michel R

    2016-07-01

    Cilia are microtubule-based organelles that project from nearly all mammalian cell types. Motile cilia generate fluid flow, whereas nonmotile (primary) cilia are required for sensory physiology and modulate various signal transduction pathways. Here we investigate the nonmotile ciliary signaling roles of parkin coregulated gene (PACRG), a protein linked to ciliary motility. PACRG is associated with the protofilament ribbon, a structure believed to dictate the regular arrangement of motility-associated ciliary components. Roles for protofilament ribbon-associated proteins in nonmotile cilia and cellular signaling have not been investigated. We show that PACRG localizes to a small subset of nonmotile cilia in Caenorhabditis elegans, suggesting an evolutionary adaptation for mediating specific sensory/signaling functions. We find that it influences a learning behavior known as gustatory plasticity, in which it is functionally coupled to heterotrimeric G-protein signaling. We also demonstrate that PACRG promotes longevity in C. elegans by acting upstream of the lifespan-promoting FOXO transcription factor DAF-16 and likely upstream of insulin/IGF signaling. Our findings establish previously unrecognized sensory/signaling functions for PACRG and point to a role for this protein in promoting longevity. Furthermore, our work suggests additional ciliary motility-signaling connections, since EFHC1 (EF-hand containing 1), a potential PACRG interaction partner similarly associated with the protofilament ribbon and ciliary motility, also positively regulates lifespan.

  13. PACRG, a protein linked to ciliary motility, mediates cellular signaling

    PubMed Central

    Loucks, Catrina M.; Bialas, Nathan J.; Dekkers, Martijn P. J.; Walker, Denise S.; Grundy, Laura J.; Li, Chunmei; Inglis, P. Nick; Kida, Katarzyna; Schafer, William R.; Blacque, Oliver E.; Jansen, Gert; Leroux, Michel R.

    2016-01-01

    Cilia are microtubule-based organelles that project from nearly all mammalian cell types. Motile cilia generate fluid flow, whereas nonmotile (primary) cilia are required for sensory physiology and modulate various signal transduction pathways. Here we investigate the nonmotile ciliary signaling roles of parkin coregulated gene (PACRG), a protein linked to ciliary motility. PACRG is associated with the protofilament ribbon, a structure believed to dictate the regular arrangement of motility-associated ciliary components. Roles for protofilament ribbon–associated proteins in nonmotile cilia and cellular signaling have not been investigated. We show that PACRG localizes to a small subset of nonmotile cilia in Caenorhabditis elegans, suggesting an evolutionary adaptation for mediating specific sensory/signaling functions. We find that it influences a learning behavior known as gustatory plasticity, in which it is functionally coupled to heterotrimeric G-protein signaling. We also demonstrate that PACRG promotes longevity in C. elegans by acting upstream of the lifespan-promoting FOXO transcription factor DAF-16 and likely upstream of insulin/IGF signaling. Our findings establish previously unrecognized sensory/signaling functions for PACRG and point to a role for this protein in promoting longevity. Furthermore, our work suggests additional ciliary motility-signaling connections, since EFHC1 (EF-hand containing 1), a potential PACRG interaction partner similarly associated with the protofilament ribbon and ciliary motility, also positively regulates lifespan. PMID:27193298

  14. Intraflagellar Transport Protein 172 is essential for primary cilia formation and plays a vital role in patterning the mammalian brain

    PubMed Central

    Gorivodsky, Marat; Mukhopadhyay, Mahua; Wilsch-Braeuninger, Michaela; Phillips, Matthew; Teufel, Andreas; Kim, Changmee; Malik, Nasir; Huttner, Wieland; Westphal, Heiner

    2008-01-01

    IFT172, also known as Selective Lim-domain Binding protein (SLB), is a component of the Intraflagellar Transport (IFT) complex. In order to evaluate the biological role of the Ift172 gene, we generated a loss-of-function mutation in the mouse. The resulting Slb mutant embryos die between E12.5–13.0, and exhibit severe cranio-facial malformations, failure to close the cranial neural tube, holoprosencephaly, heart edema and extensive hemorrhages. Cilia outgrowth in cells of the neuroepithelium is initiated but the axonemes are severely truncated and do not contain visible microtubules. Morphological and molecular analyses revealed a global brain-patterning defect along the dorsal-ventral (DV) and anterior-posterior (AP) axes. We demonstrate that Ift172 gene function is required for early regulation of Fgf8 at the midbrain-hindbrain boundary and maintenance of the isthmic organizer. In addition, Ift172 is required for proper function of the embryonic node, the early embryonic organizer and for formation of the head organizing center (the anterior mesendoderm, or AME). We propose a model suggesting that forebrain and mid-hindbrain growth and AP patterning depends on the early function of Ift172 at gastrulation. Our data suggest that the formation and function of the node and AME in the mouse embryo relies on an indispensable role of Ift172 in cilia morphogenesis and cilia-mediated signaling. PMID:18930042

  15. Fine-Tuning Motile Cilia and Flagella: Evolution of the Dynein Motor Proteins from Plants to Humans at High Resolution

    PubMed Central

    Kollmar, Martin

    2016-01-01

    The flagellum is a key innovation linked to eukaryogenesis. It provides motility by regulated cycles of bending and bend propagation, which are thought to be controlled by a complex arrangement of seven distinct dyneins in repeated patterns of outer- (OAD) and inner-arm dynein (IAD) complexes. Electron tomography showed high similarity of this axonemal repeat pattern across ciliates, algae, and animals, but the diversity of dynein sequences across the eukaryotes has not yet comprehensively been resolved and correlated with structural data. To shed light on the evolution of the axoneme I performed an exhaustive analysis of dyneins using the available sequenced genome data. Evidence from motor domain phylogeny allowed expanding the current set of nine dynein subtypes by eight additional isoforms with, however, restricted taxonomic distributions. I confirmed the presence of the nine dyneins in all eukaryotic super-groups indicating their origin predating the last eukaryotic common ancestor. The comparison of the N-terminal tail domains revealed a most likely axonemal dynein origin of the new classes, a group of chimeric dyneins in plants/algae and Stramenopiles, and the unique domain architecture and origin of the outermost OADs present in green algae and ciliates but not animals. The correlation of sequence and structural data suggests the single-headed class-8 and class-9 dyneins to localize to the distal end of the axonemal repeat and the class-7 dyneins filling the region up to the proximal heterodimeric IAD. Tracing dynein gene duplications across the eukaryotes indicated ongoing diversification and fine-tuning of flagellar functions in extant taxa and species. PMID:27880711

  16. CFAP54 is required for proper ciliary motility and assembly of the central pair apparatus in mice

    PubMed Central

    McKenzie, Casey W.; Craige, Branch; Kroeger, Tiffany V.; Finn, Rozzy; Wyatt, Todd A.; Sisson, Joseph H.; Pavlik, Jacqueline A.; Strittmatter, Lara; Hendricks, Gregory M.; Witman, George B.; Lee, Lance

    2015-01-01

    Motile cilia and flagella play critical roles in fluid clearance and cell motility, and dysfunction commonly results in the pediatric syndrome primary ciliary dyskinesia (PCD). CFAP221, also known as PCDP1, is required for ciliary and flagellar function in mice and Chlamydomonas reinhardtii, where it localizes to the C1d projection of the central microtubule apparatus and functions in a complex that regulates flagellar motility in a calcium-dependent manner. We demonstrate that the genes encoding the mouse homologues of the other C. reinhardtii C1d complex members are primarily expressed in motile ciliated tissues, suggesting a conserved function in mammalian motile cilia. The requirement for one of these C1d complex members, CFAP54, was identified in a mouse line with a gene-trapped allele. Homozygous mice have PCD characterized by hydrocephalus, male infertility, and mucus accumulation. The infertility results from defects in spermatogenesis. Motile cilia have a structural defect in the C1d projection, indicating that the C1d assembly mechanism requires CFAP54. This structural defect results in decreased ciliary beat frequency and perturbed cilia-driven flow. This study identifies a critical role for CFAP54 in proper assembly and function of mammalian cilia and flagella and establishes the gene-trapped allele as a new model of PCD. PMID:26224312

  17. Species-Specific Adaptations of Trypanosome Morphology and Motility to the Mammalian Host

    PubMed Central

    McOdimba, Francis A.; Omogo, Collins O.; Adung’a, Vincent O.; Krüger, Timothy; Masiga, Daniel K.; Engstler, Markus

    2016-01-01

    African trypanosomes thrive in the bloodstream and tissue spaces of a wide range of mammalian hosts. Infections of cattle cause an enormous socio-economic burden in sub-Saharan Africa. A hallmark of the trypanosome lifestyle is the flagellate’s incessant motion. This work details the cell motility behavior of the four livestock-parasites Trypanosoma vivax, T. brucei, T. evansi and T. congolense. The trypanosomes feature distinct swimming patterns, speeds and flagellar wave frequencies, although the basic mechanism of flagellar propulsion is conserved, as is shown by extended single flagellar beat analyses. Three-dimensional analyses of the trypanosomes expose a high degree of dynamic pleomorphism, typified by the ‘cellular waveform’. This is a product of the flagellar oscillation, the chirality of the flagellum attachment and the stiffness of the trypanosome cell body. The waveforms are characteristic for each trypanosome species and are influenced by changes of the microenvironment, such as differences in viscosity and the presence of confining obstacles. The distinct cellular waveforms may be reflective of the actual anatomical niches the parasites populate within their mammalian host. T. vivax displays waveforms optimally aligned to the topology of the bloodstream, while the two subspecies T. brucei and T. evansi feature distinct cellular waveforms, both additionally adapted to motion in more confined environments such as tissue spaces. T. congolense reveals a small and stiff waveform, which makes these parasites weak swimmers and destined for cell adherence in low flow areas of the circulation. Thus, our experiments show that the differential dissemination and annidation of trypanosomes in their mammalian hosts may depend on the distinct swimming capabilities of the parasites. PMID:26871910

  18. Primary cilia and graded Sonic Hedgehog signaling.

    PubMed

    Sasai, Noriaki; Briscoe, James

    2012-01-01

    Cilia are evolutionary-conserved microtubule-containing organelles protruding from the surface of cells. They are classified into two types--primary and motile cilia. Primary cilia are nearly ubiquitous, at least in vertebrate cells, and it has become apparent that they play an essential role in the intracellular transduction of a range of stimuli. Most notable among these is Sonic Hedgehog. In this article we briefly summarize the structure and biogenesis of primary cilia. We discuss the evidence implicating cilia in the transduction of extrinsic signals. We focus on the involvement and molecular mechanism of cilia in signaling by Sonic Hedgehog in embryonic tissues, specifically the neural tube, and we discuss how cilia play an active role in the interpretation of gradients of Sonic Hedgehog (Shh) signaling.

  19. Pericentrin, a centrosomal protein related to microcephalic primordial dwarfism, is required for olfactory cilia assembly in mice.

    PubMed

    Miyoshi, Ko; Kasahara, Kyosuke; Miyazaki, Ikuko; Shimizu, Shoko; Taniguchi, Manabu; Matsuzaki, Shinsuke; Tohyama, Masaya; Asanuma, Masato

    2009-10-01

    The Drosophila pericentrin-like protein has been shown to be essential for the formation of the sensory cilia of chemosensory and mechanosensory neurons by mutant analysis in flies, while the in vivo function of pericentrin, a well-studied mammalian centrosomal protein related to microcephalic primordial dwarfism, has been unclear. To determine whether pericentrin is required for ciliogenesis in mammals, we generated and analyzed mice with a hypomorphic mutation of Pcnt encoding the mouse pericentrin. Immunofluorescence analysis demonstrated that olfactory cilia of chemosensory neurons in the nasal olfactory epithelium were malformed in the homozygous mutant mice. On the other hand, the assembly of motile and primary cilia of non-neuronal epithelial cells and the formation of sperm flagella were not affected in the Pcnt-mutant mice. The defective assembly of olfactory cilia in the mutant was apparent from birth. The mutant animals displayed reduced olfactory performance in agreement with the compromised assembly of olfactory cilia. Our findings suggest that pericentrin is essential for the assembly of chemosensory cilia of olfactory receptor neurons, but it is not globally required for cilia formation in mammals.

  20. Cilia distribution and polarity in the epithelial lining of the mouse middle ear cavity

    PubMed Central

    Luo, Wenwei; Yi, Hong; Taylor, Jeannette; Li, Jian-dong; Chi, Fanglu; Todd, N. Wendell; Lin, Xi; Ren, Dongdong; Chen, Ping

    2017-01-01

    The middle ear conducts sound to the cochlea for hearing. Otitis media (OM) is the most common illness in childhood. Moreover, chronic OM with effusion (COME) is the leading cause of conductive hearing loss. Clinically, COME is highly associated with Primary Ciliary Dyskinesia, implicating significant contributions of cilia dysfunction to COME. The understanding of middle ear cilia properties that are critical to OM susceptibility, however, is limited. Here, we confirmed the presence of a ciliated region near the Eustachian tube orifice at the ventral region of the middle ear cavity, consisting mostly of a lumen layer of multi-ciliated and a layer of Keratin-5-positive basal cells. We also found that the motile cilia are polarized coordinately and display a planar cell polarity. Surprisingly, we also found a region of multi-ciliated cells that line the posterior dorsal pole of the middle ear cavity which was previously thought to contain only non-ciliated cells. Our study provided a more complete understanding of cilia distribution and revealed for the first time coordinated polarity of cilia in the epithelium of the mammalian middle ear, thus illustrating novel structural features that are likely critical for middle ear functions and related to OM susceptibility. PMID:28358397

  1. Selective particle capture by asynchronously beating cilia

    NASA Astrophysics Data System (ADS)

    Ding, Yang; Kanso, Eva

    2015-12-01

    Selective particle filtration is fundamental in many engineering and biological systems. For example, many aquatic microorganisms use filter feeding to capture food particles from the surrounding fluid, using motile cilia. One of the capture strategies is to use the same cilia to generate feeding currents and to intercept particles when the particles are on the downstream side of the cilia. Here, we develop a 3D computational model of ciliary bands interacting with flow suspended particles and calculate particle trajectories for a range of particle sizes. Consistent with experimental observations, we find optimal particle sizes that maximize capture rate. The optimal size depends nonlinearly on cilia spacing and cilia coordination, synchronous vs. asynchronous. These parameters affect the cilia-generated flow field, which in turn affects particle trajectories. The low capture rate of smaller particles is due to the particles' inability to cross the flow streamlines of neighboring cilia. Meanwhile, large particles have difficulty entering the sub-ciliary region once advected downstream, also resulting in low capture rates. The optimal range of particle sizes is enhanced when cilia beat asynchronously. These findings have potentially important implications on the design and use of biomimetic cilia in processes such as particle sorting in microfluidic devices.

  2. Cilia organize ependymal planar polarity.

    PubMed

    Mirzadeh, Zaman; Han, Young-Goo; Soriano-Navarro, Mario; García-Verdugo, Jose Manuel; Alvarez-Buylla, Arturo

    2010-02-17

    Multiciliated epithelial cells, called ependymal cells, line the ventricles in the adult brain. Most ependymal cells are born prenatally and are derived from radial glia. Ependymal cells have a remarkable planar polarization that determines orientation of ciliary beating and propulsion of CSF. Disruption of ependymal ciliary beating, by injury or disease, results in aberrant CSF circulation and hydrocephalus, a common disorder of the CNS. Very little is known about the mechanisms guiding ependymal planar polarity and whether this organization is acquired during ependymal cell development or is already present in radial glia. Here we show that basal bodies in ependymal cells in the lateral ventricle walls of adult mice are polarized in two ways: (1) rotational; angle of individual basal bodies with respect to their long axis and (2) translational; the position of basal bodies on the apical surface of the cell. Conditional ablation of motile cilia disrupted rotational orientation, but translational polarity was largely preserved. In contrast, translational polarity was dramatically affected when radial glial primary cilia were ablated earlier in development. Remarkably, radial glia in the embryo have a translational polarity that predicts the orientation of mature ependymal cells. These results suggest that ependymal planar cell polarity is a multistep process initially organized by primary cilia in radial glia and then refined by motile cilia in ependymal cells.

  3. Cilia organize ependymal planar polarity

    PubMed Central

    Mirzadeh, Zaman; Han, Young-Goo; Soriano-Navarro, Mario; García-Verdugo, Jose Manuel; Alvarez-Buylla, Arturo

    2010-01-01

    Multi-ciliated epithelial cells, called ependymal cells, line the ventricles in the adult brain. Most ependymal cells are born prenatally and are derived from radial glia. Ependymal cells have a remarkable planar polarization that determines orientation of ciliary beating and propulsion of cerebrospinal fluid (CSF). Disruption of ependymal ciliary beating, by injury or disease, results in aberrant CSF circulation and hydrocephalus, a common disorder of the central nervous system. Very little is known about the mechanisms guiding ependymal planar polarity and whether this organization is acquired during ependymal cell development or is already present in radial glia. Here we show that basal bodies in ependymal cells in the lateral ventricle walls of adult mice are polarized in two ways: i) rotational; angle of individual basal bodies with respect to their long axis and ii) translational; the position of basal bodies on the apical surface of the cell. Conditional ablation of motile cilia disrupted rotational orientation, but translational polarity was largely preserved. In contrast, translational polarity was dramatically affected when radial glial primary cilia were ablated earlier in development. Remarkably, radial glia in the embryo have a translational polarity that predicts the orientation of mature ependymal cells. These results suggest that ependymal planar cell polarity is a multi-step process initially organized by primary cilia in radial glia and then refined by motile cilia in ependymal cells. PMID:20164345

  4. Mechanical Properties of Primary Cilia

    NASA Astrophysics Data System (ADS)

    Battle, Christopher; Schmidt, Christoph F.

    2013-03-01

    Recent studies have shown that the primary cilium, long thought to be a vestigial cellular appendage with no function, is involved in a multitude of sensory functions. One example, interesting from both a biophysical and medical standpoint, is the primary cilium of kidney epithelial cells, which acts as a mechanosensitive flow sensor. Genetic defects in ciliary function can cause, e.g., polycystic kidney disease (PKD). The material properties of these non-motile, microtubule-based 9 +0 cilia, and the way they are anchored to the cell cytoskeleton, are important to know if one wants to understand the mechano-electrochemical response of these cells, which is mediated by their cilia. We have probed the mechanical properties, boundary conditions, and dynamics of the cilia of MDCK cells using optical traps and DIC/fluorescence microscopy. We found evidence for both elastic relaxation of the cilia themselves after bending and for compliance in the intracellular anchoring structures. Angular and positional fluctuations of the cilia reflect both thermal excitations and cellular driving forces.

  5. The sensory cilia of Caenorhabditis elegans.

    PubMed

    Inglis, Peter N; Ou, Guangshuo; Leroux, Michel R; Scholey, Jonathan M

    2007-03-08

    The non-motile cilium, once believed to be a vestigial cellular structure, is now increasingly associated with the ability of a wide variety of cells and organisms to sense their chemical and physical environments. With its limited number of sensory cilia and diverse behavioral repertoire, C. elegans has emerged as a powerful experimental system for studying how cilia are formed, function, and ultimately modulate complex behaviors. Here, we discuss the biogenesis, distribution, structures, composition and general functions of C. elegans cilia. We also briefly highlight how C. elegans is being used to provide molecular insights into various human ciliopathies, including Polycystic Kidney Disease and Bardet-Biedl Syndrome.

  6. Proteomics of Primary Cilia by Proximity Labeling

    PubMed Central

    Mick, David U.; Rodrigues, Rachel B.; Leib, Ryan D.; Adams, Christopher M.; Chien, Allis S.; Gygi, Steven P.; Nachury, Maxence V.

    2015-01-01

    SUMMARY While cilia are recognized as important signaling organelles, the extent of ciliary functions remains unknown because of difficulties in cataloguing proteins from mammalian primary cilia. We present a method that readily captures rapid snapshots of the ciliary proteome by selectively biotinylating ciliary proteins using a cilia-targeted proximity labeling enzyme (cilia-APEX). Besides identifying known ciliary proteins, cilia-APEX uncovered several ciliary signaling molecules. The kinases PKA, AMPK and LKB1 were validated as bona fide ciliary proteins and PKA was found to regulate Hedgehog signaling in primary cilia. Furthermore, proteomics profiling of Ift27/Bbs19 mutant cilia correctly detected BBSome accumulation inside Ift27−/− cilia and revealed that β-arrestin 2 and the viral receptor CAR are candidate cargoes of the BBSome. This work demonstrates that proximity labeling can be applied to proteomics of non-membrane-enclosed organelles and suggests that proteomics profiling of cilia will enable a rapid and powerful characterization of ciliopathies. PMID:26585297

  7. Primary Cilia and Intraflagellar Transport Proteins in Bone and Cartilage.

    PubMed

    Yuan, X; Yang, S

    2016-11-01

    Primary cilia, present on most mammalian cells, function as a sensor to sense the environment change and transduce signaling. Loss of primary cilia causes a group of human pleiotropic syndromes called Ciliopathies. Some of the ciliopathies display skeletal dysplasias, implying the important role of primary cilia in skeletal development and homeostasis. Emerging evidence has shown that loss or malfunction of primary cilia or ciliary proteins in bone and cartilage is associated with developmental and function defects. Intraflagellar transport (IFT) proteins are essential for cilia formation and/or function. In this review, we discuss the role of primary cilia and IFT proteins in the development of bone and cartilage, as well as the differentiation and mechanotransduction of mesenchymal stem cells, osteoblasts, osteocytes, and chondrocytes. We also include the role of primary cilia in tooth development and highlight the current advance of primary cilia and IFT proteins in the pathogenesis of cartilage diseases, including osteoarthritis, osteosarcoma, and chondrosarcoma.

  8. JAM-A is present in mammalian spermatozoa where it is essential for normal motility.

    PubMed

    Shao, Minghai; Ghosh, Ananya; Cooke, Vesselina G; Naik, Ulhas P; Martin-DeLeon, Patricia A

    2008-01-01

    Junctional adhesion molecules (JAMs) that are expressed in endothelial and epithelial cells and function in tight junction assembly, also perform important roles in testis where the closely-related JAM-A, JAM-B, and JAM-C are found. Disruption of murine Jam-B and Jam-C has varying effects on sperm development and function; however, deletion of Jam-A has not yet been studied. Here we show for the first time that in addition to expression in the Sertoli-Sertoli tight junctions in the seminiferous tubules, the approximately 32 kDa murine JAM-A is present in elongated spermatids and in the plasma membrane of the head and flagellum of sperm. Deletion of Jam-A, using the gene trap technology, results in flagellar defects at the ultrastructural level. In Jam-A-deficient mice, which have reduced litter size, both progressive and hyperactive motility are significantly affected (P<0.0001) before and, more severely, after capacitation. The findings show that JAM-A is involved in sperm tail formation and is essential for normal motility, which may occur via its signal transduction and protein phosphorylation properties. Detection of JAM-A in human sperm proteins indicates that its role may be conserved in sperm motility and that JAM-A may be a candidate gene for the analysis of idiopathic sperm motility defects resulting in male subfertility in the human population.

  9. STUDIES ON CILIA

    PubMed Central

    Satir, Peter

    1968-01-01

    This study confirms and extends previous work on the lateral cilia of the fresh-water mussel, Elliptio complanatus, in support of a "sliding filament" mechanism of ciliary motility wherein peripheral filaments (microtubules) do not change length during beat (see Satir, 1967). Short sequences of serial sections of tips are examined in control (nonbeating) and activated (metachronal wave) preparations. Several different tip types, functional rather than morphogenetic variants, are demonstrated, but similarly bent cilia have similar tips. The peripheral filaments are composed of two subfibers: a and b. The bent regions of cilia are in the form of circular arcs, and apparent differences in subfiber-b length at the tip are those predicted solely by geometry of the stroke without the necessity of assuming filament contraction. Various subfibers b apparently move with respect to one another during beat, since small systematic variations in relative position can be detected from cilium to cilium. While subfiber-b lengths are uniform throughout, subfiber-a lengths are morphologically different for each filament: 8 and 3 are about 0.8 µ longer than 1, 4 and 5, but each unique length is independent of stroke position or tip type. Subfiber-a does not contract, nor does it move, e.g. slide, with respect to subfiber-b of the same doublet. The central pair of filaments extends to the tip of the cilium where its members fuse. Subunit assembly in ciliary microtubules is evidently precise. This may be of importance in establishing the relationships needed for mechanochemical interactions that produce sliding and beat. PMID:5678451

  10. Otoacoustic emissions without somatic motility: Can stereocilia mechanics drive the mammalian cochlea?

    NASA Astrophysics Data System (ADS)

    Liberman, M. C.; Zuo, Jian; Guinan, J. J.

    2004-09-01

    Distortion product otoacoustic emissions (DPOAEs) evoked by low-level tones are a sensitive indicator of outer hair cell (OHC) function. High-level DPOAEs are less vulnerable to cochlear insult, and their dependence on the OHC function is more controversial. Here, the mechanism underlying high-level DPOAE generation is addressed using a mutant mouse line lacking prestin, the molecular motor driving OHC somatic motility, required for cochlear amplification. With prestin deletion, attenuated DPOAEs were measurable at high sound levels. DPOAE thresholds were shifted by ~50 dB, matching the loss of cochlear amplifier gain measured in compound action potentials. In contrast, at high sound levels, distortion products in the cochlear microphonic (CM) of mutants were not decreased re wildtypes (expressed re CM at the primaries). Distortion products in both CM and otoacoustic emissions disappeared rapidly after death. The results show that OHC somatic motility is not necessary for the production of DPOAEs at high SPLs. They also suggest that the small, physiologically vulnerable DPOAE that remains without prestin-based motility is due directly to the mechanical nonlinearity associated with stereociliary transduction, and that this stereocilia mechanical nonlinearity is robustly coupled to the motion of the cochlear partition to the extent that it can drive the middle ear.

  11. Role of Primary Cilia in Odontogenesis.

    PubMed

    Hampl, M; Cela, P; Szabo-Rogers, H L; Bosakova, M Kunova; Dosedelova, H; Krejci, P; Buchtova, M

    2017-08-01

    Primary cilium is a solitary organelle that emanates from the surface of most postmitotic mammalian cells and serves as a sensory organelle, transmitting the mechanical and chemical cues to the cell. Primary cilia are key coordinators of various signaling pathways during development and maintenance of tissue homeostasis. The emerging evidence implicates primary cilia function in tooth development. Primary cilia are located in the dental epithelium and mesenchyme at early stages of tooth development and later during cell differentiation and production of hard tissues. The cilia are present when interactions between both the epithelium and mesenchyme are required for normal morphogenesis. As the primary cilium coordinates several signaling pathways essential for odontogenesis, ciliary defects can interrupt the latter process. Genetic or experimental alterations of cilia function lead to various developmental defects, including supernumerary or missing teeth, enamel and dentin hypoplasia, or teeth crowding. Moreover, dental phenotypes are observed in ciliopathies, including Bardet-Biedl syndrome, Ellis-van Creveld syndrome, Weyers acrofacial dysostosis, cranioectodermal dysplasia, and oral-facial-digital syndrome, altogether demonstrating that primary cilia play a critical role in regulation of both the early odontogenesis and later differentiation of hard tissue-producing cells. Here, we summarize the current evidence for the localization of primary cilia in dental tissues and the impact of disrupted cilia signaling on tooth development in ciliopathies.

  12. Functional optical coherence tomography for high-resolution mapping of cilia beat frequency in the mouse oviduct in vivo

    NASA Astrophysics Data System (ADS)

    Wang, Shang; Burton, Jason C.; Behringer, Richard R.; Larina, Irina V.

    2016-02-01

    Since mouse is a superior model for genetic analysis of human disorders, reproductive studies in mice have significant implications on further understanding of fertility and infertility in humans. Fertilized oocytes are transported through the reproductive tract by motile cilia lining the lumen of the oviduct as well as by oviduct contractions. While the role of cilia is well recognized, ciliary dynamics in the oviduct is not well understood, largely owing to the lack of live imaging approaches. Here, we report in vivo micro-scale mapping of cilia and cilia beat frequency (CBF) in the mouse oviduct using optical coherence tomography (OCT). This functional imaging method is based on spectral analysis of the OCT speckle variations produced by the beat of cilia in the oviduct, which does not require exogenous contrast agents. Animal procedures similar to the ones used for production of transgenic mice are utilized to expose the reproductive organs for imaging in anesthetized females. In this paper, we first present in vivo structural imaging of the mouse oviduct capturing the oocyte and the preimplantation embryo and then show the result of depth-resolved high-resolution CBF mapping in the ampulla of the live mouse. These data indicate that this structural and functional OCT imaging approach can be a useful tool for a variety of live investigations of mammalian reproduction and infertility.

  13. Cilia induced cerebrospinal fluid flow in the third ventricle of brain

    NASA Astrophysics Data System (ADS)

    Wang, Yong; Westendorf, Christian; Faubel, Regina; Eichele, Gregor; Bodenschatz, Eberhard

    2016-11-01

    Cerebrospinal fluid (CSF) conveys many physiologically important signaling factors through the ventricles of the mammalian brain. The walls of the ventricles are covered with motile cilia that were thought to generate a laminar flow purely following the curvature of walls. However, we recently discovered that cilia of the ventral third ventricle (v3V) generate a complex flow network along the wall, leading to subdivision of the v3V. The contribution of such cilia induced flow to the overall three dimensional volume flow remains to be investigated by using numerical simulation, arguably the best approach for such investigations. The lattice Boltzmann method is used to study the CFS flow in a reconstructed geometry of the v3V. Simulation of CSF flow neglecting cilia in this geometry confirmed that the previous idea about pure confined flow does not reflect the reality observed in experiment. The experimentally recorded ciliary flow network along the wall was refined with the smoothed particle hydrodynamics and then adapted as boundary condition in simulation. We study the contribution of the ciliary network to overall CSF flow and identify site-specific delivery of CSF constituents with respect to the temporal changes.

  14. Switching on cilia: transcriptional networks regulating ciliogenesis.

    PubMed

    Choksi, Semil P; Lauter, Gilbert; Swoboda, Peter; Roy, Sudipto

    2014-04-01

    Cilia play many essential roles in fluid transport and cellular locomotion, and as sensory hubs for a variety of signal transduction pathways. Despite having a conserved basic morphology, cilia vary extensively in their shapes and sizes, ultrastructural details, numbers per cell, motility patterns and sensory capabilities. Emerging evidence indicates that this diversity, which is intimately linked to the different functions that cilia perform, is in large part programmed at the transcriptional level. Here, we review our understanding of the transcriptional control of ciliary biogenesis, highlighting the activities of FOXJ1 and the RFX family of transcriptional regulators. In addition, we examine how a number of signaling pathways, and lineage and cell fate determinants can induce and modulate ciliogenic programs to bring about the differentiation of distinct cilia types.

  15. Cilia gene mutations cause atrioventricular septal defects by multiple mechanisms

    PubMed Central

    Burnicka-Turek, Ozanna; Steimle, Jeffrey D.; Huang, Wenhui; Felker, Lindsay; Kamp, Anna; Kweon, Junghun; Peterson, Michael; Reeves, Roger H.; Maslen, Cheryl L.; Gruber, Peter J.; Yang, Xinan H.; Shendure, Jay; Moskowitz, Ivan P.

    2016-01-01

    Atrioventricular septal defects (AVSDs) are a common severe form of congenital heart disease (CHD). In this study we identified deleterious non-synonymous mutations in two cilia genes, Dnah11 and Mks1, in independent N-ethyl-N-nitrosourea-induced mouse mutant lines with heritable recessive AVSDs by whole-exome sequencing. Cilia are required for left/right body axis determination and second heart field (SHF) Hedgehog (Hh) signaling, and we find that cilia mutations affect these requirements differentially. Dnah11avc4 did not disrupt SHF Hh signaling and caused AVSDs only concurrently with heterotaxy, a left/right axis abnormality. In contrast, Mks1avc6 disrupted SHF Hh signaling and caused AVSDs without heterotaxy. We performed unbiased whole-genome SHF transcriptional profiling and found that cilia motility genes were not expressed in the SHF whereas cilia structural and signaling genes were highly expressed. SHF cilia gene expression predicted the phenotypic concordance between AVSDs and heterotaxy in mice and humans with cilia gene mutations. A two-step model of cilia action accurately predicted the AVSD/heterotaxyu phenotypic expression pattern caused by cilia gene mutations. We speculate that cilia gene mutations contribute to both syndromic and non-syndromic AVSDs in humans and provide a model that predicts the phenotypic consequences of specific cilia gene mutations. PMID:27340223

  16. Brucella spp. of amphibians comprise genomically diverse motile strains competent for replication in macrophages and survival in mammalian hosts

    PubMed Central

    Al Dahouk, Sascha; Köhler, Stephan; Occhialini, Alessandra; Jiménez de Bagüés, María Pilar; Hammerl, Jens Andre; Eisenberg, Tobias; Vergnaud, Gilles; Cloeckaert, Axel; Zygmunt, Michel S.; Whatmore, Adrian M.; Melzer, Falk; Drees, Kevin P.; Foster, Jeffrey T.; Wattam, Alice R.; Scholz, Holger C.

    2017-01-01

    Twenty-one small Gram-negative motile coccobacilli were isolated from 15 systemically diseased African bullfrogs (Pyxicephalus edulis), and were initially identified as Ochrobactrum anthropi by standard microbiological identification systems. Phylogenetic reconstructions using combined molecular analyses and comparative whole genome analysis of the most diverse of the bullfrog strains verified affiliation with the genus Brucella and placed the isolates in a cluster containing B. inopinata and the other non-classical Brucella species but also revealed significant genetic differences within the group. Four representative but molecularly and phenotypically diverse strains were used for in vitro and in vivo infection experiments. All readily multiplied in macrophage-like murine J774-cells, and their overall intramacrophagic growth rate was comparable to that of B. inopinata BO1 and slightly higher than that of B. microti CCM 4915. In the BALB/c murine model of infection these strains replicated in both spleen and liver, but were less efficient than B. suis 1330. Some strains survived in the mammalian host for up to 12 weeks. The heterogeneity of these novel strains hampers a single species description but their phenotypic and genetic features suggest that they represent an evolutionary link between a soil-associated ancestor and the mammalian host-adapted pathogenic Brucella species. PMID:28300153

  17. A Role for the Chemokine Receptor CCR6 in Mammalian Sperm Motility and Chemotaxis

    PubMed Central

    Caballero-Campo, Pedro; Buffone, Mariano G.; Benencia, Fabian; Conejo-García, José R.; Rinaudo, Paolo F.; Gerton, George L.

    2013-01-01

    Although recent evidence indicates that several chemokines and defensins, well-known as inflammatory mediators, are expressed in the male and female reproductive tracts, the location and functional significance of chemokine networks in sperm physiology and sperm reproductive tract interactions are poorly understood. To address this deficiency in our knowledge, we examined the expression and function in sperm of CCR6, a receptor common to several chemoattractant peptides, and screened several reproductive tract fluids for the presence of specific ligands. CCR6 protein is present in mouse and human sperm and mainly localized in the sperm tail with other minor patterns in sperm from mice (neck and acrosomal region) and men (neck and midpiece regions). As expected from the protein immunoblotting and immunofluorescence results, mouse Ccr6 mRNA is expressed in the testis. Furthermore, the Defb29 mRNA encoding the CCR6 ligand, β-defensin DEFB29, is expressed at high levels in the epididymis. As determined by protein chip analysis, several chemokines (including some that act through CCR6, such as CCL20/MIP-3α (formerly Macrophage Inflammatory Protein 3α) and protein hormones were present in human follicular fluid, endometrial secretions, and seminal plasma. In functional chemotaxis assays, capacitated human sperm exhibited a directional movement towards CCL20, and displayed modifications in motility parameters. Our data indicate that chemokine ligand/receptor interactions in the male and female genital tracts promote sperm motility and chemotaxis under non-inflammatory conditions. Therefore, some of the physiological reactions mediated by CCR6 ligands in male reproduction extend beyond a pro-inflammatory response and might find application in clinical reproduction and/or contraception. PMID:23765988

  18. Planar polarity of ependymal cilia.

    PubMed

    Kishimoto, Norihito; Sawamoto, Kazunobu

    2012-02-01

    Ependymal cells, epithelial cells that line the cerebral ventricles of the adult brain in various animals, extend multiple motile cilia from their apical surface into the ventricles. These cilia move rapidly, beating in a direction determined by the ependymal planar cell polarity (PCP). Ciliary dysfunction interferes with cerebrospinal fluid circulation and alters neuronal migration. In this review, we summarize recent studies on the cellular and molecular mechanisms underlying two distinct types of ependymal PCP. Ciliary beating in the direction of fluid flow is established by a combination of hydrodynamic forces and intracellular planar polarity signaling. The ciliary basal bodies' anterior position on the apical surface of the cell is determined in the embryonic radial glial cells, inherited by ependymal cells, and established by non-muscle myosin II in early postnatal development.

  19. IFT-Cargo Interactions and Protein Transport in Cilia.

    PubMed

    Lechtreck, Karl F

    2015-12-01

    The motile and sensory functions of cilia and flagella are indispensable for human health. Cilia assembly requires a dedicated protein shuttle, intraflagellar transport (IFT), a bidirectional motility of multi-megadalton protein arrays along ciliary microtubules. IFT functions as a protein carrier delivering hundreds of distinct proteins into growing cilia. IFT-based protein import and export continue in fully grown cilia and are required for ciliary maintenance and sensing. Large ciliary building blocks might depend on IFT to move through the transition zone, which functions as a ciliary gate. Smaller, freely diffusing proteins, such as tubulin, depend on IFT to be concentrated or removed from cilia. As I discuss here, recent work provides insights into how IFT interacts with its cargoes and how the transport is regulated.

  20. Inversin, Wnt signaling and primary cilia.

    PubMed

    Lienkamp, Soeren; Ganner, Athina; Walz, Gerd

    2012-02-01

    Mutations of the ankyrin-repeat protein Inversin, a member of a diverse family of more than 12 proteins, cause nephronophthisis (NPH), an autosomal recessive cystic kidney disease associated with extra-renal manifestations such as retinitis pigmentosa, cerebellar aplasia and situs inversus. Most NPH gene products (NPHPs) localize to the cilium, and appear to control the transport of cargo protein to the cilium by forming functional networks. Inversin interacts with NPHP1 and NPHP3, and shares with NPHP4 the ability to antagonize Dishevelled-stimulated canonical Wnt signaling, potentially through recruitment of the Anaphase Promoting Complex (APC/C). However, Dishevelled antagonism may be confined towards the basal body, thereby polarizing motile cilia on the cells of the ventral node and respiratory tract. Inversin is essential for recruiting Dishevelled to the plasma membrane in response to activated Frizzled, a crucial step in planar cell polarity signaling. During vertebrate pronephros development, the Inversin-mediated translocation of Dishevelled appears to orchestrate the migration of cells and differentiation of segments that correspond to the mammalian loop of Henle. Thus, defective tubule migration and elongation may contribute to concentration defects and cause cyst formation in patients with NPH. Copyright © 2011 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  1. Bbof1 is required to maintain cilia orientation

    PubMed Central

    Chien, Yuan-Hung; Werner, Michael E.; Stubbs, Jennifer; Joens, Matt S.; Li, Julie; Chien, Shu; Fitzpatrick, James A. J.; Mitchell, Brian J.; Kintner, Chris

    2013-01-01

    Multiciliate cells (MCCs) are highly specialized epithelial cells that employ hundreds of motile cilia to produce a vigorous directed flow in a variety of organ systems. The production of this flow requires the establishment of planar cell polarity (PCP) whereby MCCs align hundreds of beating cilia along a common planar axis. The planar axis of cilia in MCCs is known to be established via the PCP pathway and hydrodynamic cues, but the downstream steps required for cilia orientation remain poorly defined. Here, we describe a new component of cilia orientation, based on the phenotypic analysis of an uncharacterized coiled-coil protein, called bbof1. We show that the expression of bbof1 is induced during the early phases of MCC differentiation by the master regulator foxj1. MCC differentiation and ciliogenesis occurs normally in embryos where bbof1 activity is reduced, but cilia orientation is severely disrupted. We show that cilia in bbof1 mutants can still respond to patterning and hydrodynamic cues, but lack the ability to maintain their precise orientation. Misexpression of bbof1 promotes cilia alignment, even in the absence of flow or in embryos where microtubules and actin filaments are disrupted. Bbof1 appears to mediate cilia alignment by localizing to a polar structure adjacent to the basal body. Together, these results suggest that bbof1 is a basal body component required in MCCs to align and maintain cilia orientation in response to flow. PMID:23900544

  2. Paramecium BBS genes are key to presence of channels in Cilia

    PubMed Central

    2012-01-01

    Background Changes in genes coding for ciliary proteins contribute to complex human syndromes called ciliopathies, such as Bardet-Biedl Syndrome (BBS). We used the model organism Paramecium to focus on ciliary ion channels that affect the beat form and sensory function of motile cilia and evaluate the effects of perturbing BBS proteins on these channels. Methods We used immunoprecipitations and mass spectrometry to explore whether Paramecium proteins interact as in mammalian cells. We used RNA interference (RNAi) and swimming behavior assays to examine the effects of BBS depletion on ciliary ion channels that control ciliary beating. Combining RNA interference and epitope tagging, we examined the effects of BBS depletion of BBS 7, 8 and 9 on the location of three channels and a chemoreceptor in cilia. Results We found 10 orthologs of 8 BBS genes in P. tetraurelia. BBS1, 2, 4, 5, 7, 8 and 9 co-immunoprecipitate. While RNAi reduction of BBS 7 and 9 gene products caused loss and shortening of cilia, RNAi for all BBS genes except BBS2 affected patterns of ciliary motility that are governed by ciliary ion channels. Swimming behavior assays pointed to loss of ciliary K+ channel function. Combining RNAi and epitope tagged ciliary proteins we demonstrated that a calcium activated K+ channel was no longer located in the cilia upon depletion of BBS 7, 8 or 9, consistent with the cells’ swimming behavior. The TRPP channel PKD2 was also lost from the cilia. In contrast, the ciliary voltage gated calcium channel was unaffected by BBS depletion, consistent with behavioral assays. The ciliary location of a chemoreceptor for folate was similarly unperturbed by the depletion of BBS 7, 8 or 9. Conclusions The co-immunoprecipitation of BBS 1,2,4,5,7,8, and 9 suggests a complex of BBS proteins. RNAi for BBS 7, 8 or 9 gene products causes the selective loss of K+ and PKD2 channels from the cilia while the critical voltage gated calcium channel and a peripheral receptor protein remain

  3. Primary cilia in the basal cells of equine epididymis: a serendipitous finding.

    PubMed

    Arrighi, Silvana

    2013-04-01

    Occurrence of a solitary cilium was an unexpected discovery while studying the ultrastructure of epididymal epithelium in equidae. Primary cilia were detected in epididymal basal cells of all individuals of the equines studied - horses, donkey and mules - independently from age and tract of the duct, emerging from the basal cell surface and insinuating into the intercellular spaces. More rarely solitary cilia occurred also at the luminal surface of the principal cells. The ciliary apparatus was constituted by a structurally typical basal body continuous with the finger-like ciliary shaft extending from the cell surface, and an adjacent centriole oriented at right angles to the basal body. The cilium was structured as the typical primary, non-motile cilia found in many mammalian cells, having a 9+0 microtubular pattern. The basal diplosome was randomly associated with other cellular organelles including the Golgi complex, the endoplasmic reticulum, the microfilament network, the plasma membrane, vesicles and pits. Primary ciliogenesis is a new and unexpected finding in the epididymal epithelium. A monitoring role of luminal factors and extracellular liquids might be attributed to this organelle, likely acting as chemical receptor of the luminal environment, thus modulating the epithelial function by a cell-to-cell crosstalk involving the entire epithelium.

  4. An age of enlightenment for cilia: The FASEB Summer Research Conference on the “Biology of Cilia and Flagella”

    PubMed Central

    Tran, Pamela V.; Lechtreck, Karl F.

    2015-01-01

    From July 19–24, 2015, 169 clinicians and basic scientists gathered in the vertiginous heights of Snowmass, Colorado (2,502 m) for the fourth FASEB summer research conference on the ‘Biology of Cilia and Flagella’. Organizers Maureen Barr (Rutgers University), Iain Drummond (Massachusetts General Hospital/Harvard Medical School), and Jagesh Shah (Brigham and Women’s Hospital/Harvard Medical School) assembled a program filled with new data and forward-thinking ideas documenting the ongoing growth of the field. Sixty oral presentations and 77 posters covered novel aspects of cilia structure, ciliogenesis, cilia motility, cilia-mediated signaling, and cilia-related disease. In this report, we summarize the meeting, highlight exciting developments and discuss open questions. PMID:26597000

  5. An age of enlightenment for cilia: The FASEB summer research conference on the "Biology of Cilia and Flagella".

    PubMed

    Tran, Pamela V; Lechtreck, Karl F

    2016-01-15

    From July 19-24, 2015, 169 clinicians and basic scientists gathered in the vertiginous heights of Snowmass, Colorado (2502 m) for the fourth FASEB summer research conference on the 'Biology of Cilia and Flagella'. Organizers Maureen Barr (Rutgers University), Iain Drummond (Massachusetts General Hospital/Harvard Medical School), and Jagesh Shah (Brigham and Women's Hospital/Harvard Medical School) assembled a program filled with new data and forward-thinking ideas documenting the ongoing growth of the field. Sixty oral presentations and 77 posters covered novel aspects of cilia structure, ciliogenesis, cilia motility, cilia-mediated signaling, and cilia-related disease. In this report, we summarize the meeting, highlight exciting developments and discuss open questions.

  6. Dynamics of cilia length in left–right development

    PubMed Central

    2017-01-01

    Reduction in the length of motile cilia in the zebrafish left–right organizer (LRO), also known as Kupffer's vesicle, has a large impact on left–right development. Here we demonstrate through genetic overexpression in zebrafish embryos and mathematical modelling that the impact of increased motile cilia length in embryonic LRO fluid flow is milder than that of short cilia. Through Arl13b overexpression, which increases cilia length without impacting cilia beat frequency, we show that the increase in cilium length is associated with a decrease in beat amplitude, resulting in similar flow strengths for Arl13b overexpression and wild-type (WT) embryos, which were not predicted by current theory. Longer cilia exhibit pronounced helical beat patterns and, consequently, lower beat amplitudes relative to WT, a result of an elastohydrodynamic shape transition. For long helical cilia, fluid dynamics modelling predicts a mild (approx. 12%) reduction in the torque exerted on the fluid relative to the WT, resulting in a proportional reduction in flow generation. This mild reduction is corroborated by experiments, providing a mechanism for the mild impact on organ situs. PMID:28405397

  7. STUDIES ON CILIA

    PubMed Central

    Satir, Peter

    1963-01-01

    Upon excision into spring water, the lateral cilia of the gill of the freshwater mussel Elliptio complanatus (Solander) stop beating, but 0.04 M potassium ion can activate the gill so that these cilia again beat with metachronal rhythm. One per cent osmium tetroxide quickly pipetted onto a fully activated gill fixes the lateral cilia in a pattern that preserves the form and arrangement of the metachronal wave, and permits the cilia to be studied with the electron microscope in all stages of their beat cycle. Changes are seen in the fixed active preparation that are not present in the inactive control, i.e., in the packing of the cilia, the position of the axis of the ciliary cross-section, and the diameter of the ring of peripheral filaments. Analysis of these parameters may lead to new correlations between ciliary fine structure and function. PMID:14079494

  8. Left-right organizer flow dynamics: how much cilia activity reliably yields laterality?

    PubMed

    Sampaio, Pedro; Ferreira, Rita R; Guerrero, Adán; Pintado, Petra; Tavares, Bárbara; Amaro, Joana; Smith, Andrew A; Montenegro-Johnson, Thomas; Smith, David J; Lopes, Susana S

    2014-06-23

    Internal organs are asymmetrically positioned inside the body. Embryonic motile cilia play an essential role in this process by generating a directional fluid flow inside the vertebrate left-right organizer. Detailed characterization of how fluid flow dynamics modulates laterality is lacking. We used zebrafish genetics to experimentally generate a range of flow dynamics. By following the development of each embryo, we show that fluid flow in the left-right organizer is asymmetric and provides a good predictor of organ laterality. This was tested in mosaic organizers composed of motile and immotile cilia generated by dnah7 knockdowns. In parallel, we used simulations of fluid dynamics to analyze our experimental data. These revealed that fluid flow generated by 30 or more cilia predicts 90% situs solitus, similar to experimental observations. We conclude that cilia number, dorsal anterior motile cilia clustering, and left flow are critical to situs solitus via robust asymmetric charon expression.

  9. Sensing a Sensor: Identifying the Mechanosensory Function of Primary Cilia

    PubMed Central

    Prasad, Rahul M.; Jin, Xingjian; Nauli, Surya M.

    2014-01-01

    Over the past decade, primary cilia have emerged as the premier means by which cells sense and transduce mechanical stimuli. Primary cilia are sensory organelles that have been shown to be vitally involved in the mechanosensation of urine in the renal nephron, bile in the hepatic biliary system, digestive fluid in the pancreatic duct, dentin in dental pulp, lacunocanalicular fluid in bone and cartilage, and blood in vasculature. The prevalence of primary cilia among mammalian cell types is matched by the tremendously varied disease states caused by both structural and functional defects in cilia. In the process of delineating the mechanisms behind these disease states, calcium fluorimetry has been widely utilized as a means of quantifying ciliary function to both fluid flow and pharmacological agents. In this review, we will discuss the approaches used in associating calcium levels to cilia function. PMID:24839551

  10. FOXJ1 prevents cilia growth inhibition by cigarette smoke in human airway epithelium in vitro.

    PubMed

    Brekman, Angelika; Walters, Matthew S; Tilley, Ann E; Crystal, Ronald G

    2014-11-01

    Airway epithelium ciliated cells play a central role in clearing the lung of inhaled pathogens and xenobiotics, and cilia length and coordinated beating are important for airway clearance. Based on in vivo studies showing that the airway epithelium of healthy smokers has shorter cilia than that of healthy nonsmokers, we investigated the mechanisms involved in cigarette smoke-mediated inhibition of ciliogenesis by assessing normal human airway basal cell differentiation in air-liquid interface (ALI) cultures in the presence of nontoxic concentrations of cigarette smoke extract (CSE). Measurements of cilia length from Day 28 ALI cultures demonstrated that CSE exposure was associated with shorter cilia (P < 0.05), reproducing the effect of cigarette smoking on cilia length observed in vivo. This phenotype correlated with a broad CSE-mediated suppression of genes involved in cilia-related transcriptional regulation, intraflagellar transport, cilia motility, structural integrity, and basal body development but not of control genes or epithelial barrier integrity. The CSE-mediated inhibition of cilia growth could be prevented by lentivirus-mediated overexpression of FOXJ1, the major cilia-related transcription factor, which led to partial reversal of expression of cilia-related genes suppressed by CSE. Together, the data suggest that components of cigarette smoke are responsible for a broad suppression of genes involved in cilia growth, but, by stimulating ciliogenesis with the transcription factor FOXJ1, it may be possible to maintain close to normal cilia length despite the stress of cigarette smoking.

  11. Visualizing renal primary cilia.

    PubMed

    Deane, James A; Verghese, Elizabeth; Martelotto, Luciano G; Cain, Jason E; Galtseva, Alya; Rosenblum, Norman D; Watkins, D Neil; Ricardo, Sharon D

    2013-03-01

    Renal primary cilia are microscopic sensory organelles found on the apical surface of epithelial cells of the nephron and collecting duct. They are based upon a microtubular cytoskeleton, bounded by a specialized membrane, and contain an array of proteins that facilitate their assembly, maintenance and function. Cilium-based signalling is important for the control of epithelial differentiation and has been implicated in the pathogenesis of various cystic kidney diseases and in renal repair. As such, visualizing renal primary cilia and understanding their composition has become an essential component of many studies of inherited kidney disease and mechanisms of epithelial regeneration. Primary cilia were initially identified in the kidney using electron microscopy and this remains a useful technique for the high resolution examination of these organelles. New reagents and techniques now also allow the structure and composition of primary cilia to be analysed in detail using fluorescence microscopy. Primary cilia can be imaged in situ in sections of kidney, and many renal-derived cell lines produce primary cilia in culture providing a simplified and accessible system in which to investigate these organelles. Here we outline microscopy-based techniques commonly used for studying renal primary cilia.

  12. Novel roles for the radial spoke head protein 9 in neural and neurosensory cilia

    PubMed Central

    Sedykh, Irina; TeSlaa, Jessica J.; Tatarsky, Rose L.; Keller, Abigail N.; Toops, Kimberly A.; Lakkaraju, Aparna; Nyholm, Molly K.; Wolman, Marc A.; Grinblat, Yevgenya

    2016-01-01

    Cilia are cell surface organelles with key roles in a range of cellular processes, including generation of fluid flow by motile cilia. The axonemes of motile cilia and immotile kinocilia contain 9 peripheral microtubule doublets, a central microtubule pair, and 9 connecting radial spokes. Aberrant radial spoke components RSPH1, 3, 4a and 9 have been linked with primary ciliary dyskinesia (PCD), a disorder characterized by ciliary dysmotility; yet, radial spoke functions remain unclear. Here we show that zebrafish Rsph9 is expressed in cells bearing motile cilia and kinocilia, and localizes to both 9 + 2 and 9 + 0 ciliary axonemes. Using CRISPR mutagenesis, we show that rsph9 is required for motility of presumptive 9 + 2 olfactory cilia and, unexpectedly, 9 + 0 neural cilia. rsph9 is also required for the structural integrity of 9 + 2 and 9 + 0 ciliary axonemes. rsph9 mutant larvae exhibit reduced initiation of the acoustic startle response consistent with hearing impairment, suggesting a novel role for Rsph9 in the kinocilia of the inner ear and/or lateral line neuromasts. These data identify novel roles for Rsph9 in 9 + 0 motile cilia and in sensory kinocilia, and establish a useful zebrafish PCD model. PMID:27687975

  13. Small GTPases and cilia.

    PubMed

    Li, Yujie; Hu, Jinghua

    2011-01-01

    Small GTPases are key molecular switches that bind and hydrolyze GTP in diverse membrane- and cytoskeleton-related cellular processes. Recently, mounting evidences have highlighted the role of various small GTPases, including the members in Arf/Arl, Rab, and Ran subfamilies, in cilia formation and function. Once overlooked as an evolutionary vestige, the primary cilium has attracted more and more attention in last decade because of its role in sensing various extracellular signals and the association between cilia dysfunction and a wide spectrum of human diseases, now called ciliopathies. Here we review recent advances about the function of small GTPases in the context of cilia, and the correlation between the functional impairment of small GTPases and ciliopathies. Understanding of these cellular processes is of fundamental importance for broadening our view of cilia development and function in normal and pathological states and for providing valuable insights into the role of various small GTPases in disease processes, and their potential as therapeutic targets.

  14. Cildb: a knowledgebase for centrosomes and cilia.

    PubMed

    Arnaiz, Olivier; Malinowska, Agata; Klotz, Catherine; Sperling, Linda; Dadlez, Michal; Koll, France; Cohen, Jean

    2009-01-01

    Ciliopathies, pleiotropic diseases provoked by defects in the structure or function of cilia or flagella, reflect the multiple roles of cilia during development, in stem cells, in somatic organs and germ cells. High throughput studies have revealed several hundred proteins that are involved in the composition, function or biogenesis of cilia. The corresponding genes are potential candidates for orphan ciliopathies. To study ciliary genes, model organisms are used in which particular questions on motility, sensory or developmental functions can be approached by genetics. In the course of high throughput studies of cilia in Paramecium tetraurelia, we were confronted with the problem of comparing our results with those obtained in other model organisms. We therefore developed a novel knowledgebase, Cildb, that integrates ciliary data from heterogeneous sources. Cildb links orthology relationships among 18 species to high throughput ciliary studies, and to OMIM data on human hereditary diseases. The web interface of Cildb comprises three tools, BioMart for complex queries, BLAST for sequence homology searches and GBrowse for browsing the human genome in relation to OMIM information for human diseases. Cildb can be used for interspecies comparisons, building candidate ciliary proteomes in any species, or identifying candidate ciliopathy genes.Database URL:http://cildb.cgm.cnrs-gif.fr.

  15. Emergence of multiple synchronization modes in hydrodynamically-coupled cilia

    NASA Astrophysics Data System (ADS)

    Guo, Hanliang; Kanso, Eva

    2016-11-01

    Motile cilia and flagella exhibit different phase coordinations. For example, closely swimming spermatozoa are observed to synchronize together; bi-flagellates Chlamydomonas regulate the flagella in a "breast-stroke" fashion; cilia on the surface of Paramecium beat in a fixed phase lag in an orchestrated wave like fashion. Experimental evidence suggests that phase coordinations can be achieved solely via hydrodynamical interactions. However, the exact mechanisms behind it remain illusive. Here, adapting a "geometric switch" model, we observe different synchronization modes in pairs of hydrodynamically-coupled cilia by changing physical parameters such as the strength of the cilia internal motor and the separation distance between cilia. Interestingly, we find regions in the parameter space where the coupled cilia reach stable phase coordinations and regions where the phase coordinations are sensitive to perturbations. We also find that leaning into the fluid reduces the sensitivity to perturbations, and produces stable phase coordination that is neither in-phase nor anti-phase, which could explain the origin of metachronal waves in large cilia populations.

  16. Primary Cilia Are Not Calcium-Responsive Mechanosensors

    PubMed Central

    Delling, M.; Indzhykulian, A. A.; Liu, X.; Liu, Y.; Xie, T.; Corey, D. P.; Clapham, D. E.

    2016-01-01

    Primary cilia are solitary, generally non-motile, hair-like protrusions that extend from the surface of cells between cell divisions. Their antenna-like structure leads naturally to the assumption that they sense the surrounding environment, the most common hypothesis being sensation of mechanical force through calcium-permeable ion channels within the cilium1. This Ca2+- Responsive MechanoSensor (CaRMS) hypothesis for primary cilia has been invoked to explain a large range of biological responses, from control of left-right axis determination in embryonic development to adult progression of polycystic kidney disease and some cancers2,3. Here, we report the complete lack of mechanically induced calcium increases in primary cilia, in tissues upon which this hypothesis has been based. First, we developed a transgenic mouse, Arl13b-mCherry-GECO1.2, expressing a ratiometric genetically encoded calcium indicator (GECI) in all primary cilia. We then measured responses to flow in primary cilia of cultured kidney epithelial cells, kidney thick ascending tubules, crown cells of the embryonic node, kinocilia of inner ear hair cells, and several cell lines. Cilia-specific Ca2+ influxes were not observed in physiological or even highly supraphysiological levels of fluid flow. We conclude that mechanosensation, if it originates in primary cilia, is not via calcium signaling. PMID:27007841

  17. [The importance of model organisms to study cilia and flagella biology].

    PubMed

    Vincensini, Laetitia; Blisnick, Thierry; Bastin, Philippe

    2011-01-01

    Cilia and flagella are ubiquitous organelles that protrude from the surfaces of many cells, and whose architecture is highly conserved from protists to humans. These complex organelles, composed of over 500 proteins, can be either immotile or motile. They are involved in a myriad of biological processes, including sensing (non-motile cilia) and/or cell motility or movement of extracellular fluids (motile cilia). The ever-expanding list of human diseases linked to defective cilia illustrates the functional importance of cilia and flagella. These ciliopathies are characterised by an impressive diversity of symptoms and an often complex genetic etiology. A precise knowledge of cilia and flagella biology is thus critical to better understand these pathologies. However, multi-ciliated cells are terminally differentiated and difficult to manipulate, and a primary cilium is assembled only when the cell exits from the cell cycle. In this context the use of model organisms, that relies on the high degree of structural but also of molecular conservation of these organelles across evolution, is instrumental to decipher the many facets of cilia and flagella biology. In this review, we highlight the specific strengths of the main model organisms to investigate the molecular composition, mode of assembly, sensing and motility mechanisms and functions of cilia and flagella. Pioneering studies carried out in the green alga Chlamydomonas established the link between cilia and several genetic diseases. Moreover, multicellular organisms such as mouse, zebrafish, Xenopus, C. elegans or Drosophila, and protists like Paramecium, Tetrahymena and Trypanosoma or Leishmania each bring specific advantages to the study of cilium biology. For example, the function of genes involved in primary ciliary dyskinesia (due to defects in ciliary motility) can be efficiently assessed in trypanosomes. © Société de Biologie, 2011.

  18. In vivo micro-scale tomography of ciliary behavior in the mammalian oviduct

    PubMed Central

    Wang, Shang; Burton, Jason C.; Behringer, Richard R.; Larina, Irina V.

    2015-01-01

    Motile cilia in the mammalian oviduct play a key role in reproduction, such as transporting fertilized oocytes to the uterus for implantation. Due to their small size (~5–10 μm in length and ~300 nm in diameter), live visualization of cilia and their activity in the lumen of the oviduct through tissue layers represents a major challenge not yet overcome. Here, we report a functional low-coherence optical imaging technique that allows in vivo depth-resolved mapping of the cilia location and cilia beat frequency (CBF) in the intact mouse oviduct with micro-scale spatial resolution. We validate our approach with widely-used microscopic imaging methods, present the first in vivo mapping of the oviduct CBF in its native context, and demonstrate the ability of this approach to differentiate CBF in different locations of the oviduct at different post-conception stages. This technique opens a range of opportunities for live studies in reproductive medicine as well as other areas focused on cilia activity and related ciliopathies. PMID:26279472

  19. Primary cilia disappear in rat podocytes during glomerular development.

    PubMed

    Ichimura, Koichiro; Kurihara, Hidetake; Sakai, Tatsuo

    2010-07-01

    Most tubular epithelial cell types express primary cilia, and mutations of primary-cilium-associated proteins are well known to cause several kinds of cystic renal disease. However, until now, it has been unclear whether mammalian podocytes express primary cilia in vivo. In this study, we determined whether primary cilia are present in the podocytes of rat immature and mature glomeruli by means of transmission electron microscopy of serial ultrathin sections. In immature glomeruli of fetal rats, podocytes express the primary cilia with high percentages at the S-shaped body (88 +/- 5%, n = 3), capillary loop (95 +/- 4%, n = 4), and maturing glomerulus (76 +/- 13%, n = 5) stages. The percentage of ciliated podocytes was significantly lower at the maturing glomerulus stage than at the former two stages. In mature glomeruli of adult rats, ciliated podocytes were not found at all (0 +/- 0%, n = 11). These findings indicate that the primary cilia gradually disappear in rat podocytes during glomerular development. Since glomerular filtration rate increases during development, the primary cilia on the podocytes are subjected to a stronger bending force. Thus, the disappearance of the primary cilia presumably prevents the entry of excessive calcium-ions via the cilium-associated polycystin complexes and the disturbance of intracellular signaling cascades in mature podocytes.

  20. Primary cilia regulate hippocampal neurogenesis by mediating sonic hedgehog signaling

    PubMed Central

    Breunig, Joshua J.; Sarkisian, Matthew R.; Arellano, Jon I.; Morozov, Yury M.; Ayoub, Albert E.; Sojitra, Sonal; Wang, Baolin; Flavell, Richard A.; Rakic, Pasko; Town, Terrence

    2008-01-01

    Primary cilia are present on mammalian neurons and glia, but their function is largely unknown. We generated conditional homozygous mutant mice for a gene we termed Stumpy. Mutants lack cilia and have conspicuous abnormalities in postnatally developing brain regions, including a hypoplasic hippocampus characterized by a primary deficiency in neural stem cells known as astrocyte-like neural precursors (ALNPs). Previous studies suggested that primary cilia mediate sonic hedgehog (Shh) signaling. Here, we find that loss of ALNP cilia leads to abrogated Shh activity, increased cell cycle exit, and morphological abnormalities in ALNPs. Processing of Gli3, a mediator of Shh signaling, is also altered in the absence of cilia. Further, key mediators of the Shh pathway localize to ALNP cilia. Thus, selective targeting of Shh machinery to primary cilia confers to ALNPs the ability to differentially respond to Shh mitogenic signals compared to neighboring cells. Our data suggest these organelles are cellular “antennae” critically required to modulate ALNP behavior. PMID:18728187

  1. Cilia biology: stop overeating now!

    PubMed

    Satir, Peter

    2007-11-20

    Knocking out primary cilia of adult mouse tissues or a specific subset of cilia from POMC-expressing neurons in the brain initiates uncontrolled eating. This behavior leads to obesity and kidney disease.

  2. Cilia and Diseases

    PubMed Central

    Brown, Jason M.; Witman, George B.

    2014-01-01

    In recent decades, cilia have moved from relative obscurity to a position of importance for understanding multiple complex human diseases. Now termed the ciliopathies, these diseases inflict devastating effects on millions of people worldwide. In this review, written primarily for teachers and students who may not yet be aware of the recent exciting developments in this field, we provide a general overview of our current understanding of cilia and human disease. We start with an introduction to cilia structure and assembly and indicate where they are found in the human body. We then discuss the clinical features of selected ciliopathies, with an emphasis on primary ciliary dyskinesia, polycystic kidney disease, and retinal degeneration. The history of ciliopathy research involves a fascinating interplay between basic and clinical sciences, highlighted in a timeline. Finally, we summarize the relative strengths of individual model organisms for ciliopathy research; many of these are suitable for classroom use. PMID:25960570

  3. Early eukaryotic origins for cilia-associated bioactive peptide-amidating activity.

    PubMed

    Kumar, Dhivya; Blaby-Haas, Crysten E; Merchant, Sabeeha S; Mains, Richard E; King, Stephen M; Eipper, Betty A

    2016-03-01

    Ciliary axonemes and basal bodies were present in the last eukaryotic common ancestor and play crucial roles in sensing and responding to environmental cues. Peptidergic signaling, generally considered a metazoan innovation, is essential for organismal development and homeostasis. Peptidylglycine α-amidating monooxygenase (PAM) is crucial for the last step of bioactive peptide biosynthesis. However, identification of a complete PAM-like gene in green algal genomes suggests ancient evolutionary roots for bioactive peptide signaling. We demonstrate that the Chlamydomonas reinhardtii PAM gene encodes an active peptide-amidating enzyme (CrPAM) that shares key structural and functional features with the mammalian enzyme, indicating that components of the peptide biosynthetic pathway predate multicellularity. In addition to its secretory pathway localization, CrPAM localizes to cilia and tightly associates with the axonemal superstructure, revealing a new axonemal enzyme activity. This localization pattern is conserved in mammals, with PAM present in both motile and immotile sensory cilia. The conserved ciliary localization of PAM adds to the known signaling capabilities of the eukaryotic cilium and provides a potential mechanistic link between peptidergic signaling and endocrine abnormalities commonly observed in ciliopathies.

  4. Early eukaryotic origins for cilia-associated bioactive peptide-amidating activity

    PubMed Central

    Kumar, Dhivya; Blaby-Haas, Crysten E.; Merchant, Sabeeha S.; Mains, Richard E.; King, Stephen M.; Eipper, Betty A.

    2016-01-01

    ABSTRACT Ciliary axonemes and basal bodies were present in the last eukaryotic common ancestor and play crucial roles in sensing and responding to environmental cues. Peptidergic signaling, generally considered a metazoan innovation, is essential for organismal development and homeostasis. Peptidylglycine α-amidating monooxygenase (PAM) is crucial for the last step of bioactive peptide biosynthesis. However, identification of a complete PAM-like gene in green algal genomes suggests ancient evolutionary roots for bioactive peptide signaling. We demonstrate that the Chlamydomonas reinhardtii PAM gene encodes an active peptide-amidating enzyme (CrPAM) that shares key structural and functional features with the mammalian enzyme, indicating that components of the peptide biosynthetic pathway predate multicellularity. In addition to its secretory pathway localization, CrPAM localizes to cilia and tightly associates with the axonemal superstructure, revealing a new axonemal enzyme activity. This localization pattern is conserved in mammals, with PAM present in both motile and immotile sensory cilia. The conserved ciliary localization of PAM adds to the known signaling capabilities of the eukaryotic cilium and provides a potential mechanistic link between peptidergic signaling and endocrine abnormalities commonly observed in ciliopathies. PMID:26787743

  5. Sensory cilia and integration of signal transduction in human health and disease.

    PubMed

    Christensen, Søren T; Pedersen, Lotte B; Schneider, Linda; Satir, Peter

    2007-02-01

    The primary cilium is a hallmark of mammalian tissue cells. Recent research has shown that these organelles display unique sets of selected signal transduction modules including receptors, ion channels, effector proteins and transcription factors that relay chemical and physical stimuli from the extracellular environment in order to control basic cellular processes during embryonic and postnatal development, as well as in tissue homeostasis in adulthood. Consequently, defects in building of the cilium or in transport or function of ciliary signal proteins are associated with a series of pathologies, including developmental disorders and cancer. In this review, we highlight recent examples of the mechanisms by which signal components are selectively targeted and transported to the ciliary membrane and we present an overview of the signal transduction pathways associated with primary and motile cilia in vertebrate cells, including platelet-derived growth factor receptor-alpha (PDGFRalpha), hedgehog and Wnt signaling pathways. Finally, we discuss the functions of these cilia-associated signal transduction pathways and their role in human health and development.

  6. Identification of protein tyrosine phosphatases and dual-specificity phosphatases in mammalian spermatozoa and their role in sperm motility and protein tyrosine phosphorylation.

    PubMed

    González-Fernández, L; Ortega-Ferrusola, C; Macias-Garcia, B; Salido, G M; Peña, F J; Tapia, J A

    2009-06-01

    rapid and reversible. Pervanadate also increased tyrosine phosphorylation of different proteins in capacitated and noncapacitated spermatozoa. Results showed that the phosphatases PTPN11, DUSP4, and DUSP3 are present in boar, stallion, and dog spermatozoa. PTPRB is also present in boar and stallion spermatozoa but was not detected in dog. The subcellular distribution of the identified phosphatases is diverse, suggesting that they likely have specific roles in sperm. Finally, PTP activity has a positive role in the regulation of motility and is involved in protein tyrosine phosphorylation in mammalian sperm.

  7. Primary cilia and autophagic dysfunction in Huntington's disease

    PubMed Central

    Kaliszewski, M; Knott, A B; Bossy-Wetzel, E

    2015-01-01

    Huntington's disease (HD) is an inherited, neurodegenerative disorder caused by a single-gene mutation: a CAG expansion in the huntingtin (HTT) gene that results in production of a mutated protein, mutant HTT, with a polyglutamine tail (polyQ-HTT). Although the molecular pathways of polyQ-HTT toxicity are not fully understood, because protein misfolding and aggregation are central features of HD, it has long been suspected that cellular housekeeping processes such as autophagy might be important to disease pathology. Indeed, multiple lines of research have identified abnormal autophagy in HD, characterized generally by increased autophagic induction and inefficient clearance of substrates. To date, the origin of autophagic dysfunction in HD remains unclear and the search for actors involved continues. To that end, recent studies have suggested a bidirectional relationship between autophagy and primary cilia, signaling organelles of most mammalian cells. Interestingly, primary cilia structure is defective in HD, suggesting a potential link between autophagic dysfunction, primary cilia and HD pathogenesis. In addition, because polyQ-HTT also accumulates in primary cilia, the possibility exists that primary cilia might play additional roles in HD: perhaps by disrupting signaling pathways or acting as a reservoir for secretion and propagation of toxic, misfolded polyQ-HTT fragments. Here, we review recent research suggesting potential links between autophagy, primary cilia and HD and speculate on possible pathogenic mechanisms and future directions for the field. PMID:26160070

  8. HIF Stabilization Weakens Primary Cilia

    PubMed Central

    Resnick, Andrew

    2016-01-01

    Although solitary or sensory cilia are present in most cells of the body and their existence has been known since the sixties, very little is known about their functions. One suspected function is fluid flow sensing- physical bending of cilia produces an influx of Ca++, which can then result in a variety of activated signaling pathways. Defective cilia and ciliary-associated proteins have been shown to result in cystic diseases. Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a progressive disease, typically appearing in the 5th decade of life and is one of the most common monogenetic inherited human diseases, affecting approximately 600,000 people in the United States. Because the mechanical properties of cilia impact their response to applied flow, we asked how the stiffness of cilia can be controlled pharmacologically. We performed an experiment subjecting cilia to Taxol (a microtubule stabilizer) and CoCl2 (a HIF stabilizer to model hypoxia). Madin-Darby Canine Kidney (MDCK) cells were selected as our model system. After incubation with a selected pharmacological agent, cilia were optically trapped and the bending modulus measured. We found that HIF stabilization significantly weakens cilia. These results illustrate a method to alter the mechanical properties of primary cilia and potentially alter the flow sensing properties of cilia. PMID:27812213

  9. HIF Stabilization Weakens Primary Cilia.

    PubMed

    Resnick, Andrew

    2016-01-01

    Although solitary or sensory cilia are present in most cells of the body and their existence has been known since the sixties, very little is known about their functions. One suspected function is fluid flow sensing- physical bending of cilia produces an influx of Ca++, which can then result in a variety of activated signaling pathways. Defective cilia and ciliary-associated proteins have been shown to result in cystic diseases. Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a progressive disease, typically appearing in the 5th decade of life and is one of the most common monogenetic inherited human diseases, affecting approximately 600,000 people in the United States. Because the mechanical properties of cilia impact their response to applied flow, we asked how the stiffness of cilia can be controlled pharmacologically. We performed an experiment subjecting cilia to Taxol (a microtubule stabilizer) and CoCl2 (a HIF stabilizer to model hypoxia). Madin-Darby Canine Kidney (MDCK) cells were selected as our model system. After incubation with a selected pharmacological agent, cilia were optically trapped and the bending modulus measured. We found that HIF stabilization significantly weakens cilia. These results illustrate a method to alter the mechanical properties of primary cilia and potentially alter the flow sensing properties of cilia.

  10. Evolutionarily Ancient Association of the FoxJ1 Transcription Factor with the Motile Ciliogenic Program

    PubMed Central

    Ho, Hao Kee; Babu, Deepak; Eitel, Michael; Narasimhan, Vijayashankaranarayanan; Tiku, Varnesh; Westbrook, Jody; Schierwater, Bernd; Roy, Sudipto

    2012-01-01

    It is generally believed that the last eukaryotic common ancestor (LECA) was a unicellular organism with motile cilia. In the vertebrates, the winged-helix transcription factor FoxJ1 functions as the master regulator of motile cilia biogenesis. Despite the antiquity of cilia, their highly conserved structure, and their mechanism of motility, the evolution of the transcriptional program controlling ciliogenesis has remained incompletely understood. In particular, it is presently not known how the generation of motile cilia is programmed outside of the vertebrates, and whether and to what extent the FoxJ1-dependent regulation is conserved. We have performed a survey of numerous eukaryotic genomes and discovered that genes homologous to foxJ1 are restricted only to organisms belonging to the unikont lineage. Using a mis-expression assay, we then obtained evidence of a conserved ability of FoxJ1 proteins from a number of diverse phyletic groups to activate the expression of a host of motile ciliary genes in zebrafish embryos. Conversely, we found that inactivation of a foxJ1 gene in Schmidtea mediterranea, a platyhelminth (flatworm) that utilizes motile cilia for locomotion, led to a profound disruption in the differentiation of motile cilia. Together, all of these findings provide the first evolutionary perspective into the transcriptional control of motile ciliogenesis and allow us to propose a conserved FoxJ1-regulated mechanism for motile cilia biogenesis back to the origin of the metazoans. PMID:23144623

  11. Antennas of organ morphogenesis: the roles of cilia in vertebrate kidney development

    PubMed Central

    Marra, Amanda N.; Li, Yue

    2016-01-01

    Abstract Cilia arose early during eukaryotic evolution, and their structural components are highly conserved from the simplest protists to complex metazoan species. In recent years, the role of cilia in the ontogeny of vertebrate organs has received increasing attention due to a staggering correlation between human disease and dysfunctional cilia. In particular, the presence of cilia in both the developing and mature kidney has become a deep area of research due to ciliopathies common to the kidney, such as polycystic kidney disease (PKD). Interestingly, mutations in genes encoding proteins that localize to the cilia cause similar cystic phenotypes in kidneys of various vertebrates, suggesting an essential role for cilia in kidney organogenesis and homeostasis as well. Importantly, the genes so far identified in kidney disease have conserved functions across species, whose kidneys include both primary and motile cilia. Here, we aim to provide a comprehensive description of cilia and their role in kidney development, as well as highlight the usefulness of the zebrafish embryonic kidney as a model to further understand the function of cilia in kidney health. PMID:27389733

  12. Novel Insights into the Development and Function of Cilia Using the Advantages of the Paramecium Cell and Its Many Cilia

    PubMed Central

    Yano, Junji; Valentine, Megan S.; Van Houten, Judith L.

    2015-01-01

    Paramecium species, especially P. tetraurelia and caudatum, are model organisms for modern research into the form and function of cilia. In this review, we focus on the ciliary ion channels and other transmembrane proteins that control the beat frequency and wave form of the cilium by controlling the signaling within the cilium. We put these discussions in the context of the advantages that Paramecium brings to the understanding of ciliary motility: mutants for genetic dissections of swimming behavior, electrophysiology, structural analysis, abundant cilia for biochemistry and modern proteomics, genomics and molecular biology. We review the connection between behavior and physiology, which allows the cells to broadcast the function of their ciliary channels in real time. We build a case for the important insights and advantages that this model organism continues to bring to the study of cilia. PMID:26230712

  13. Conservation of ciliary proteins in plants with no cilia

    PubMed Central

    2011-01-01

    Background Eukaryotic cilia are complex, highly conserved microtubule-based organelles with a broad phylogenetic distribution. Cilia were present in the last eukaryotic common ancestor and many proteins involved in cilia function have been conserved through eukaryotic diversification. However, cilia have also been lost multiple times in different lineages, with at least two losses occurring within the land plants. Whereas all non-seed plants produce cilia for motility of male gametes, some gymnosperms and all angiosperms lack cilia. During these evolutionary losses, proteins with ancestral ciliary functions may be lost or co-opted into different functions. Results Here we identify a core set of proteins with an inferred ciliary function that are conserved in ciliated eukaryotic species. We interrogate this genomic dataset to identify proteins with a predicted ancestral ciliary role that have been maintained in non-ciliated land plants. In support of our prediction, we demonstrate that several of these proteins have a flagellar localisation in protozoan trypanosomes. The phylogenetic distribution of these genes within the land plants indicates evolutionary scenarios of either sub- or neo-functionalisation and expression data analysis shows that these genes are highly expressed in Arabidopsis thaliana pollen cells. Conclusions A large number of proteins possess a phylogenetic ciliary profile indicative of ciliary function. Remarkably, many genes with an ancestral ciliary role are maintained in non-ciliated land plants. These proteins have been co-opted to perform novel functions, most likely before the loss of cilia, some of which appear related to the formation of the male gametes. PMID:22208660

  14. Effect of viscosity on metachrony in mucus propelling cilia.

    PubMed

    Gheber, L; Korngreen, A; Priel, Z

    1998-01-01

    In the present work we report that increasing the viscosity of the medium caused not only a decrease in the ciliary beat frequency but also changes in the metachrony and correlation between cilia. The study was performed using double and triple simultaneous photoelectric measurements on cultured ciliary cells from the frog esophagus in the viscosity range of 1-2,000 cp. We observed that increasing the viscosity intensified the fluctuations in all the measured parameters. Ciliary beat frequency decreased moderately. Even at quite high viscosities (circa 2000 cp.), cilia were still active with beating frequencies of 3-5 Hz. In addition, the degree of correlation between cilia parallel to the effective stroke direction (ESD) decreased, while that perpendicular to the ESD at a low range of viscosities remained unchanged and even increased at high viscosities. Medium viscosities in the range of 30-1,500 cp. altered the metachronal wave properties of cultured frog esophagus. The metachronal wavelength increased by up to 50%, and the wave direction changed towards more orthoplectic type of coordination. According to our recently suggested model [Gheber and Priel, 1990: Cell Motil. Cytoskeleton 16:167-181], these effects can be explained by a decrease in the temporal asymmetry of the ciliary beat. Since similar results were observed in water propelling cilia of Paramecium subjected to medium viscosity ranges of up to 40 cp. [Machemer, 1972: J. Exp. Biol. 57:239-259], we conclude that hydrodynamic interactions govern the metachronal wave properties of both mucus and water propelling cilia, though mucus propelling cilia, with their better adaptation to increased load, are affected at much higher viscosities than water propelling cilia.

  15. Primary cilia enhance kisspeptin receptor signaling on gonadotropin-releasing hormone neurons

    PubMed Central

    Koemeter-Cox, Andrew I.; Sherwood, Thomas W.; Green, Jill A.; Steiner, Robert A.; Berbari, Nicolas F.; Yoder, Bradley K.; Kauffman, Alexander S.; Monsma, Paula C.; Brown, Anthony; Askwith, Candice C.; Mykytyn, Kirk

    2014-01-01

    Most central neurons in the mammalian brain possess an appendage called a primary cilium that projects from the soma into the extracellular space. The importance of these organelles is highlighted by the fact that primary cilia dysfunction is associated with numerous neuropathologies, including hyperphagia-induced obesity, hypogonadism, and learning and memory deficits. Neuronal cilia are enriched for signaling molecules, including certain G protein-coupled receptors (GPCRs), suggesting that neuronal cilia sense and respond to neuromodulators in the extracellular space. However, the impact of cilia on signaling to central neurons has never been demonstrated. Here, we show that the kisspeptin receptor (Kiss1r), a GPCR that is activated by kisspeptin to regulate the onset of puberty and adult reproductive function, is enriched in cilia projecting from mouse gonadotropin-releasing hormone (GnRH) neurons. Interestingly, GnRH neurons in adult animals are multiciliated and the percentage of GnRH neurons possessing multiple Kiss1r-positive cilia increases during postnatal development in a progression that correlates with sexual maturation. Remarkably, disruption of cilia selectively on GnRH neurons leads to a significant reduction in kisspeptin-mediated GnRH neuronal activity. To our knowledge, this result is the first demonstration of cilia disruption affecting central neuronal activity and highlights the importance of cilia for proper GPCR signaling. PMID:24982149

  16. Biomimetic cilia arrays - fabrication, magnetic actuation, and driven fluid transport phenomena

    NASA Astrophysics Data System (ADS)

    Shields, Adam

    The cilium is one of biology's most basic functional nanostructures, present on nearly every cell and increasingly realized as vital to many aspects of human health. A fundamental reason for the ubiquity of cilia is their ability to effectively interact with fluids at the microscale, where the Reynolds number is low and thus inertia is irrelevant. This ability makes cilia an attractive and popular candidate for an engineered biomimic with potential applications in microfluidics and sensing. In addition, biological ciliated systems are difficult to study for many reasons, and so I demonstrate how a functional biomimetic system can also serve as a model platform for highly controlled studies of biologically relevant, cilia-driven hydrodynamics. Using the template-based microfabrication of a magnetic nanoparticle/polymer composite, I fabricate arrays of magnetically actuated biomimetic cilia at the scale of their biological analogues. I will discuss this fabrication technique and the magnetic actuation of these arrays to mimic the beat of biological cilia. I also report on the nature of the fluid flows driven by the cilia beat, and demonstrate how these cilia arrays can simultaneously generate long-range fluid transport and mixing in distinct fluid flow regimes. Finally, I present these results within the context of canonical hydrodynamics problems and discuss the implications for biological systems, such as the motile cilia recently discovered in the embryonic node.

  17. Cilia are required for asymmetric nodal induction in the sea urchin embryo.

    PubMed

    Tisler, Matthias; Wetzel, Franziska; Mantino, Sabrina; Kremnyov, Stanislav; Thumberger, Thomas; Schweickert, Axel; Blum, Martin; Vick, Philipp

    2016-08-23

    Left-right (LR) organ asymmetries are a common feature of metazoan animals. In many cases, laterality is established by a conserved asymmetric Nodal signaling cascade during embryogenesis. In most vertebrates, asymmetric nodal induction results from a cilia-driven leftward fluid flow at the left-right organizer (LRO), a ciliated epithelium present during gastrula/neurula stages. Conservation of LRO and flow beyond the vertebrates has not been reported yet. Here we study sea urchin embryos, which use nodal to establish larval LR asymmetry as well. Cilia were found in the archenteron of embryos undergoing gastrulation. Expression of foxj1 and dnah9 suggested that archenteron cilia were motile. Cilia were polarized to the posterior pole of cells, a prerequisite of directed flow. High-speed videography revealed rotating cilia in the archenteron slightly before asymmetric nodal induction. Removal of cilia through brief high salt treatments resulted in aberrant patterns of nodal expression. Our data demonstrate that cilia - like in vertebrates - are required for asymmetric nodal induction in sea urchin embryos. Based on these results we argue that the anterior archenteron represents a bona fide LRO and propose that cilia-based symmetry breakage is a synapomorphy of the deuterostomes.

  18. Primary cilia disappear in rat podocytes during glomerular development

    PubMed Central

    Kurihara, Hidetake; Sakai, Tatsuo

    2010-01-01

    Most tubular epithelial cell types express primary cilia, and mutations of primary-cilium-associated proteins are well known to cause several kinds of cystic renal disease. However, until now, it has been unclear whether mammalian podocytes express primary cilia in vivo. In this study, we determined whether primary cilia are present in the podocytes of rat immature and mature glomeruli by means of transmission electron microscopy of serial ultrathin sections. In immature glomeruli of fetal rats, podocytes express the primary cilia with high percentages at the S-shaped body (88 ± 5%, n = 3), capillary loop (95 ± 4%, n =  4), and maturing glomerulus (76 ± 13%, n = 5) stages. The percentage of ciliated podocytes was significantly lower at the maturing glomerulus stage than at the former two stages. In mature glomeruli of adult rats, ciliated podocytes were not found at all (0 ± 0%, n = 11). These findings indicate that the primary cilia gradually disappear in rat podocytes during glomerular development. Since glomerular filtration rate increases during development, the primary cilia on the podocytes are subjected to a stronger bending force. Thus, the disappearance of the primary cilia presumably prevents the entry of excessive calcium-ions via the cilium-associated polycystin complexes and the disturbance of intracellular signaling cascades in mature podocytes. Electronic supplementary material The online version of this article (doi:10.1007/s00441-010-0983-7) contains supplementary material, which is available to authorized users. PMID:20495826

  19. The formation and positioning of cilia in Ciona intestinalis embryos in relation to the generation and evolution of chordate left-right asymmetry.

    PubMed

    Thompson, Helen; Shaw, Michael K; Dawe, Helen R; Shimeld, Sebastian M

    2012-04-15

    In the early mouse embryo monocilia on the ventral node rotate to generate a leftward flow of fluid. This nodal flow is essential for the left-sided expression of nodal and pitx2, and for subsequent asymmetric organ patterning. Equivalent left fluid flow has been identified in other vertebrates, including Xenopus and zebrafish, indicating it is an ancient vertebrate mechanism. Asymmetric nodal and Pitx expression have also been identified in several invertebrates, including the vertebrates' nearest relatives, the urochordates. However whether cilia regulate this asymmetric gene expression remains unknown, and previous studies in urochordates have not identified any cilia prior to the larval stage, when asymmetry is already long established. Here we use Scanning and Transmission Electron Microscopy and immunofluorescence to investigate cilia in the urochordate Ciona intestinalis. We show that single cilia are transiently present on each ectoderm cell of the late neurula/early tailbud stage embryo, a time point just before onset of asymmetric nodal expression. Mapping the position of each cilium on these cells shows they are posteriorly positioned, something also described for mouse node cilia. The C. intestinalis cilia have a 9+0 ring ultrastructure, however we find no evidence of structures associated with motility such as dynein arms, radial spokes or nexin. Furthermore the 9+0 ring structure becomes disorganised immediately after the cilia have exited the cell, indicative of cilia which are not capable of motility. Our results indicate that although cilia are present prior to molecular asymmetries, they are not motile and hence cannot be operating in the same way as the flow-generating cilia of the vertebrate node. We conclude that the cilia may have a role in the development of C. intestinalis left-right asymmetry but that this would have to be in a sensory capacity, perhaps as mechanosensors as hypothesised in two-cilia physical models of vertebrate cilia

  20. Axonemal Positioning and Orientation in 3-D Space for Primary Cilia: What is Known, What is Assumed, and What Needs Clarification

    PubMed Central

    Farnum, Cornelia E.; Wilsman, Norman J.

    2012-01-01

    Two positional characteristics of the ciliary axoneme – its location on the plasma membrane as it emerges from the cell, and its orientation in three-dimensional space – are known to be critical for optimal function of actively motile cilia (including nodal cilia), as well as for modified cilia associated with special senses. However, these positional characteristics have not been analyzed to any significant extent for primary cilia. This review briefly summarizes the history of knowledge of these two positional characteristics across a wide spectrum of cilia, emphasizing their importance for proper function. Then the review focuses what is known about these same positional characteristics for primary cilia in all major tissue types where they have been reported. The review emphasizes major areas that would be productive for future research for understanding how positioning and 3-D orientation of primary cilia may be related to their hypothesized signaling roles within different cellular populations. PMID:22012592

  1. Symmetry Breaking in a Model for Nodal Cilia

    NASA Astrophysics Data System (ADS)

    Brokaw, Charles J.

    2005-03-01

    Nodal cilia are very short cilia found in the embryonic node on the ventral surface of early mammalian embryos. They create a right to left fluid flow that is responsible for determining the normal asymmetry of the internal organs of the mammalian body. To do this, the distal end of the cilium must circle in a counterclockwise sense. Computer simulations with 3-dimensional models of flagella allow examination of 3-dimensional movements such as those of nodal cilia. 3-dimensional circling motions of short cilia can be achieved with velocity controlled models, in which dynein activity is regulated by sliding velocity. If dyneins on one outer doublet are controlled by the sliding velocity experienced by that doublet, the system is symmetric, and the 3-dimensional models can show either clockwise or counterclockwise circling. My computer simulations have examined two possible symmetry breaking mechanisms: 1) dyneins on doublet N are regulated by a mixture of the sliding velocities experienced by doublets N and N+1 (numbered in a clockwise direction, looking from the base). or 2) symmetry is broken by an off-axis force that produces a right-handed twist of the axoneme, consistent with observations that some dyneins can rotate their substrate microtubules in a clockwise direction.

  2. STUDIES ON CILIA

    PubMed Central

    Satir, Peter

    1965-01-01

    Termination of peripheral filaments of the axoneme of gill cilia of fresh-water mussels (Elliptio or Anodonta) occurs in characteristic fashion: (a) subfiber b of certain doublets ends leaving a single simplified tubular unit; (b) the wall of the unit becomes thick and may even obliterate the interior; and (c) the filament drops out of the 9 + 2 pattern. The order in which doublets begin simplifying is also characteristic. This may be determined by numbering the filaments, those with the bridge being 5–6, with the direction of numbering determined by the apparent enantiomorphic configuration (I to IV) of the cross-section. Shorter filaments can be identified in simplifying tips with mixed double and single peripheral units. In this material, laterofrontal cirri show a morphological specialization in the region where individual cilia simplify. The cilia studied run frontally from the body of the cirrus and point in the direction of effective stroke. The longest filaments (Nos. 3, 4, 5, 6, 7) appear as the doublets at the bottom of the cross-section, nearest the surface of the cell of origin. Above them, and above the central pair, a dark band (a section of a dense rod) runs through the matrix. The remaining filaments are the single units. Effective-pointing frontal and lateral ciliary tips end in a fashion similar to laterofrontal tips, although no dense band is present. For all effective-pointing tips studied, the order in which the peripheral filaments end appears to be Nos. (9, 1), 8, 2, 7, 6, 3, 4, 5. However, recovery-pointing lateral tips show a different order: Nos. 7, 6, 8, 5, 9, 4, 1 (3, 2), although the longer filaments are still at the bottom of the cross-section. In simple models of ciliary movement involving contraction of the peripheral filaments, filaments at the top of the cross-section should be longer, if any are. Such models are not supported by the evidence here. These results can be interpreted as supporting sliding-filament models of movement

  3. Hydrodynamic Phase Locking in Mouse Node Cilia

    NASA Astrophysics Data System (ADS)

    Takamatsu, Atsuko; Shinohara, Kyosuke; Ishikawa, Takuji; Hamada, Hiroshi

    2013-06-01

    Rotational movement of mouse node cilia generates leftward fluid flow in the node cavity, playing an important role in left-right determination in the embryo. Although rotation of numerous cilia was believed necessary to trigger the determination, recent reports indicate the action of two cilia to be sufficient. We examine cooperative cilia movement via hydrodynamic interaction. Results show cilia to be cooperative, having phases locked in a certain relation; a system with a pair of nonidentical cilia can achieve phase-locked states more easily than one with a pair of identical cilia.

  4. Piracy of adhesins: attachment of superinfecting pathogens to respiratory cilia by secreted adhesins of Bordetella pertussis.

    PubMed

    Tuomanen, E

    1986-12-01

    Two proteins secreted by Bordetella pertussis are known to mediate adherence of these bacteria to mammalian respiratory cilia. When either ciliated cells or other pathogenic bacteria were pretreated with these adhesins, Streptococcus pneumoniae, Haemophilus influenzae, and Staphylococcus aureus acquired the ability to adhere to cilia in vitro and in vivo. Such piracy of adhesins may contribute to superinfection in mucosal diseases such as whooping cough.

  5. Force Generation and Dynamics of Individual Cilia under External Loading

    PubMed Central

    Hill, David B.; Swaminathan, Vinay; Estes, Ashley; Cribb, Jeremy; O'Brien, E. Timothy; Davis, C. William; Superfine, R.

    2010-01-01

    Abstract Motile cilia are unique multimotor systems that display coordination and periodicity while imparting forces to biological fluids. They play important roles in normal physiology, and ciliopathies are implicated in a growing number of human diseases. In this work we measure the response of individual human airway cilia to calibrated forces transmitted via spot-labeled magnetic microbeads. Cilia respond to applied forces by 1), a reduction in beat amplitude (up to an 85% reduction by 160–170 pN of force); 2), a decreased tip velocity proportionate to applied force; and 3), no significant change in beat frequency. Tip velocity reduction occurred in each beat direction, independently of the direction of applied force, indicating that the cilium is “driven” in both directions at all times. By applying a quasistatic force model, we deduce that axoneme stiffness is dominated by the rigidity of the microtubules, and that cilia can exert 62 ± 18 pN of force at the tip via the generation of 5.6 ± 1.6 pN/dynein head. PMID:20085719

  6. Analysis of properties of cilia using Tetrahymena thermophila.

    PubMed

    Rajagopalan, Vidyalakshmi; Corpuz, Elizabeth O; Hubenschmidt, Mark J; Townsend, Caroline R; Asai, David J; Wilkes, David E

    2009-01-01

    Cilia and eukaryotic flagella are important structures required for the motility of cells, the movement of medium across the surfaces of cells, and the connections between the receptor and synthetic portions of sensory cells. The axoneme forms the cytoskeleton of the cilium comprising several hundreds of proteins that assemble into the 9 + 2 arrangement of outer doublet and central pair microtubules, the inner and outer rows of dynein arms, and many other structures. Tetrahymena thermophila is an excellent model organism for the study of cilia and ciliogenesis. The cell is covered by about 1,000 cilia which are essential for survival. Additionally, the Tetrahymena genome is available and targeted genetic manipulations are straightforward. In this chapter, we describe five protocols that examine properties of cilia: (a) measuring mRNA levels to see the effect of deciliation on gene expression; (b) swimming velocity and linearity; (c) ciliary length and density; (d) phagocytosis that occurs through the ciliated oral apparatus; and (e) depolarization-induced ciliary reversal.

  7. Direct recording and molecular identification of the calcium channel of primary cilia

    NASA Astrophysics Data System (ADS)

    Decaen, Paul G.; Delling, Markus; Vien, Thuy N.; Clapham, David E.

    2013-12-01

    A primary cilium is a solitary, slender, non-motile protuberance of structured microtubules (9+0) enclosed by plasma membrane. Housing components of the cell division apparatus between cell divisions, primary cilia also serve as specialized compartments for calcium signalling and hedgehog signalling pathways. Specialized sensory cilia such as retinal photoreceptors and olfactory cilia use diverse ion channels. An ion current has been measured from primary cilia of kidney cells, but the responsible genes have not been identified. The polycystin proteins (PC and PKD), identified in linkage studies of polycystic kidney disease, are candidate channels divided into two structural classes: 11-transmembrane proteins (PKD1, PKD1L1 and PKD1L2) remarkable for a large extracellular amino terminus of putative cell adhesion domains and a G-protein-coupled receptor proteolytic site, and the 6-transmembrane channel proteins (PKD2, PKD2L1 and PKD2L2; TRPPs). Evidence indicates that the PKD1 proteins associate with the PKD2 proteins via coiled-coil domains. Here we use a transgenic mouse in which only cilia express a fluorophore and use it to record directly from primary cilia, and demonstrate that PKD1L1 and PKD2L1 form ion channels at high densities in several cell types. In conjunction with an accompanying manuscript, we show that the PKD1L1-PKD2L1 heteromeric channel establishes the cilia as a unique calcium compartment within cells that modulates established hedgehog pathways.

  8. Be together, not the same: Spatiotemporal organization of different cilia types generates distinct transport functions

    NASA Astrophysics Data System (ADS)

    Nawroth, Janna; Guo, Hanliang; Ruby, Edward; Dabiri, John; McFall-Ngai, Margaret; Kanso, Eva

    2016-11-01

    Motile cilia are microscopic, hair-like structures on the cell surface that can sense and propel the extracellular fluid environment. Cilia are often thought to be limited to stereotypic morphologies, beat kinematics and non-discriminatory clearance functions, but we find that the spatiotemporal organization of different cilia types and beat behaviors can generate complex flow patterns and transport functions. Here, we present a case study in the Hawaiian bobtail squid where collective ciliary activity and resulting flow fields help recruit symbiont bacteria to the animal host. In particular, we demonstrate empirically and computationally how the squid's internal cilia act like a microfluidic device that actively filters the water for potential bacterial candidates and also provides a sheltered zone allowing for accumulation of mucus and bacteria into a biofilm. Moreover, in this sheltered zone, different cilia-driven flows enhance diffusion of biochemical signals, which could accelerate specific bacteria-host recognition. These results suggest that studying cilia activity on the population level might reveal a diverse range of biological transport and sensing functions. Moreover, understanding cilia as functional building blocks could inspire the design of ciliated robots and devices.

  9. Microfluidic characterization of cilia-driven fluid flow using optical coherence tomography-based particle tracking velocimetry

    PubMed Central

    Jonas, Stephan; Bhattacharya, Dipankan; Khokha, Mustafa K.; Choma, Michael A.

    2011-01-01

    Motile cilia are cellular organelles that generate directional fluid flow across various epithelial surfaces including the embryonic node and respiratory mucosa. The proper functioning of cilia is necessary for normal embryo development and, for the respiratory system, the clearance of mucus and potentially harmful particulate matter. Here we show that optical coherence tomography (OCT) is well-suited for quantitatively characterizing the microfluidic-scale flow generated by motile cilia. Our imaging focuses on the ciliated epithelium of Xenopus tropicalis embryos, a genetically manipulable and experimentally tractable animal model of human disease. We show qualitative flow profile characterization using OCT-based particle pathline imaging. We show quantitative, two-dimensional, two-component flow velocity field characterization using OCT-based particle tracking velocimetry. Quantitative imaging and phenotyping of cilia-driven fluid flow using OCT will enable more detailed research in ciliary biology and in respiratory medicine. PMID:21750777

  10. Deletion of airway cilia results in noninflammatory bronchiectasis and hyperreactive airways

    PubMed Central

    Gilley, Sandra K.; Stenbit, Antine E.; Pasek, Raymond C.; Sas, Kelli M.; Steele, Stacy L.; Amria, May; Bunni, Marlene A.; Estell, Kimberly P.; Schwiebert, Lisa M.; Flume, Patrick; Gooz, Monika; Haycraft, Courtney J.; Yoder, Bradley K.; Miller, Caroline; Pavlik, Jacqueline A.; Turner, Grant A.; Sisson, Joseph H.

    2013-01-01

    The mechanisms for the development of bronchiectasis and airway hyperreactivity have not been fully elucidated. Although genetic, acquired diseases and environmental influences may play a role, it is also possible that motile cilia can influence this disease process. We hypothesized that deletion of a key intraflagellar transport molecule, IFT88, in mature mice causes loss of cilia, resulting in airway remodeling. Airway cilia were deleted by knockout of IFT88, and airway remodeling and pulmonary function were evaluated. In IFT88− mice there was a substantial loss of airway cilia on respiratory epithelium. Three months after the deletion of cilia, there was clear evidence for bronchial remodeling that was not associated with inflammation or apparent defects in mucus clearance. There was evidence for airway epithelial cell hypertrophy and hyperplasia. IFT88− mice exhibited increased airway reactivity to a methacholine challenge and decreased ciliary beat frequency in the few remaining cells that possessed cilia. With deletion of respiratory cilia there was a marked increase in the number of club cells as seen by scanning electron microscopy. We suggest that airway remodeling may be exacerbated by the presence of club cells, since these cells are involved in airway repair. Club cells may be prevented from differentiating into respiratory epithelial cells because of a lack of IFT88 protein that is necessary to form a single nonmotile cilium. This monocilium is a prerequisite for these progenitor cells to transition into respiratory epithelial cells. In conclusion, motile cilia may play an important role in controlling airway structure and function. PMID:24213915

  11. Oscillatory fluid flow influences primary cilia and microtubule mechanics.

    PubMed

    Espinha, Lina C; Hoey, David A; Fernandes, Paulo R; Rodrigues, Hélder C; Jacobs, Christopher R

    2014-07-01

    Many tissues are sensitive to mechanical stimuli; however, the mechanotransduction mechanism used by cells remains unknown in many cases. The primary cilium is a solitary, immotile microtubule-based extension present on nearly every mammalian cell which extends from the basal body. The cilium is a mechanosensitive organelle and has been shown to transduce fluid flow-induced shear stress in tissues, such as the kidney and bone. The majority of microtubules assemble from the mother centriole (basal body), contributing significantly to the anchoring of the primary cilium. Several studies have attempted to quantify the number of microtubules emanating from the basal body and the results vary depending on the cell type. It has also been shown that cellular response to shear stress depends on microtubular integrity. This study hypothesizes that changing the microtubule attachment of primary cilia in response to a mechanical stimulus could change primary cilia mechanics and, possibly, mechanosensitivity. Oscillatory fluid flow was applied to two different cell types and the microtubule attachment to the ciliary base was quantified. For the first time, an increase in microtubules around primary cilia both with time and shear rate in response to oscillatory fluid flow stimulation was demonstrated. Moreover, it is presented that the primary cilium is required for this loading-induced cellular response. This study has demonstrated a new role for the cilium in regulating alterations in the cytoplasmic microtubule network in response to mechanical stimulation, and therefore provides a new insight into how cilia may regulate its mechanics and thus the cells mechanosensitivity.

  12. Pericentrin forms a complex with intraflagellar transport proteins and polycystin-2 and is required for primary cilia assembly

    PubMed Central

    Jurczyk, Agata; Gromley, Adam; Redick, Sambra; Agustin, Jovenal San; Witman, George; Pazour, Gregory J.; Peters, Dorien J.M.; Doxsey, Stephen

    2004-01-01

    Primary cilia are nonmotile microtubule structures that assemble from basal bodies by a process called intraflagellar transport (IFT) and are associated with several human diseases. Here, we show that the centrosome protein pericentrin (Pcnt) colocalizes with IFT proteins to the base of primary and motile cilia. Immunogold electron microscopy demonstrates that Pcnt is on or near basal bodies at the base of cilia. Pcnt depletion by RNA interference disrupts basal body localization of IFT proteins and the cation channel polycystin-2 (PC2), and inhibits primary cilia assembly in human epithelial cells. Conversely, silencing of IFT20 mislocalizes Pcnt from basal bodies and inhibits primary cilia assembly. Pcnt is found in spermatocyte IFT fractions, and IFT proteins are found in isolated centrosome fractions. Pcnt antibodies coimmunoprecipitate IFT proteins and PC2 from several cell lines and tissues. We conclude that Pcnt, IFTs, and PC2 form a complex in vertebrate cells that is required for assembly of primary cilia and possibly motile cilia and flagella. PMID:15337773

  13. Behavior of Primary Cilia and Tricellular Tight Junction Proteins during Differentiation in Temperature-Sensitive Mouse Cochlear Precursor Hair Cells.

    PubMed

    Kakuki, Takuya; Kaneko, Yakuto; Takano, Kenichi; Ninomiya, Takafumi; Kohno, Takayuki; Kojima, Takashi; Himi, Tetsuo

    2016-01-01

    In the sensory hair cells of the mammalian cochlea, the primary cilia in the planar cell polarity as well as the tight junctions in the epithelial cell polarity and the barrier are important to maintain normal hearing. Temperature-sensitive mouse cochlear precursor hair cells were used to investigate the behavior of primary cilia and tricellular tight junction proteins during the differentiation of sensory hair cells. In undifferentiated cells (incubated at 33°C), many acetylated tubulin-positive primary cilia were observed, and each was accompanied with an x03B3;-tubulin-positive basal body. The primary cilia had a '9 + 0' architecture with nine outer microtubule doublets but lacking a central pair of microtubules. In differentiated cells (incubated at 39°C), acetylated tubulin-positive primary cilia as well as acetylated tubulin-positive cilia-like structures were partially observed on the cell surface. In differentiated cells, the number of primary cilia was markedly reduced compared with undifferentiated cells, and innumerable cilia-like structures with no ciliary pockets were partially observed on the cell surface. In undifferentiated cells, few tricellulin molecules and lipolysis-stimulated lipoprotein receptors (LSRs) were observed in the cytoplasm. In differentiated cells, many tricellulin molecules and LSRs were observed on the membranes and within the cytoplasm. Conditional immortalized mouse cochlear precursor hair cells may be useful to investigate the roles of primary cilia and tricellular tight junctions during cellular differentiation and degeneration such as apoptosis.

  14. Fluid pumping using magnetic cilia

    NASA Astrophysics Data System (ADS)

    Hanasoge, Srinivas; Ballard, Matt; Alexeev, Alexander; Hesketh, Peter; Woodruff School of Mechanical Engineering Team

    2016-11-01

    Using experiments and computer simulations, we examine fluid pumping by artificial magnetic cilia fabricated using surface micromachining techniques. An asymmetry in forward and recovery strokes of the elastic cilia causes the net pumping in a creeping flow regime. We show this asymmetry in the ciliary strokes is due to the change in magnetization of the elastic cilia combined with viscous force due to the fluid. Specifically, the time scale for forward stroke is mostly governed by the magnetic forces, whereas the time scale for the recovery stroke is determined by the elastic and viscous forces. These different time scales result in different cilia deformation during forward and backward strokes which in turn lead to the asymmetry in the ciliary motion. To disclose the physics of magnetic cilia pumping we use a hybrid lattice Boltzmann and lattice spring method. We validate our model by comparing the simulation results with the experimental data. The results of our study will be useful to design microfluidic systems for fluid mixing and particle manipulation including different biological particles. USDA and NSF.

  15. hemingway is required for sperm flagella assembly and ciliary motility in Drosophila.

    PubMed

    Soulavie, Fabien; Piepenbrock, David; Thomas, Joëlle; Vieillard, Jennifer; Duteyrat, Jean-Luc; Cortier, Elisabeth; Laurençon, Anne; Göpfert, Martin C; Durand, Bénédicte

    2014-04-01

    Cilia play major functions in physiology and development, and ciliary dysfunctions are responsible for several diseases in humans called ciliopathies. Cilia motility is required for cell and fluid propulsion in organisms. In humans, cilia motility deficiencies lead to primary ciliary dyskinesia, with upper-airways recurrent infections, left-right asymmetry perturbations, and fertility defects. In Drosophila, we identified hemingway (hmw) as a novel component required for motile cilia function. hmw encodes a 604-amino acid protein characterized by a highly conserved coiled-coil domain also found in the human orthologue, KIAA1430. We show that HMW is conserved in species with motile cilia and that, in Drosophila, hmw is expressed in ciliated sensory neurons and spermatozoa. We created hmw-knockout flies and found that they are hearing impaired and male sterile. hmw is implicated in the motility of ciliated auditory sensory neurons and, in the testis, is required for elongation and maintenance of sperm flagella. Because HMW is absent from mature flagella, we propose that HMW is not a structural component of the motile axoneme but is required for proper acquisition of motile properties. This identifies HMW as a novel, evolutionarily conserved component necessary for motile cilium function and flagella assembly.

  16. hemingway is required for sperm flagella assembly and ciliary motility in Drosophila

    PubMed Central

    Soulavie, Fabien; Piepenbrock, David; Thomas, Joëlle; Vieillard, Jennifer; Duteyrat, Jean-Luc; Cortier, Elisabeth; Laurençon, Anne; Göpfert, Martin C.; Durand, Bénédicte

    2014-01-01

    Cilia play major functions in physiology and development, and ciliary dysfunctions are responsible for several diseases in humans called ciliopathies. Cilia motility is required for cell and fluid propulsion in organisms. In humans, cilia motility deficiencies lead to primary ciliary dyskinesia, with upper-airways recurrent infections, left–right asymmetry perturbations, and fertility defects. In Drosophila, we identified hemingway (hmw) as a novel component required for motile cilia function. hmw encodes a 604–amino acid protein characterized by a highly conserved coiled-coil domain also found in the human orthologue, KIAA1430. We show that HMW is conserved in species with motile cilia and that, in Drosophila, hmw is expressed in ciliated sensory neurons and spermatozoa. We created hmw-knockout flies and found that they are hearing impaired and male sterile. hmw is implicated in the motility of ciliated auditory sensory neurons and, in the testis, is required for elongation and maintenance of sperm flagella. Because HMW is absent from mature flagella, we propose that HMW is not a structural component of the motile axoneme but is required for proper acquisition of motile properties. This identifies HMW as a novel, evolutionarily conserved component necessary for motile cilium function and flagella assembly. PMID:24554765

  17. Alcohol drives S-nitrosylation and redox activation of protein phosphatase 1, causing bovine airway cilia dysfunction.

    PubMed

    Price, Michael E; Pavlik, Jacqueline A; Liu, Miao; Ding, Shi-Jian; Wyatt, Todd A; Sisson, Joseph H

    2017-03-01

    Individuals with alcohol (ethanol)-use disorders are at increased risk for lung infections, in part, due to defective mucociliary clearance driven by motile cilia in the airways. We recently reported that isolated, demembranated bovine cilia (axonemes) are capable of producing nitric oxide ((∙)NO) when exposed to biologically relevant concentrations of alcohol. This increased presence of (∙)NO can lead to protein S-nitrosylation, a posttranslational modification signaling mechanism involving reversible adduction of nitrosonium cations or (∙)NO to thiolate or thiyl radicals, respectively, of proteins forming S-nitrosothiols (SNOs). We quantified and compared SNO content between isolated, demembranated axonemes extracted from bovine tracheae, with or without in situ alcohol exposure (100 mM × 24 h). We demonstrate that relevant concentrations of alcohol exposure shift the S-nitrosylation status of key cilia regulatory proteins, including 20-fold increases in S-nitrosylation of proteins that include protein phosphatase 1 (PP1). With the use of an ATP-reactivated axoneme motility system, we demonstrate that alcohol-driven S-nitrosylation of PP1 is associated with PP1 activation and dysfunction of axoneme motility. These new data demonstrate that alcohol can shift the S-nitrothiol balance at the level of the cilia organelle and highlight S-nitrosylation as a novel signaling mechanism to regulate PP1 and cilia motility.

  18. Role of cilia in structural birth defects: insights from ciliopathy mutant mouse models.

    PubMed

    Rao Damerla, Rama; Gabriel, George C; Li, You; Klena, Nikolai T; Liu, Xiaoqin; Chen, Yu; Cui, Cheng; Pazour, Gregory J; Lo, Cecilia W

    2014-06-01

    Structural birth defect (SBD) is a major cause of morbidity and mortality in the newborn period. Although the etiology of SBD is diverse, a wide spectrum of SBD associated with ciliopathies points to the cilium as having a central role in the pathogenesis of SBDs. Ciliopathies are human diseases arising from disruption of cilia structure and/or function. They are associated with developmental anomalies in one or more organ systems and can involve defects in motile cilia, such as those in the airway epithelia or from defects in nonmotile (primary cilia) that have sensory and cell signaling function. Availability of low cost next generation sequencing has allowed for explosion of new knowledge in genetic etiology of ciliopathies. This has led to the appreciation that many genes are shared in common between otherwise clinically distinct ciliopathies. Further insights into the relevance of the cilium in SBD has come from recovery of pathogenic mutations in cilia-related genes from many large-scale mouse forward genetic screens with differing developmental phenotyping focus. Our mouse mutagenesis screen for congenital heart disease (CHD) using noninvasive fetal echocardiography has yielded a marked enrichment for pathogenic mutations in genes required for motile or primary cilia function. These novel mutant mouse models will be invaluable for modeling human ciliopathies and further interrogating the role of the cilium in the pathogenesis of SBD and CHD. Overall, these findings suggest a central role for the cilium in the pathogenesis of a wide spectrum of developmental anomalies associated with CHD and SBDs.

  19. Quantitative description of fluid flows produced by left-right cilia in zebrafish.

    PubMed

    Fox, Craig; Manning, M Lisa; Amack, Jeffrey D

    2015-01-01

    Motile cilia generate directional flows that move mucus through airways, cerebrospinal fluid through brain ventricles, and oocytes through fallopian tubes. In addition, specialized monocilia beat in a rotational pattern to create asymmetric flows that are involved in establishing the left-right (LR) body axis during embryogenesis. These monocilia, which we refer to as "left-right cilia," produce a leftward flow of extraembryonic fluid in a transient "organ of asymmetry" that directs asymmetric signaling and development of LR asymmetries in the cardiovascular system and gastrointestinal tract. The asymmetric flows are thought to establish a chemical gradient and/or activate mechanosensitive cilia to initiate calcium ion signals and a conserved Nodal (TGFβ) pathway on the left side of the embryo, but the mechanisms underlying this process remain unclear. The zebrafish organ of asymmetry, called Kupffer's vesicle, provides a useful model system for investigating LR cilia and cilia-powered fluid flows. Here, we describe methods to visualize flows in Kupffer's vesicle using fluorescent microspheres and introduce a new and freely available MATLAB particle tracking code to quantitatively describe these flows. Analysis of normal and aberrant flows indicates this approach is useful for characterizing flow properties that impact LR asymmetry and may be more broadly applicable for quantifying other cilia flows.

  20. TTC26/DYF13 is an intraflagellar transport protein required for transport of motility-related proteins into flagella.

    PubMed

    Ishikawa, Hiroaki; Ide, Takahiro; Yagi, Toshiki; Jiang, Xue; Hirono, Masafumi; Sasaki, Hiroyuki; Yanagisawa, Haruaki; Wemmer, Kimberly A; Stainier, Didier Yr; Qin, Hongmin; Kamiya, Ritsu; Marshall, Wallace F

    2014-01-01

    Cilia/flagella are assembled and maintained by the process of intraflagellar transport (IFT), a highly conserved mechanism involving more than 20 IFT proteins. However, the functions of individual IFT proteins are mostly unclear. To help address this issue, we focused on a putative IFT protein TTC26/DYF13. Using live imaging and biochemical approaches we show that TTC26/DYF13 is an IFT complex B protein in mammalian cells and Chlamydomonas reinhardtii. Knockdown of TTC26/DYF13 in zebrafish embryos or mutation of TTC26/DYF13 in C. reinhardtii, produced short cilia with abnormal motility. Surprisingly, IFT particle assembly and speed were normal in dyf13 mutant flagella, unlike in other IFT complex B mutants. Proteomic and biochemical analyses indicated a particular set of proteins involved in motility was specifically depleted in the dyf13 mutant. These results support the concept that different IFT proteins are responsible for different cargo subsets, providing a possible explanation for the complexity of the IFT machinery. DOI: http://dx.doi.org/10.7554/eLife.01566.001.

  1. Adipogenic Differentiation of hMSCs is Mediated by Recruitment of IGF-1r Onto the Primary Cilium Associated With Cilia Elongation.

    PubMed

    Dalbay, Melis T; Thorpe, Stephen D; Connelly, John T; Chapple, J Paul; Knight, Martin M

    2015-06-01

    Primary cilia are single non-motile organelles that provide a highly regulated compartment into which specific proteins are trafficked as a critical part of various signaling pathways. The absence of primary cilia has been shown to prevent differentiation of human mesenchymal stem cells (hMSCs). Changes in primary cilia length are crucial for regulating signaling events; however it is not known how alterations in cilia structure relate to differentiation. This study tested the hypothesis that changes in primary cilia structure are required for stem cell differentiation. hMSCs expressed primary cilia that were labeled with acetylated alpha tubulin and visualized by confocal microscopy. Chemically induced differentiation resulted in lineage specific changes in cilia length and prevalence which were independent of cell cycle. In particular, adipogenic differentiation resulted in cilia elongation associated with the presence of dexamethasone, while insulin had an inhibitory effect on cilia length. Over a 7-day time course, adipogenic differentiation media resulted in cilia elongation within 2 days followed by increased nuclear PPARγ levels; an early marker of adipogenesis. Cilia elongation was associated with increased trafficking of insulin-like growth factor-1 receptor β (IGF-1Rβ) into the cilium. This was reversed on inhibition of elongation by IFT-88 siRNA transfection, which also decreased nuclear PPARγ. This is the first study to show that adipogenic differentiation requires primary cilia elongation associated with the recruitment of IGF-1Rβ onto the cilium. This study may lead to the development of cilia-targeted therapies for controlling adipogenic differentiation and associated conditions such as obesity.

  2. Characterization of cancer stem cells and primary cilia in medulloblastoma.

    PubMed

    Gate, David; Danielpour, Moise; Bannykh, Serguei; Town, Terrence

    2015-01-01

    Medulloblastoma, a tumor of the cerebellum, is the most common pediatric central nervous system malignancy. These tumors are etiologically linked to mutations in the Sonic hedgehog (Shh) pathway, which signals through the primary, non-motile cilium. The growth of these aggressive tumors relies on self-renewal of tumor-propagating cells known as cancer stem cells (CSCs). Previous reports have implicated CD133-expressing cells as CSCs in brain tumors, while those expressing CD15 have been shown to propagate medulloblastoma. Here, we demonstrate that CD133+ and CD15+ cells are distinct medulloblastoma populations. CD15+ cells comprise approximately 0.5-1% of total human medulloblastoma cells, display CSC properties in culture and are detected in the Smoothened A1 transgenic mouse model of medulloblastoma. Additionally, we report on a medulloblastoma patient with enriched CD15+ cells in recurrent vs primary medulloblastoma. We also demonstrate that human medulloblastoma cells critically rely on establishment of primary cilia to drive Shh-mediated cell division. Primary cilia are found in external granule cells of human fetal cerebellum and in 12/14 medulloblastoma samples. Yet, CD15+ medulloblastoma cells lack primary cilia, suggesting that this CSC population signals independently of Shh. These results are important when considering the effects of current and prospective treatment modalities on medulloblastoma CSC populations.

  3. Primary cilia in energy balance signaling and metabolic disorder.

    PubMed

    Lee, Hankyu; Song, Jieun; Jung, Joo Hyun; Ko, Hyuk Wan

    2015-12-01

    Energy homeostasis in our body system is maintained by balancing the intake and expenditure of energy. Excessive accumulation of fat by disrupting the balance system causes overweight and obesity, which are increasingly becoming global health concerns. Understanding the pathogenesis of obesity focused on studying the genes related to familial types of obesity. Recently, a rare human genetic disorder, ciliopathy, links the role for genes regulating structure and function of a cellular organelle, the primary cilium, to metabolic disorder, obesity and type II diabetes. Primary cilia are microtubule based hair-like membranous structures, lacking motility and functions such as sensing the environmental cues, and transducing extracellular signals within the cells. Interestingly, the subclass of ciliopathies, such as Bardet-Biedle and Alström syndrome, manifest obesity and type II diabetes in human and mouse model systems. Moreover, studies on genetic mouse model system indicate that more ciliary genes affect energy homeostasis through multiple regulatory steps such as central and peripheral actions of leptin and insulin. In this review, we discuss the latest findings in primary cilia and metabolic disorders, and propose the possible interaction between primary cilia and the leptin and insulin signal pathways which might enhance our understanding of the unambiguous link of a cell's antenna to obesity and type II diabetes.

  4. Primary cilia in energy balance signaling and metabolic disorder

    PubMed Central

    Lee, Hankyu; Song, Jieun; Jung, Joo Hyun; Ko, Hyuk Wan

    2015-01-01

    Energy homeostasis in our body system is maintained by balancing the intake and expenditure of energy. Excessive accumulation of fat by disrupting the balance system causes overweight and obesity, which are increasingly becoming global health concerns. Understanding the pathogenesis of obesity focused on studying the genes related to familial types of obesity. Recently, a rare human genetic disorder, ciliopathy, links the role for genes regulating structure and function of a cellular organelle, the primary cilium, to metabolic disorder, obesity and type II diabetes. Primary cilia are microtubule based hair-like membranous structures, lacking motility and functions such as sensing the environmental cues, and transducing extracellular signals within the cells. Interestingly, the subclass of ciliopathies, such as Bardet-Biedle and Alström syndrome, manifest obesity and type II diabetes in human and mouse model systems. Moreover, studies on genetic mouse model system indicate that more ciliary genes affect energy homeostasis through multiple regulatory steps such as central and peripheral actions of leptin and insulin. In this review, we discuss the latest findings in primary cilia and metabolic disorders, and propose the possible interaction between primary cilia and the leptin and insulin signal pathways which might enhance our understanding of the unambiguous link of a cell’s antenna to obesity and type II diabetes. [BMB Reports 2015; 48(12): 647-654] PMID:26538252

  5. Centrin 2 is required for mouse olfactory ciliary trafficking and development of ependymal cilia planar polarity.

    PubMed

    Ying, Guoxin; Avasthi, Prachee; Irwin, Mavis; Gerstner, Cecilia D; Frederick, Jeanne M; Lucero, Mary T; Baehr, Wolfgang

    2014-04-30

    Centrins are ancient calmodulin-related Ca(2+)-binding proteins associated with basal bodies. In lower eukaryotes, Centrin2 (CETN2) is required for basal body replication and positioning, although its function in mammals is undefined. We generated a germline CETN2 knock-out (KO) mouse presenting with syndromic ciliopathy including dysosmia and hydrocephalus. Absence of CETN2 leads to olfactory cilia loss, impaired ciliary trafficking of olfactory signaling proteins, adenylate cyclase III (ACIII), and cyclic nucleotide-gated (CNG) channel, as well as disrupted basal body apical migration in postnatal olfactory sensory neurons (OSNs). In mutant OSNs, cilia base-anchoring of intraflagellar transport components IFT88, the kinesin-II subunit KIF3A, and cytoplasmic dynein 2 appeared compromised. Although the densities of mutant ependymal and respiratory cilia were largely normal, the planar polarity of mutant ependymal cilia was disrupted, resulting in uncoordinated flow of CSF. Transgenic expression of GFP-CETN2 rescued the Cetn2-deficiency phenotype. These results indicate that mammalian basal body replication and ciliogenesis occur independently of CETN2; however, mouse CETN2 regulates protein trafficking of olfactory cilia and participates in specifying planar polarity of ependymal cilia.

  6. The C. elegans mRNA decapping enzyme shapes morphology of cilia.

    PubMed

    Adachi, Takeshi; Nagahama, Keigo; Izumi, Susumu

    2017-09-05

    Cilia and flagella are evolutionarily conserved organelles that protrude from cell surfaces. Most cilia and flagella are single rod-shaped but some cilia show a variety of shapes. For example, human airway epithelial cells are multiciliated, flagella of crayfish spermatozoon are star-like shaped, and fruit fly spermatozoon extends long flagella. In Caenorhabditis elegans, cilia display morphological diversity of shapes (single, dual rod-type and wing-like and highly-branched shapes). Here we show that DCAP-1 and DCAP-2, which are the homologues of mammalian DCP1 and DCP2 mRNA decapping enzymes, respectively, are involved in formation of dual rod-type and wing-like shaped cilia in C. elegans. mRNA decapping enzyme catalyzes hydrolysis of 5' cap structure of mRNA, which leads to degradation of mRNA. Rescue experiments showed that DCAP-2 acts not in glial cells surrounding cilia but in neurons. This is the first evidence to demonstrate that mRNA decapping is involved in ciliary shape formation. Copyright © 2017. Published by Elsevier Inc.

  7. Centrin 2 Is Required for Mouse Olfactory Ciliary Trafficking and Development of Ependymal Cilia Planar Polarity

    PubMed Central

    Avasthi, Prachee; Irwin, Mavis; Gerstner, Cecilia D.; Frederick, Jeanne M.; Lucero, Mary T.

    2014-01-01

    Centrins are ancient calmodulin-related Ca2+-binding proteins associated with basal bodies. In lower eukaryotes, Centrin2 (CETN2) is required for basal body replication and positioning, although its function in mammals is undefined. We generated a germline CETN2 knock-out (KO) mouse presenting with syndromic ciliopathy including dysosmia and hydrocephalus. Absence of CETN2 leads to olfactory cilia loss, impaired ciliary trafficking of olfactory signaling proteins, adenylate cyclase III (ACIII), and cyclic nucleotide-gated (CNG) channel, as well as disrupted basal body apical migration in postnatal olfactory sensory neurons (OSNs). In mutant OSNs, cilia base-anchoring of intraflagellar transport components IFT88, the kinesin-II subunit KIF3A, and cytoplasmic dynein 2 appeared compromised. Although the densities of mutant ependymal and respiratory cilia were largely normal, the planar polarity of mutant ependymal cilia was disrupted, resulting in uncoordinated flow of CSF. Transgenic expression of GFP-CETN2 rescued the Cetn2-deficiency phenotype. These results indicate that mammalian basal body replication and ciliogenesis occur independently of CETN2; however, mouse CETN2 regulates protein trafficking of olfactory cilia and participates in specifying planar polarity of ependymal cilia. PMID:24790208

  8. 3D structure of eukaryotic flagella/cilia by cryo-electron tomography.

    PubMed

    Ishikawa, Takashi

    2013-01-01

    Flagella/cilia are motile organelles with more than 400 proteins. To understand the mechanism of such complex systems, we need methods to describe molecular arrange-ments and conformations three-dimensionally in vivo. Cryo-electron tomography enabled us such a 3D structural analysis. Our group has been working on 3D structure of flagella/cilia using this method and revealed highly ordered and beautifully organized molecular arrangement. 3D structure gave us insights into the mechanism to gener-ate bending motion with well defined waveforms. In this review, I summarize our recent structural studies on fla-gella/cilia by cryo-electron tomography, mainly focusing on dynein microtubule-based ATPase motor proteins and the radial spoke, a regulatory protein complex.

  9. Primary cilia in neurodevelopmental disorders

    PubMed Central

    Valente, Enza Maria; Rosti, Rasim O.; Gibbs, Elizabeth; Gleeson, Joseph G.

    2014-01-01

    Primary cilia are generally solitary organelles that emanate from the surface of almost all vertebrate cell types. Until recently, details regarding the function of these structures were lacking; however, extensive evidence now suggests that primary cilia have critical roles in sensing the extracellular environment, and in coordinating developmental and homeostatic signalling pathways. Furthermore, disruption of these functions seems to underlie a diverse spectrum of disorders, known as primary ciliopathies. These disorders are characterized by wide-ranging clinical and genetic heterogeneity, but with substantial overlap among distinct conditions. Indeed, ciliopathies are associated with a large variety of manifestations that often include distinctive neurological findings. Herein, we review neurological features associated with primary ciliopathies, highlight genotype–phenotype correlations, and discuss potential mechanisms underlying these findings. PMID:24296655

  10. Oscillatory Fluid Flow Influences Primary Cilia and Microtubule Mechanics

    PubMed Central

    Espinha, Lina C.; Hoey, David A.; Fernandes, Paulo R.; Rodrigues, Hélder C.; Jacobs, Christopher R.

    2014-01-01

    Many tissues are sensitive to mechanical stimuli; however, the mechanotransduction mechanism used by cells remains unknown in many cases. The primary cilium is a solitary, immotile microtubule-based extension present on nearly every mammalian cell which extends from the basal body. The cilium is a mechanosensitive organelle and has been shown to transduce fluid flow-induced shear stress in tissues such as the kidney and bone. The majority of microtubules assemble from the mother centriole (basal body), contributing significantly to the anchoring of the primary cilium. Several studies have attempted to quantify the number of microtubules emanating from the basal body and the results vary depending on the cell type. It has also been shown that cellular response to shear stress depends on microtubular integrity. This study hypothesizes that changing the microtubule attachment of primary cilia in response to a mechanical stimulus could change primary cilia mechanics and, possibly, mechanosensitivity. Oscillatory fluid flow was applied to two different cell types and the microtubule attachment to the ciliary base was quantified. For the first time, an increase in microtubules around primary cilia both with time and shear rate in response to oscillatory fluid flow stimulation was demonstrated. Moreover, it is presented that the primary cilium is required for this loading-induced cellular response. This study has demonstrated a new role for the cilium in regulating alterations in the cytoplasmic microtubule network in response to mechanical stimulation, and therefore provides a new insight into how cilia may regulate its mechanics and thus the cells mechanosensitivity. PMID:25044764

  11. Emergence of metachronal waves in cilia arrays

    PubMed Central

    Elgeti, Jens; Gompper, Gerhard

    2013-01-01

    Propulsion by cilia is a fascinating and universal mechanism in biological organisms to generate fluid motion on the cellular level. Cilia are hair-like organelles, which are found in many different tissues and many uni- and multicellular organisms. Assembled in large fields, cilia beat neither randomly nor completely synchronously—instead they display a striking self-organization in the form of metachronal waves (MCWs). It was speculated early on that hydrodynamic interactions provide the physical mechanism for the synchronization of cilia motion. Theory and simulations of physical model systems, ranging from arrays of highly simplified actuated particles to a few cilia or cilia chains, support this hypothesis. The main questions are how the individual cilia interact with the flow field generated by their neighbors and synchronize their beats for the metachronal wave to emerge and how the properties of the metachronal wave are determined by the geometrical arrangement of the cilia, like cilia spacing and beat direction. Here, we address these issues by large-scale computer simulations of a mesoscopic model of 2D cilia arrays in a 3D fluid medium. We show that hydrodynamic interactions are indeed sufficient to explain the self-organization of MCWs and study beat patterns, stability, energy expenditure, and transport properties. We find that the MCW can increase propulsion velocity more than 3-fold and efficiency almost 10-fold—compared with cilia all beating in phase. This can be a vital advantage for ciliated organisms and may be interesting to guide biological experiments as well as the design of efficient microfluidic devices and artificial microswimmers. PMID:23487771

  12. Emergence of metachronal waves in cilia arrays.

    PubMed

    Elgeti, Jens; Gompper, Gerhard

    2013-03-19

    Propulsion by cilia is a fascinating and universal mechanism in biological organisms to generate fluid motion on the cellular level. Cilia are hair-like organelles, which are found in many different tissues and many uni- and multicellular organisms. Assembled in large fields, cilia beat neither randomly nor completely synchronously--instead they display a striking self-organization in the form of metachronal waves (MCWs). It was speculated early on that hydrodynamic interactions provide the physical mechanism for the synchronization of cilia motion. Theory and simulations of physical model systems, ranging from arrays of highly simplified actuated particles to a few cilia or cilia chains, support this hypothesis. The main questions are how the individual cilia interact with the flow field generated by their neighbors and synchronize their beats for the metachronal wave to emerge and how the properties of the metachronal wave are determined by the geometrical arrangement of the cilia, like cilia spacing and beat direction. Here, we address these issues by large-scale computer simulations of a mesoscopic model of 2D cilia arrays in a 3D fluid medium. We show that hydrodynamic interactions are indeed sufficient to explain the self-organization of MCWs and study beat patterns, stability, energy expenditure, and transport properties. We find that the MCW can increase propulsion velocity more than 3-fold and efficiency almost 10-fold--compared with cilia all beating in phase. This can be a vital advantage for ciliated organisms and may be interesting to guide biological experiments as well as the design of efficient microfluidic devices and artificial microswimmers.

  13. Magnetically Actuated Cilia for Microfluidic Manipulation

    NASA Astrophysics Data System (ADS)

    Hanasoge, Srinivas; Owen, Drew; Ballard, Matt; Hesketh, Peter J.; Alexeev, Alexander; Woodruff School of Mechanical Engineering Collaboration; Petit InstituteBioengineering; Biosciences Collaboration

    2015-11-01

    We demonstrate magnetic micro-cilia based microfluidic mixing and capture techniques. For this, we use a simple and easy to fabricate high aspect ratio cilia, which are actuated magnetically. These micro-features are fabricated by evaporating NiFe alloy at room temperature, on to patterned photoresist. The evaporated alloy curls upwards when the seed layer is removed to release the cilia, thus making a free standing `C' shaped magnetic microstructure. This is actuated using an external electromagnet or a rotating magnet. The artificial cilia can be actuated upto 20Hz. We demonstrate the active mixing these cilia can produce in the microchannel. Also, we demonstrate the capture of target species in a sample using these fast oscillating cilia. The surface of the cilia is functionalized by streptavidin which binds to biotin labelled fluorescent microspheres and mimic the capture of bacteria. We show very high capture efficiencies by using these methods. These simple to fabricate micro cilia can easily be incorporated into many microfluidic systems which require high mixing and capture efficiencies.

  14. Microfluidic manipulation with artificial/bioinspired cilia.

    PubMed

    den Toonder, Jaap M J; Onck, Patrick R

    2013-02-01

    A recent development, inspired by nature, is the use of 'artificial cilia' to create pumping and/or mixing in microfluidic devices. Cilia are small hairs that can be found in biology and are used for (fluid) actuation and sensing. Microscopic actuators resembling cilia, actuated to move under the influence of various stimuli such as electrostatic field, magnetic field, and even light, have been developed by a number of groups and shown to be capable of generating flow and mixing in microfluidic environments. The research on artificial cilia started about a decade ago and is rapidly expanding. In addition to being relevant for potential application in lab-on-a-chip devices, the work on artificial cilia forms a beautiful example of how a biological system can form the successful basis for both scientific research and technological applications. In this review, we will give an overview of the most important approaches in this exciting field. Copyright © 2012 Elsevier Ltd. All rights reserved.

  15. Cilia ultrastructure in children with Down syndrome.

    PubMed

    McLean, Laurie; MacCormick, Johnna; Robb, Ian; Carpenter, Blair; Pothos, Mary

    2003-12-01

    Chronic sinusitis, otitis media with effusion, and upper respiratory tract infections are commonly found in patients with Down syndrome. These diseases are generally felt to be secondary to depressed immune function and altered craniofacial dimensions. Recently, a cilia ultrastructure abnormality was found in a child with Down syndrome. This study is the first to be carried out to determine if cilia ultrastructure abnormalities are prevalent in the population with Down syndrome. Four of 10 patients had documented cilia abnormalities, but these were present in the background of normal cilia, suggesting that they were the result rather than the cause of chronic sinusitis. Similarly, nasal epithelium metaplasia was detected in 50% of the patients. Chronic sinusitis, otitis media with effusion, and recurrent upper respiratory tract infections in children with Down syndrome cannot generally be attributed to primary cilia ultrastructure abnormalities.

  16. Swimming like algae: biomimetic soft artificial cilia

    PubMed Central

    Sareh, Sina; Rossiter, Jonathan; Conn, Andrew; Drescher, Knut; Goldstein, Raymond E.

    2013-01-01

    Cilia are used effectively in a wide variety of biological systems from fluid transport to thrust generation. Here, we present the design and implementation of artificial cilia, based on a biomimetic planar actuator using soft-smart materials. This actuator is modelled on the cilia movement of the alga Volvox, and represents the cilium as a piecewise constant-curvature robotic actuator that enables the subsequent direct translation of natural articulation into a multi-segment ionic polymer metal composite actuator. It is demonstrated how the combination of optimal segmentation pattern and biologically derived per-segment driving signals reproduce natural ciliary motion. The amenability of the artificial cilia to scaling is also demonstrated through the comparison of the Reynolds number achieved with that of natural cilia. PMID:23097503

  17. Swimming like algae: biomimetic soft artificial cilia.

    PubMed

    Sareh, Sina; Rossiter, Jonathan; Conn, Andrew; Drescher, Knut; Goldstein, Raymond

    2013-01-06

    Cilia are used effectively in a wide variety of biological systems from fluid transport to thrust generation. Here, we present the design and implementation of artificial cilia, based on a biomimetic planar actuator using soft-smart materials. This actuator is modelled on the cilia movement of the alga Volvox, and represents the cilium as a piecewise constant-curvature robotic actuator that enables the subsequent direct translation of natural articulation into a multi-segment ionic polymer metal composite actuator. It is demonstrated how the combination of optimal segmentation pattern and biologically derived per-segment driving signals reproduce natural ciliary motion. The amenability of the artificial cilia to scaling is also demonstrated through the comparison of the Reynolds number achieved with that of natural cilia.

  18. Cystogenesis and elongated primary cilia in Tsc1-deficient distal convoluted tubules.

    PubMed

    Armour, Eric A; Carson, Robert P; Ess, Kevin C

    2012-08-15

    Tuberous sclerosis complex (TSC) is a multiorgan hamartomatous disease caused by loss of function mutations of either the TSC1 or TSC2 genes. Neurological symptoms of TSC predominate in younger patients, but renal pathologies are a serious aspect of the disease in older children and adults. To study TSC pathogenesis in the kidney, we inactivated the mouse Tsc1 gene in the distal convoluted tubules (DCT). At young ages, Tsc1 conditional knockout (CKO) mice have enlarged kidneys and mild cystogenesis with increased mammalian target of rapamycin complex (mTORC)1 but decreased mTORC2 signaling. Treatment with the mTORC1 inhibitor rapamycin reduces kidney size and cystogenesis. Rapamycin withdrawal led to massive cystogenesis involving both distal as well as proximal tubules. To assess the contribution of decreased mTORC2 signaling in kidney pathogenesis, we also generated Rictor CKO mice. These animals did not have any detectable kidney pathology. Finally, we examined primary cilia in the DCT. Cilia were longer in Tsc1 CKO mice, and rapamycin treatment returned cilia length to normal. Rictor CKO mice had normal cilia in the DCT. Overall, our findings suggest that loss of the Tsc1 gene in the DCT is sufficient for renal cystogenesis. This cytogenesis appears to be mTORC1 but not mTORC2 dependent. Intriguingly, the mechanism may be cell autonomous as well as non-cell autonomous and possibly involves the length and function of primary cilia.

  19. An Essential Role for Dermal Primary Cilia in Hair Follicle Morphogenesis

    PubMed Central

    Lehman, Jonathan; Laag, Essam; Michaud, Edward J.; Yoder, Bradley K.

    2009-01-01

    The primary cilium is a microtubule-based organelle implicated as an essential component of a number of signaling pathways. It is present on cells throughout the mammalian body; however, its functions in most tissues remain largely unknown. Herein we demonstrate that primary cilia are present on cells in murine skin and hair follicles throughout morphogenesis and during hair follicle cycling in postnatal life. Using the Cre-lox system, we disrupted cilia assembly in the ventral dermis and evaluated the effects on hair follicle development. Mice with disrupted dermal cilia have severe hypotrichosis (lack of hair) in affected areas. Histological analyses reveal that most follicles in the mutants arrest at stage 2 of hair development and have small or absent dermal condensates. This phenotype is reminiscent of that seen in the skin of mice lacking Shh or Gli2. In situ hybridization and quantitative RT-PCR analysis indicates that the hedgehog pathway is downregulated in the dermis of the cilia mutant hair follicles. Thus, these data establish cilia as a critical signaling component required for normal hair morphogenesis and suggest that this organelle is needed on cells in the dermis for reception of signals such as sonic hedgehog. PMID:18987668

  20. Entropy-based measures of in vivo cilia-driven microfluidic mixing derived from quantitative optical imaging

    NASA Astrophysics Data System (ADS)

    Chandrasekera, Kenny; Jonas, Stephan; Bhattacharya, Dipankan; Khokha, Mustafa; Choma, Michael A.

    2012-02-01

    Motile cilia are cellular organelles that project from different epithelial surfaces including respiratory epithelium. They generate directional fluid flow that removes harmful pathogens and particulate matter from the respiratory system. While it has been known that primary ciliary dyskinesia increases the risk of recurrent pulmonary infections, there is now heightened interest in understanding the role that cilia play in a wide-variety of respiratory diseases. Different optical imaging technologies are being investigated to visualize cilia-driven fluid flow, and quantitative image analysis is used to generate measures of ciliary performance. Here, we demonstrate the quantification of in vivo cilia-driven microfluidic mixing using spatial and temporal measures of Shannon information entropy. Using videomicroscopy, we imaged in vivo cilia-driven fluid flow generated by the epidermis of the Xenopus tropicalis embryo. Flow was seeded with either dyes or microparticles. Both spatial and temporal measures of entropy show significant levels of mixing, with maximum entropy measures of ~6.5 (out of a possible range of 0 to 8). Spatial entropy measures showed localization of mixing "hot-spots" and "cold-spots" and temporal measures showed mixing throughout.In sum, entropy-based measures of microfluidic mixing can characterize in vivo cilia-driven fluid flow and hold the potential for better characterization of ciliary dysfunction.

  1. Live Imaging of the Ependymal Cilia in the Lateral Ventricles of the Mouse Brain.

    PubMed

    Al Omran, Alzahra J; Saternos, Hannah C; Liu, Tongyu; Nauli, Surya M; AbouAlaiwi, Wissam A

    2015-06-01

    Multiciliated ependymal cells line the ventricles in the adult brain. Abnormal function or structure of ependymal cilia is associated with various neurological deficits. The current ex vivo live imaging of motile ependymal cilia technique allows for a detailed study of ciliary dynamics following several steps. These steps include: mice euthanasia with carbon dioxide according to protocols of The University of Toledo's Institutional Animal Care and Use Committee (IACUC); craniectomy followed by brain removal and sagittal brain dissection with a vibratome or sharp blade to obtain very thin sections through the brain lateral ventricles, where the ependymal cilia can be visualized. Incubation of the brain's slices in a customized glass-bottom plate containing Dulbecco's Modified Eagle's Medium (DMEM)/High-Glucose at 37 °C in the presence of 95%/5% O2/CO2 mixture is essential to keep the tissue alive during the experiment. A video of the cilia beating is then recorded using a high-resolution differential interference contrast microscope. The video is then analyzed frame by frame to calculate the ciliary beating frequency. This allows distinct classification of the ependymal cells into three categories or types based on their ciliary beating frequency and angle. Furthermore, this technique allows the use of high-speed fluorescence imaging analysis to characterize the unique intracellular calcium oscillation properties of ependymal cells as well as the effect of pharmacological agents on the calcium oscillations and the ciliary beating frequency. In addition, this technique is suitable for immunofluorescence imaging for ciliary structure and ciliary protein localization studies. This is particularly important in disease diagnosis and phenotype studies. The main limitation of the technique is attributed to the decrease in live motile cilia movement as the brain tissue starts to die.

  2. Swimming with protists: perception, motility and flagellum assembly.

    PubMed

    Ginger, Michael L; Portman, Neil; McKean, Paul G

    2008-11-01

    In unicellular and multicellular eukaryotes, fast cell motility and rapid movement of material over cell surfaces are often mediated by ciliary or flagellar beating. The conserved defining structure in most motile cilia and flagella is the '9+2' microtubule axoneme. Our general understanding of flagellum assembly and the regulation of flagellar motility has been led by results from seminal studies of flagellate protozoa and algae. Here we review recent work relating to various aspects of protist physiology and cell biology. In particular, we discuss energy metabolism in eukaryotic flagella, modifications to the canonical assembly pathway and flagellum function in parasite virulence.

  3. Interactive computer-assisted approach for evaluation of ultrastructural cilia abnormalities

    NASA Astrophysics Data System (ADS)

    Palm, Christoph; Siegmund, Heiko; Semmelmann, Matthias; Grafe, Claudia; Evert, Matthias; Schroeder, Josef A.

    2016-03-01

    Introduction - Diagnosis of abnormal cilia function is based on ultrastructural analysis of axoneme defects, especialy the features of inner and outer dynein arms which are the motors of ciliar motility. Sub-optimal biopsy material, methodical, and intrinsic electron microscopy factors pose difficulty in ciliary defects evaluation. We present a computer-assisted approach based on state-of-the-art image analysis and object recognition methods yielding a time-saving and efficient diagnosis of cilia dysfunction. Method - The presented approach is based on a pipeline of basal image processing methods like smoothing, thresholding and ellipse fitting. However, integration of application specific knowledge results in robust segmentations even in cases of image artifacts. The method is build hierarchically starting with the detection of cilia within the image, followed by the detection of nine doublets within each analyzable cilium, and ending with the detection of dynein arms of each doublet. The process is concluded by a rough classification of the dynein arms as basis for a computer-assisted diagnosis. Additionally, the interaction possibilities are designed in a way, that the results are still reproducible given the completion report. Results - A qualitative evaluation showed reasonable detection results for cilia, doublets and dynein arms. However, since a ground truth is missing, the variation of the computer-assisted diagnosis should be within the subjective bias of human diagnosticians. The results of a first quantitative evaluation with five human experts and six images with 12 analyzable cilia showed, that with default parameterization 91.6% of the cilia and 98% of the doublets were found. The computer-assisted approach rated 66% of those inner and outer dynein arms correct, where all human experts agree. However, especially the quality of the dynein arm classification may be improved in future work.

  4. Out of the cleanroom, self-assembled magnetic artificial cilia.

    PubMed

    Wang, Ye; Gao, Yang; Wyss, Hans; Anderson, Patrick; den Toonder, Jaap

    2013-09-07

    Micro-sized hair-like structures, such as cilia, are abundant in nature and have various functionalities. Many efforts have been made to mimic the fluid pumping function of cilia, but most of the fabrication processes for these "artificial cilia" are tedious and expensive, hindering their practical application. In this paper a cost-effective in situ fabrication technique for artificial cilia is demonstrated. The cilia are constructed by self-assembly of micron sized magnetic beads and encapsulated with soft polymer coatings. Actuation of the cilia induces an effective fluid flow, and the cilia lengths and distribution can be adjusted by varying the magnetic bead concentration and fabrication parameters.

  5. Primary Cilia in Pancreatic Development and Disease

    PubMed Central

    Lodh, Sukanya; O’Hare, Elizabeth A.; Zaghloul, Norann A.

    2014-01-01

    Primary cilia and their anchoring basal bodies are important regulators of a growing list of signaling pathways. Consequently, dysfunction in proteins associated with these structures results in perturbation of the development and function of a spectrum of tissue and cell types. Here, we review the role of cilia in mediating the development and function of the pancreas. We focus on ciliary regulation of major pathways involved in pancreatic development, including Shh, Wnt, TGF-β, Notch, and fibroblast growth factor. We also discuss pancreatic phenotypes associated with ciliary dysfunction, including pancreatic cysts and defects in glucose homeostasis, and explore the potential role of cilia in such defects. PMID:24864023

  6. HEATR2 plays a conserved role in assembly of the ciliary motile apparatus.

    PubMed

    Diggle, Christine P; Moore, Daniel J; Mali, Girish; zur Lage, Petra; Ait-Lounis, Aouatef; Schmidts, Miriam; Shoemark, Amelia; Garcia Munoz, Amaya; Halachev, Mihail R; Gautier, Philippe; Yeyati, Patricia L; Bonthron, David T; Carr, Ian M; Hayward, Bruce; Markham, Alexander F; Hope, Jilly E; von Kriegsheim, Alex; Mitchison, Hannah M; Jackson, Ian J; Durand, Bénédicte; Reith, Walter; Sheridan, Eamonn; Jarman, Andrew P; Mill, Pleasantine

    2014-09-01

    Cilia are highly conserved microtubule-based structures that perform a variety of sensory and motility functions during development and adult homeostasis. In humans, defects specifically affecting motile cilia lead to chronic airway infections, infertility and laterality defects in the genetically heterogeneous disorder Primary Ciliary Dyskinesia (PCD). Using the comparatively simple Drosophila system, in which mechanosensory neurons possess modified motile cilia, we employed a recently elucidated cilia transcriptional RFX-FOX code to identify novel PCD candidate genes. Here, we report characterization of CG31320/HEATR2, which plays a conserved critical role in forming the axonemal dynein arms required for ciliary motility in both flies and humans. Inner and outer arm dyneins are absent from axonemes of CG31320 mutant flies and from PCD individuals with a novel splice-acceptor HEATR2 mutation. Functional conservation of closely arranged RFX-FOX binding sites upstream of HEATR2 orthologues may drive higher cytoplasmic expression of HEATR2 during early motile ciliogenesis. Immunoprecipitation reveals HEATR2 interacts with DNAI2, but not HSP70 or HSP90, distinguishing it from the client/chaperone functions described for other cytoplasmic proteins required for dynein arm assembly such as DNAAF1-4. These data implicate CG31320/HEATR2 in a growing intracellular pre-assembly and transport network that is necessary to deliver functional dynein machinery to the ciliary compartment for integration into the motile axoneme.

  7. Primary Cilia on Horizontal Basal Cells Regulate Regeneration of the Olfactory Epithelium.

    PubMed

    Joiner, Ariell M; Green, Warren W; McIntyre, Jeremy C; Allen, Benjamin L; Schwob, James E; Martens, Jeffrey R

    2015-10-07

    The olfactory epithelium (OE) is one of the few tissues to undergo constitutive neurogenesis throughout the mammalian lifespan. It is composed of multiple cell types including olfactory sensory neurons (OSNs) that are readily replaced by two populations of basal stem cells, frequently dividing globose basal cells and quiescent horizontal basal cells (HBCs). However, the precise mechanisms by which these cells mediate OE regeneration are unclear. Here, we show for the first time that the HBC subpopulation of basal stem cells uniquely possesses primary cilia that are aligned in an apical orientation in direct apposition to sustentacular cell end feet. The positioning of these cilia suggests that they function in the detection of growth signals and/or differentiation cues. To test this idea, we generated an inducible, cell type-specific Ift88 knock-out mouse line (K5rtTA;tetOCre;Ift88(fl/fl)) to disrupt cilia formation and maintenance specifically in HBCs. Surprisingly, the loss of HBC cilia did not affect the maintenance of the adult OE but dramatically impaired the regeneration of OSNs following lesion. Furthermore, the loss of cilia during development resulted in a region-specific decrease in neurogenesis, implicating HBCs in the establishment of the OE. Together, these results suggest a novel role for primary cilia in HBC activation, proliferation, and differentiation. We show for the first time the presence of primary cilia on a quiescent population of basal stem cells, the horizontal basal cells (HBCs), in the olfactory epithelium (OE). Importantly, our data demonstrate that cilia on HBCs are necessary for regeneration of the OE following injury. Moreover, the disruption of HBC cilia alters neurogenesis during the development of the OE, providing evidence that HBCs participate in the establishment of this tissue. These data suggest that the mechanisms of penetrance for ciliopathies in the OE extend beyond that of defects in olfactory sensory neurons and may

  8. Identification of ICIS-1, a new protein involved in cilia stability.

    PubMed

    Ponsard, Cecile; Skowron-Zwarg, Marie; Seltzer, Virginie; Perret, Eric; Gallinger, Julia; Fisch, Cathy; Dupuis-Williams, Pascale; Caruso, Nathalie; Middendorp, Sandrine; Tournier, Frederic

    2007-01-01

    Cilia are specialized organelles that exert critical functions in numerous organisms, including that of cell motility, fluid transport and protozoan locomotion. Ciliary architecture and function strictly depend on basal body formation, migration and axoneme elongation. Numerous ultrastructural studies have been undertaken in different species to elucidate the process of ciliogenesis. Recent analyses have led to identification of genes specifically expressed in ciliated organisms, but most proteins involved in ciliogenesis remain uncharacterized. Using human nasal epithelial cells capable of ciliary differentiation in vitro, differential display was carried out to identify new proteins associated with ciliogenesis. We isolated a new gene, ICIS-1 (Involved in CIlia Stability-1), upregulated during mucociliary differentiation. This gene is localized within the TGF-beta1 promoter and is ubiquitously expressed in human tissues. Functional analyses of gene expression inhibition by RNA interference in Paramecium tetraurelia indicated that the ICIS-1 homologue interfered with cilia stability or formation. These findings demonstrate that ICIS-1 is a new protein associated with ciliated cells and potentially related to cilia stability.

  9. Primary Cilia in Breast Cancer Progression

    DTIC Science & Technology

    2010-06-01

    mechanical signals (Satir and Christensen, 2007). The process of Intraflagellar Transport ( IFT ) is responsible for building and maintaining the...Zhang, Q., Song, B., Jackson, W.S., Detloff, P.J., Serra, R., and Yoder, B.K. (2007). Intraflagellar transport is essential for endochondral bone...structure and function of cilia. The absence of Ift88/Tg737/Polaris, a core molecular component of the IFT machinery, results in the loss of cilia

  10. Cilia in vertebrate left-right patterning.

    PubMed

    Dasgupta, Agnik; Amack, Jeffrey D

    2016-12-19

    Understanding how left-right (LR) asymmetry is generated in vertebrate embryos is an important problem in developmental biology. In humans, a failure to align the left and right sides of cardiovascular and/or gastrointestinal systems often results in birth defects. Evidence from patients and animal models has implicated cilia in the process of left-right patterning. Here, we review the proposed functions for cilia in establishing LR asymmetry, which include creating transient leftward fluid flows in an embryonic 'left-right organizer'. These flows direct asymmetric activation of a conserved Nodal (TGFβ) signalling pathway that guides asymmetric morphogenesis of developing organs. We discuss the leading hypotheses for how cilia-generated asymmetric fluid flows are translated into asymmetric molecular signals. We also discuss emerging mechanisms that control the subcellular positioning of cilia and the cellular architecture of the left-right organizer, both of which are critical for effective cilia function during left-right patterning. Finally, using mosaic cell-labelling and time-lapse imaging in the zebrafish embryo, we provide new evidence that precursor cells maintain their relative positions as they give rise to the ciliated left-right organizer. This suggests the possibility that these cells acquire left-right positional information prior to the appearance of cilia.This article is part of the themed issue 'Provocative questions in left-right asymmetry'. © 2016 The Author(s).

  11. Mutation of the MAP kinase DYF-5 affects docking and undocking of kinesin-2 motors and reduces their speed in the cilia of Caenorhabditis elegans

    PubMed Central

    Burghoorn, Jan; Dekkers, Martijn P. J.; Rademakers, Suzanne; de Jong, Ton; Willemsen, Rob; Jansen, Gert

    2007-01-01

    In the cilia of the nematode Caenorhabditis elegans, anterograde intraflagellar transport (IFT) is mediated by two kinesin-2 complexes, kinesin II and OSM-3 kinesin. These complexes function together in the cilia middle segments, whereas OSM-3 alone mediates transport in the distal segments. Not much is known about the mechanisms that compartmentalize the kinesin-2 complexes or how transport by both kinesins is coordinated. Here, we identify DYF-5, a conserved MAP kinase that plays a role in these processes. Fluorescence microscopy and EM revealed that the cilia of dyf-5 loss-of-function (lf) animals are elongated and are not properly aligned into the amphid channel. Some cilia do enter the amphid channel, but the distal ends of these cilia show accumulation of proteins. Consistent with these observations, we found that six IFT proteins accumulate in the cilia of dyf-5(lf) mutants. In addition, using genetic analyses and live imaging to measure the motility of IFT proteins, we show that dyf-5 is required to restrict kinesin II to the cilia middle segments. Finally, we show that, in dyf-5(lf) mutants, OSM-3 moves at a reduced speed and is not attached to IFT particles. We propose that DYF-5 plays a role in the undocking of kinesin II from IFT particles and in the docking of OSM-3 onto IFT particles. PMID:17420466

  12. Absence of Radial Spokes in Mouse Node Cilia Is Required for Rotational Movement but Confers Ultrastructural Instability as a Trade-Off.

    PubMed

    Shinohara, Kyosuke; Chen, Duanduan; Nishida, Tomoki; Misaki, Kazuyo; Yonemura, Shigenobu; Hamada, Hiroshi

    2015-10-26

    Determination of left-right asymmetry in mouse embryos is established by a leftward fluid flow that is generated by clockwise rotation of node cilia. How node cilia achieve stable unidirectional rotation has remained unknown, however. Here we show that brief exposure to the microtubule-stabilizing drug paclitaxel (Taxol) induces randomly directed rotation and changes the ultrastructure of node cilia. In vivo observations and a computer simulation revealed that a regular 9+0 arrangement of doublet microtubules is essential for stable unidirectional rotation of node cilia. The 9+2 motile cilia of the airway, which manifest planar beating, are resistant to Taxol treatment. However, the airway cilia of mice lacking the radial spoke head protein Rsph4a undergo rotational movement instead of planar beating, are prone to microtubule rearrangement, and are sensitive to Taxol. Our results suggest that the absence of radial spokes allows node cilia to rotate unidirectionally but, as a trade-off, renders them ultrastructurally fragile. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Ultrastructural characterization of primary cilia in pathologically characterized human glioblastoma multiforme (GBM) tumors.

    PubMed

    Moser, Joanna J; Fritzler, Marvin J; Rattner, Jerome B

    2014-01-01

    Primary cilia are non-motile sensory cytoplasmic organelles that are involved in cell cycle progression. Ultrastructurally, the primary cilium region is complex, with normal ciliogenesis progressing through five distinct morphological stages in human astrocytes. Defects in early stages of ciliogenesis are key features of astrocytoma/glioblastoma cell lines and provided the impetus for the current study which describes the morphology of primary cilia in molecularly characterized human glioblastoma multiforme (GBM) tumors. Seven surgically resected human GBM tissue samples were molecularly characterized according to IDH1/2 mutation status, EGFR amplification status and MGMT promoter methylation status and were examined for primary cilia expression and structure using indirect immunofluorescence and electron microscopy. We report for the first time that primary cilia are disrupted in the early stages of ciliogenesis in human GBM tumors. We confirm that immature primary cilia and basal bodies/centrioles have aberrant ciliogenesis characteristics including absent paired vesicles, misshaped/swollen vesicular hats, abnormal configuration of distal appendages, and discontinuity of centriole microtubular blades. Additionally, the transition zone plate is able to form in the absence of paired vesicles on the distal end of the basal body and when a cilium progresses beyond the early stages of ciliogenesis, it has electron dense material clumped along the transition zone and a darkening of the microtubules at the proximal end of the cilium. Primary cilia play a role in a variety of human cancers. Previously primary cilia structure was perturbed in cultured cell lines derived from astrocytomas/glioblastomas; however there was always some question as to whether these findings were a cell culture phenomena. In this study we confirm that disruptions in ciliogenesis at early stages do occur in GBM tumors and that these ultrastructural findings bear resemblance to those previously

  14. Learn About GI Motility

    MedlinePlus

    ... Disorders of the Large Intestine Disorders of the Pelvic Floor Motility Testing Personal Stories Contact About GI Motility ... Disorders of the Large Intestine Disorders of the Pelvic Floor Motility Testing Personal Stories Contact About GI Motility ...

  15. About GI Motility

    MedlinePlus

    ... Disorders of the Large Intestine Disorders of the Pelvic Floor Motility Testing Personal Stories Contact About GI Motility ... Disorders of the Large Intestine Disorders of the Pelvic Floor Motility Testing Personal Stories Contact About GI Motility ...

  16. Primary cilia signaling mediates intraocular pressure sensation.

    PubMed

    Luo, Na; Conwell, Michael D; Chen, Xingjuan; Kettenhofen, Christine Insinna; Westlake, Christopher J; Cantor, Louis B; Wells, Clark D; Weinreb, Robert N; Corson, Timothy W; Spandau, Dan F; Joos, Karen M; Iomini, Carlo; Obukhov, Alexander G; Sun, Yang

    2014-09-02

    Lowe syndrome is a rare X-linked congenital disease that presents with congenital cataracts and glaucoma, as well as renal and cerebral dysfunction. OCRL, an inositol polyphosphate 5-phosphatase, is mutated in Lowe syndrome. We previously showed that OCRL is involved in vesicular trafficking to the primary cilium. Primary cilia are sensory organelles on the surface of eukaryotic cells that mediate mechanotransduction in the kidney, brain, and bone. However, their potential role in the trabecular meshwork (TM) in the eye, which regulates intraocular pressure, is unknown. Here, we show that TM cells, which are defective in glaucoma, have primary cilia that are critical for response to pressure changes. Primary cilia in TM cells shorten in response to fluid flow and elevated hydrostatic pressure, and promote increased transcription of TNF-α, TGF-β, and GLI1 genes. Furthermore, OCRL is found to be required for primary cilia to respond to pressure stimulation. The interaction of OCRL with transient receptor potential vanilloid 4 (TRPV4), a ciliary mechanosensory channel, suggests that OCRL may act through regulation of this channel. A novel disease-causing OCRL allele prevents TRPV4-mediated calcium signaling. In addition, TRPV4 agonist GSK 1016790A treatment reduced intraocular pressure in mice; TRPV4 knockout animals exhibited elevated intraocular pressure and shortened cilia. Thus, mechanotransduction by primary cilia in TM cells is implicated in how the eye senses pressure changes and highlights OCRL and TRPV4 as attractive therapeutic targets for the treatment of glaucoma. Implications of OCRL and TRPV4 in primary cilia function may also shed light on mechanosensation in other organ systems.

  17. Primary cilia signaling mediates intraocular pressure sensation

    PubMed Central

    Luo, Na; Conwell, Michael D.; Chen, Xingjuan; Kettenhofen, Christine Insinna; Westlake, Christopher J.; Cantor, Louis B.; Wells, Clark D.; Weinreb, Robert N.; Corson, Timothy W.; Spandau, Dan F.; Joos, Karen M.; Iomini, Carlo; Obukhov, Alexander G.; Sun, Yang

    2014-01-01

    Lowe syndrome is a rare X-linked congenital disease that presents with congenital cataracts and glaucoma, as well as renal and cerebral dysfunction. OCRL, an inositol polyphosphate 5-phosphatase, is mutated in Lowe syndrome. We previously showed that OCRL is involved in vesicular trafficking to the primary cilium. Primary cilia are sensory organelles on the surface of eukaryotic cells that mediate mechanotransduction in the kidney, brain, and bone. However, their potential role in the trabecular meshwork (TM) in the eye, which regulates intraocular pressure, is unknown. Here, we show that TM cells, which are defective in glaucoma, have primary cilia that are critical for response to pressure changes. Primary cilia in TM cells shorten in response to fluid flow and elevated hydrostatic pressure, and promote increased transcription of TNF-α, TGF-β, and GLI1 genes. Furthermore, OCRL is found to be required for primary cilia to respond to pressure stimulation. The interaction of OCRL with transient receptor potential vanilloid 4 (TRPV4), a ciliary mechanosensory channel, suggests that OCRL may act through regulation of this channel. A novel disease-causing OCRL allele prevents TRPV4-mediated calcium signaling. In addition, TRPV4 agonist GSK 1016790A treatment reduced intraocular pressure in mice; TRPV4 knockout animals exhibited elevated intraocular pressure and shortened cilia. Thus, mechanotransduction by primary cilia in TM cells is implicated in how the eye senses pressure changes and highlights OCRL and TRPV4 as attractive therapeutic targets for the treatment of glaucoma. Implications of OCRL and TRPV4 in primary cilia function may also shed light on mechanosensation in other organ systems. PMID:25143588

  18. Cilia in the nervous system: linking cilia function and neurodevelopmental disorders

    PubMed Central

    Lee, Ji E.; Gleeson, Joseph G.

    2014-01-01

    Purpose of review Ciliopathies are genetic disorders caused by defects of primary ciliary structure and/or function and are characterized by pleiotropic clinical features. The ciliopathies include several partially overlapping syndromes such as Joubert syndrome, Bardet–Biedl syndrome and Meckel–Gruber syndrome, all of which have pronounced neurodevelopmental features. Here we focus on potential roles of cilia in central nervous system function, to explore how impairments may cause brain malformation and neurodevelopmental disease. Recent findings Cilia have long been considered as ‘sensory cellular antennae’, responding as chemo-sensors, mechano-sensors and thermo-sensors, although their roles in development were not well understood until recently. The surprising finding that disparate syndromes are all due to defects of the primary cilia, along with the recent advances in genetics, has helped elucidate further roles of primary cilia beyond sensory functions. Several molecules that are associated with key signaling pathways have been discovered in primary cilia. These include sonic hedgehog, wingless, planar cell polarity and fibroblast growth factor, which are essential for many cellular processes. Additionally, mutations in ‘ciliome’ genes have largely shown developmental defects such as abnormal body axis and brain malformation, implying disrupted cilia-related signaling pathways. Accordingly, the emerging theme is that primary cilia may play roles as modulators of signal transduction to help shape cellular responses within the environmental context during both development and homeostasis. Summary The link between cilia and signal pathways has become a framework for understanding the pathogenesis of ciliopathies. Despite recent progress in ciliary biology, fundamental questions remain about how cilia regulate neuronal function in the central nervous system. Therefore, investigation of ciliary function in the nervous system may reveal cilia

  19. Cilia in the nervous system: linking cilia function and neurodevelopmental disorders.

    PubMed

    Lee, Ji E; Gleeson, Joseph G

    2011-04-01

    Ciliopathies are genetic disorders caused by defects of primary ciliary structure and/or function and are characterized by pleiotropic clinical features. The ciliopathies include several partially overlapping syndromes such as Joubert syndrome, Bardet-Biedl syndrome and Meckel-Gruber syndrome, all of which have pronounced neurodevelopmental features. Here we focus on potential roles of cilia in central nervous system function, to explore how impairments may cause brain malformation and neurodevelopmental disease. Cilia have long been considered as 'sensory cellular antennae', responding as chemo-sensors, mechano-sensors and thermo-sensors, although their roles in development were not well understood until recently. The surprising finding that disparate syndromes are all due to defects of the primary cilia, along with the recent advances in genetics, has helped elucidate further roles of primary cilia beyond sensory functions. Several molecules that are associated with key signaling pathways have been discovered in primary cilia. These include sonic hedgehog, wingless, planar cell polarity and fibroblast growth factor, which are essential for many cellular processes. Additionally, mutations in 'ciliome' genes have largely shown developmental defects such as abnormal body axis and brain malformation, implying disrupted cilia-related signaling pathways. Accordingly, the emerging theme is that primary cilia may play roles as modulators of signal transduction to help shape cellular responses within the environmental context during both development and homeostasis. The link between cilia and signal pathways has become a framework for understanding the pathogenesis of ciliopathies. Despite recent progress in ciliary biology, fundamental questions remain about how cilia regulate neuronal function in the central nervous system. Therefore, investigation of ciliary function in the nervous system may reveal cilia-modulating mechanisms in neurodevelopmental processes, as well

  20. Expression and localization of the Parkin co-regulated gene in mouse CNS suggests a role in ependymal cilia function.

    PubMed

    Wilson, Gabrielle R; Tan, Jacqueline T; Brody, Kate M; Taylor, Juliet M; Delatycki, Martin B; Lockhart, Paul J

    2009-08-21

    Parkin Co-Regulated Gene (PACRG) is a gene that shares a bi-directional promoter with the Parkinson's disease associated gene parkin. The functional role of PACRG is not well understood, although the gene has been associated with parkinsonian syndromes and more recently with eukaryotic cilia and flagella. We investigated the expression of Pacrg in the mouse brain by in situ hybridization and observed robust expression of Pacrg in the cells associated with the lateral, third and fourth ventricle, in addition to the aqueduct of Sylvius and choroid plexus. For all regions of Pacrg expression identified, strong expression was observed in the newborn period and this was maintained into adulthood. Immunohistochemical analysis showed that Pacrg was a component of the ependymal cells and cilia lining the ventricles. Based on our results and the previous association of PACRG homologues with cilia and flagella, we propose that Pacrg is a component of the ependymal cilia and may play an important role in motile cilia development and/or function in the CNS.

  1. Kinesin-II is preferentially targeted to assembling cilia and is required for ciliogenesis and normal cytokinesis in Tetrahymena.

    PubMed

    Brown, J M; Marsala, C; Kosoy, R; Gaertig, J

    1999-10-01

    We cloned two genes, KIN1 and KIN2, encoding kinesin-II homologues from the ciliate Tetrahymena thermophila and constructed strains lacking either KIN1 or KIN2 or both genes. Cells with a single disruption of either gene showed partly overlapping sets of defects in cell growth, motility, ciliary assembly, and thermoresistance. Deletion of both genes resulted in loss of cilia and arrests in cytokinesis. Mutant cells were unable to assemble new cilia or to maintain preexisting cilia. Double knockout cells were not viable on a standard medium but could be grown on a modified medium on which growth does not depend on phagocytosis. Double knockout cells could be rescued by transformation with a gene encoding an epitope-tagged Kin1p. In growing cells, epitope-tagged Kin1p preferentially accumulated in cilia undergoing active assembly. Kin1p was also detected in the cell body but did not show any association with the cleavage furrow. The cell division arrests observed in kinesin-II knockout cells appear to be induced by the loss of cilia and resulting cell paralysis.

  2. Analysis of Soluble Protein Entry into Primary Cilia Using Semi-Permeabilized Cells

    PubMed Central

    Breslow, David K.; Nachury, Maxence V.

    2016-01-01

    The primary cilium is a protrusion from the cell surface that serves as a specialized compartment for signal transduction. Many signaling factors are known to be dynamically concentrated within cilia and to require cilia for their function. Yet protein entry into primary cilia remains poorly understood. To enable a mechanistic analysis of soluble protein entry into cilia, we developed a method for semi-permeabilization of mammalian cells in which the plasma membrane is permeabilized while the ciliary membrane remains intact. Using semi-permeabilized cells as the basis for an in vitro diffusion-to-capture assay, we uncovered a size-dependent diffusion barrier that restricts soluble protein exchange between the cytosol and the cilium. The manipulability of this in vitro system enabled an extensive characterization of the ciliary diffusion barrier and led us to show that the barrier is mechanistically distinct from those at the axon initial segment and the nuclear pore complex. Because semi-permeabilized cells enable a range of experimental perturbations that would not be easily feasible in intact cells, we believe this methodology will provide a unique resource for investigating primary cilium function in development and disease. PMID:25837393

  3. C2cd3 is required for cilia formation and Hedgehog signaling in mouse.

    PubMed

    Hoover, Amber N; Wynkoop, Aaron; Zeng, Huiqing; Jia, Jinping; Niswander, Lee A; Liu, Aimin

    2008-12-01

    Cilia are essential for mammalian embryonic development as well as for the physiological activity of various adult organ systems. Despite the multiple crucial roles that cilia play, the mechanisms underlying ciliogenesis in mammals remain poorly understood. Taking a forward genetic approach, we have identified Hearty (Hty), a recessive lethal mouse mutant with multiple defects, including neural tube defects, abnormal dorsal-ventral patterning of the spinal cord, a defect in left-right axis determination and severe polydactyly (extra digits). By genetic mapping, sequence analysis of candidate genes and characterization of a second mutant allele, we identify Hty as C2cd3, a novel gene encoding a vertebrate-specific C2 domain-containing protein. Target gene expression and double-mutant analyses suggest that C2cd3 is an essential regulator of intracellular transduction of the Hedgehog signal. Furthering a link between Hedgehog signaling and cilia function, we find that cilia formation and proteolytic processing of Gli3 are disrupted in C2cd3 mutants. Finally, we observe C2cd3 protein at the basal body, consistent with its essential function in ciliogenesis. Interestingly, the human ortholog for this gene lies in proximity to the critical regions of Meckel-Gruber syndrome 2 (MKS2) and Joubert syndrome 2 (JBTS2), making it a potential candidate for these two human genetic disorders.

  4. Mouse infection and pathogenesis by Trypanosoma brucei motility mutants.

    PubMed

    Kisalu, Neville K; Langousis, Gerasimos; Bentolila, Laurent A; Ralston, Katherine S; Hill, Kent L

    2014-06-01

    The flagellum of Trypanosoma brucei is an essential and multifunctional organelle that drives parasite motility and is receiving increased attention as a potential drug target. In the mammalian host, parasite motility is suspected to contribute to infection and disease pathogenesis. However, it has not been possible to test this hypothesis owing to lack of motility mutants that are viable in the bloodstream life cycle stage that infects the mammalian host. We recently identified a bloodstream-form motility mutant in 427-derived T. brucei in which point mutations in the LC1 dynein subunit disrupt propulsive motility but do not affect viability. These mutants have an actively beating flagellum, but cannot translocate. Here we demonstrate that the LC1 point mutant fails to show enhanced cell motility upon increasing viscosity of the surrounding medium, which is a hallmark of wild type T. brucei, thus indicating that motility of the mutant is fundamentally altered compared with wild type cells. We next used the LC1 point mutant to assess the influence of trypanosome motility on infection in mice. Wesurprisingly found that disrupting parasite motility has no discernible effect on T. brucei bloodstream infection. Infection time-course, maximum parasitaemia, number of waves of parasitaemia, clinical features and disease outcome are indistinguishable between motility mutant and control parasites. Our studies provide an important step toward understanding the contribution of parasite motility to infection and a foundation for future investigations of T. brucei interaction with the mammalian host.

  5. ciliaFA: a research tool for automated, high-throughput measurement of ciliary beat frequency using freely available software.

    PubMed

    Smith, Claire M; Djakow, Jana; Free, Robert C; Djakow, Petr; Lonnen, Rana; Williams, Gwyneth; Pohunek, Petr; Hirst, Robert A; Easton, Andrew J; Andrew, Peter W; O'Callaghan, Christopher

    2012-01-01

    Analysis of ciliary function for assessment of patients suspected of primary ciliary dyskinesia (PCD) and for research studies of respiratory and ependymal cilia requires assessment of both ciliary beat pattern and beat frequency. While direct measurement of beat frequency from high-speed video recordings is the most accurate and reproducible technique it is extremely time consuming. The aim of this study was to develop a freely available automated method of ciliary beat frequency analysis from digital video (AVI) files that runs on open-source software (ImageJ) coupled to Microsoft Excel, and to validate this by comparison to the direct measuring high-speed video recordings of respiratory and ependymal cilia. These models allowed comparison to cilia beating between 3 and 52 Hz. Digital video files of motile ciliated ependymal (frequency range 34 to 52 Hz) and respiratory epithelial cells (frequency 3 to 18 Hz) were captured using a high-speed digital video recorder. To cover the range above between 18 and 37 Hz the frequency of ependymal cilia were slowed by the addition of the pneumococcal toxin pneumolysin. Measurements made directly by timing a given number of individual ciliary beat cycles were compared with those obtained using the automated ciliaFA system. The overall mean difference (± SD) between the ciliaFA and direct measurement high-speed digital imaging methods was -0.05 ± 1.25 Hz, the correlation coefficient was shown to be 0.991 and the Bland-Altman limits of agreement were from -1.99 to 1.49 Hz for respiratory and from -2.55 to 3.25 Hz for ependymal cilia. A plugin for ImageJ was developed that extracts pixel intensities and performs fast Fourier transformation (FFT) using Microsoft Excel. The ciliaFA software allowed automated, high throughput measurement of respiratory and ependymal ciliary beat frequency (range 3 to 52 Hz) and avoids operator error due to selection bias. We have included free access to the ciliaFA plugin and

  6. Cilia and coordination of signaling networks during heart development

    PubMed Central

    Koefoed, Karen; Veland, Iben Rønn; Pedersen, Lotte Bang; Larsen, Lars Allan; Christensen, Søren Tvorup

    2014-01-01

    Primary cilia are unique sensory organelles that coordinate a wide variety of different signaling pathways to control cellular processes during development and in tissue homeostasis. Defects in function or assembly of these antenna-like structures are therefore associated with a broad range of developmental disorders and diseases called ciliopathies. Recent studies have indicated a major role of different populations of cilia, including nodal and cardiac primary cilia, in coordinating heart development, and defects in these cilia are associated with congenital heart disease. Here, we present an overview of the role of nodal and cardiac primary cilia in heart development. PMID:24345806

  7. Regeneration of cilia in heavily irradiated sea urchin embryos

    SciTech Connect

    Rustad, R.C.

    1981-12-01

    Cilia were removed from blastulae, gastrulae, and plutei of the sea urchins Arbacia punctulata and Lytechinus variegatus by shaking the embryos in hypertonic media. Exposure to 50 krad (and in some experiments 100 krad) of ..gamma.. radiation either before or after deciliation had no effect on the time of appearance of regenerating cilia. There were no visually obvious differences in the rate of growth of the cilia in control and irradiated embryos. The cilia commenced beating at the same time, but the initial beating sometimes seemed less vigorous following irradiation. The data support the hypothesis that radiation has no major effect on the assembly from mature basal bodies of the microtubules of cilia.

  8. Development and Distribution of Neuronal Cilia in Mouse Neocortex

    PubMed Central

    Arellano, Jon I.; Guadiana, Sarah M.; Breunig, Joshua J.; Rakic, Pasko; Sarkisian, Matthew R.

    2011-01-01

    Neuronal primary cilia are not generally recognized, but they are considered to extend from most, if not all, neurons in the neocortex. However, when and how cilia develop in neurons are not known. This study used immunohistochemistry for adenylyl cyclase III (ACIII), a marker of primary cilia, and electron microscopic analysis to describe the development and maturation of cilia in mouse neocortical neurons. Our results indicate that ciliogenesis is initiated in late fetal stages after neuroblast migration, when the mother centriole docks with the plasma membrane, becomes a basal body, and grows a cilia bud that we call a procilium. This procilium consists of a membranous protrusion extending from the basal body but lacking axonemal structure and remains undifferentiated until development of the axoneme and cilia elongation starts at about postnatal day 4. Neuronal cilia elongation and final cilia length depend on layer position, and the process extends for a long time, lasting 8–12 weeks. We show that, in addition to pyramidal neurons, inhibitory interneurons also grow cilia of comparable length, suggesting that cilia are indeed present in all neocortical neuron subtypes. Furthermore, the study of mice with defective ciliogenesis suggested that failed elongation of cilia is not essential for proper neuronal migration and laminar organization or establishment of neuronal polarity. Thus, the function of this organelle in neocortical neurons remains elusive. PMID:22020803

  9. The Roles of Primary cilia in Polycystic Kidney Disease

    PubMed Central

    Kathem, Sarmed H.; Mohieldin, Ashraf M.; Nauli, Surya M.

    2014-01-01

    Autosomal dominant polycystic kidney disease (ADPKD) is an inherited genetic disorder that results in progressive renal cyst formation with ultimate loss of renal function and other systemic disorders. These systemic disorders include abnormalities in cardiovascular, portal, pancreatic and gastrointestinal systems. ADPKD is considered to be among the ciliopathy diseases due to the association with abnormal primary cilia function. In order to understand the full course of primary cilia and its association with ADPKD, the structure, functions and role of primary cilia have been meticulously investigated. As a result, the focus on primary cilia has emerged to support the vital roles of primary cilia in ADPKD. The primary cilia have been shown to have not only a mechanosensory function but also a chemosensory function. Both structural and functional defects in primary cilia result in cystic kidney disease and vascular hypertension. Thus, the mechanosenory and chemosensory functions will be analyzed in regards to ADPKD. PMID:25599087

  10. WD60/FAP163 is a dynein intermediate chain required for retrograde intraflagellar transport in cilia.

    PubMed

    Patel-King, Ramila S; Gilberti, Renée M; Hom, Erik F Y; King, Stephen M

    2013-09-01

    Retrograde intraflagellar transport (IFT) is required for assembly of cilia. We identify a Chlamydomonas flagellar protein (flagellar-associated protein 163 [FAP163]) as being closely related to the D1bIC(FAP133) intermediate chain (IC) of the dynein that powers this movement. Biochemical analysis revealed that FAP163 is present in the flagellar matrix and is actively trafficked by IFT. Furthermore, FAP163 copurified with D1bIC(FAP133) and the LC8 dynein light chain, indicating that it is an integral component of the retrograde IFT dynein. To assess the functional role of FAP163, we generated an RNA interference knockdown of the orthologous protein (WD60) in planaria. The Smed-wd60(RNAi) animals had a severe ciliary assembly defect that dramatically compromised whole-organism motility. Most cilia were present as short stubs that had accumulated large quantities of IFT particle-like material between the doublet microtubules and the membrane. The few remaining approximately full-length cilia had a chaotic beat with a frequency reduced from 24 to ∼10 Hz. Thus WD60/FAP163 is a dynein IC that is absolutely required for retrograde IFT and ciliary assembly.

  11. WD60/FAP163 is a dynein intermediate chain required for retrograde intraflagellar transport in cilia

    PubMed Central

    Patel-King, Ramila S.; Gilberti, Renée M.; Hom, Erik F. Y.; King, Stephen M.

    2013-01-01

    Retrograde intraflagellar transport (IFT) is required for assembly of cilia. We identify a Chlamydomonas flagellar protein (flagellar-associated protein 163 [FAP163]) as being closely related to the D1bIC(FAP133) intermediate chain (IC) of the dynein that powers this movement. Biochemical analysis revealed that FAP163 is present in the flagellar matrix and is actively trafficked by IFT. Furthermore, FAP163 copurified with D1bIC(FAP133) and the LC8 dynein light chain, indicating that it is an integral component of the retrograde IFT dynein. To assess the functional role of FAP163, we generated an RNA interference knockdown of the orthologous protein (WD60) in planaria. The Smed-wd60(RNAi) animals had a severe ciliary assembly defect that dramatically compromised whole-organism motility. Most cilia were present as short stubs that had accumulated large quantities of IFT particle–like material between the doublet microtubules and the membrane. The few remaining approximately full-length cilia had a chaotic beat with a frequency reduced from 24 to ∼10 Hz. Thus WD60/FAP163 is a dynein IC that is absolutely required for retrograde IFT and ciliary assembly. PMID:23864713

  12. Learn About GI Motility

    MedlinePlus

    ... eNewsletter Sidebar × MOBILE MENU About Us Learn About GI Motility Digestive Tract Disorders of the Esophagus Disorders ... Pelvic Floor Motility Testing Personal Stories Contact About GI Motility Twitter Facebook YouTube Search Search ... About Us ...

  13. About GI Motility

    MedlinePlus

    ... eNewsletter Sidebar × MOBILE MENU About Us Learn About GI Motility Digestive Tract Disorders of the Esophagus Disorders ... Pelvic Floor Motility Testing Personal Stories Contact About GI Motility Twitter Facebook YouTube Search Search ... About Us ...

  14. Drosophila sensory cilia lacking MKS proteins exhibit striking defects in development but only subtle defects in adults

    PubMed Central

    Titlow, Joshua S.; Davis, Ilan; Barker, Amy R.; Dawe, Helen R.

    2016-01-01

    ABSTRACT Cilia are conserved organelles that have important motility, sensory and signalling roles. The transition zone (TZ) at the base of the cilium is crucial for cilia function, and defects in several TZ proteins are associated with human congenital ciliopathies such as nephronophthisis (NPHP) and Meckel–Gruber syndrome (MKS). In several species, MKS and NPHP proteins form separate complexes that cooperate with Cep290 to assemble the TZ, but flies seem to lack core components of the NPHP module. We show that MKS proteins in flies are spatially separated from Cep290 at the TZ, and that flies mutant for individual MKS genes fail to recruit other MKS proteins to the TZ, whereas Cep290 seems to be recruited normally. Although there are abnormalities in microtubule and membrane organisation in developing MKS mutant cilia, these defects are less apparent in adults, where sensory cilia and sperm flagella seem to function quite normally. Thus, localising MKS proteins to the cilium or flagellum is not essential for viability or fertility in flies. PMID:27577095

  15. Magnetically-actuated artificial cilia for microfluidic propulsion.

    PubMed

    Khaderi, S N; Craus, C B; Hussong, J; Schorr, N; Belardi, J; Westerweel, J; Prucker, O; Rühe, J; den Toonder, J M J; Onck, P R

    2011-06-21

    In this paper we quantitatively analyse the performance of magnetically-driven artificial cilia for lab-on-a-chip applications. The artificial cilia are fabricated using thin polymer films with embedded magnetic nano-particles and their deformation is studied under different external magnetic fields and flows. A coupled magneto-mechanical solid-fluid model that accurately captures the interaction between the magnetic field, cilia and fluid is used to simulate the cilia motion. The elastic and magnetic properties of the cilia are obtained by fitting the results of the computational model to the experimental data. The performance of the artificial cilia with a non-uniform cross-section is characterised using the numerical model for two channel configurations that are of practical importance: an open-loop and a closed-loop channel. We predict that the flow and pressure head generated by the artificial cilia can be as high as 18 microlitres per minute and 3 mm of water, respectively. We also study the effect of metachronal waves on the flow generated and show that the fluid propelled increases drastically compared to synchronously beating cilia, and is unidirectional. This increase is significant even when the phase difference between adjacent cilia is small. The obtained results provide guidelines for the optimal design of magnetically-driven artificial cilia for microfluidic propulsion.

  16. Ciliae-based actuator with piezoelectric excitation

    NASA Astrophysics Data System (ADS)

    Pott, Peter P.; Carrasco, Alvaro; Schlaak, Helmut F.

    2012-06-01

    Small actuators based on the inverse piezoelectric effect are successfully deployed in commercial applications. Usually, ultrasonic motors are used. Based on resonance effects these motors provide a pronounced nonlinearity at low speeds and thus put high demands on the control algorithm. In contrast, piezoelectric stepping motors are mechanically complex and provide only low speeds. The contribution at hand describes a proposed design for a new piezoelectric motor based on cilia friction that can be manufactured at low costs. The cilia are made from uniaxial carbon-fibre reinforced plastics. The derived CFRP-brushes are pressed perpendicularly to the rotor surface to produce force or torque. First experiments prove the feasibility of the concept. A net pushing force of 500 mN is achieved.

  17. Autophagy and primary cilia: dual interplay

    PubMed Central

    Pampliega, Olatz; Cuervo, Ana Maria

    2016-01-01

    Primary cilia are microtubule-based organelles for sensing of the extracellular milieu and transducing this information into the cell through a variety of molecular signaling pathways. Functioning of the primary cilium has been recently connected to autophagy, a pathway for degradation of cellular components in lysosomes. Autophagy regulates the length of the cilia by removing proteins required for ciliogenesis, a phenomenon that is molecularly different if performed by basal autophagy or when autophagy is induced in response to various stressors. Here we review the current knowledge about the dual interaction between autophagy and ciliogenesis, and discuss the potential role that deregulated ciliary autophagy could have in pathologies with alterations in autophagy and ciliogenesis. PMID:26826446

  18. Disruption of Mks1 localization to the mother centriole causes cilia defects and developmental malformations in Meckel-Gruber syndrome

    PubMed Central

    Cui, Cheng; Chatterjee, Bishwanath; Francis, Deanne; Yu, Qing; SanAgustin, Jovenal T.; Francis, Richard; Tansey, Terry; Henry, Charisse; Wang, Baolin; Lemley, Bethan; Pazour, Gregory J.; Lo, Cecilia W.

    2011-01-01

    SUMMARY Meckel-Gruber syndrome (MKS) is a recessive disorder resulting in multiple birth defects that are associated with mutations affecting ciliogenesis. We recovered a mouse mutant with a mutation in the Mks1 gene (Mks1del64-323) that caused a 260-amino-acid deletion spanning nine amino acids in the B9 domain, a protein motif with unknown function conserved in two other basal body proteins. We showed that, in wild-type cells, Mks1 was localized to the mother centriole from which the cilium was generated. However, in mutant Mks1del64-323 cells, Mks1 was not localized to the centriole, even though it maintained a punctate distribution. Resembling MKS patients, Mks1 mutants had craniofacial defects, polydactyly, congenital heart defects, polycystic kidneys and randomized left-right patterning. These defects reflected disturbance of functions subserved by motile and non-motile cilia. In the kidney, glomerular and tubule cysts were observed along with short cilia, and cilia were reduced in number to a near-complete loss. Underlying the left-right patterning defects were fewer and shorter nodal cilia, and analysis with fluorescent beads showed no directional flow at the embryonic node. In the cochlea, the stereocilia were mal-patterned, with the kinocilia being abnormally positioned. Together, these defects suggested disruption of planar cell polarity, which is known to regulate node, kidney and cochlea development. In addition, we also showed that Shh signaling was disrupted. Thus, in the neural tube, the floor plate was not specified posteriorly even as expression of the Shh mediator Gli2 increased. By contrast, the Shh signaling domain was expanded in the anterior neural tube and anterior limb bud, consistent with reduced Gli3-repressor (Gli3R) function. The latter probably accounted for the preaxial digit duplication exhibited by the Mks1del64-323 mutants. Overall, these findings indicate that centriole localization of Mks1 is required for ciliogenesis of motile

  19. Disruption of Mks1 localization to the mother centriole causes cilia defects and developmental malformations in Meckel-Gruber syndrome.

    PubMed

    Cui, Cheng; Chatterjee, Bishwanath; Francis, Deanne; Yu, Qing; SanAgustin, Jovenal T; Francis, Richard; Tansey, Terry; Henry, Charisse; Wang, Baolin; Lemley, Bethan; Pazour, Gregory J; Lo, Cecilia W

    2011-01-01

    Meckel-Gruber syndrome (MKS) is a recessive disorder resulting in multiple birth defects that are associated with mutations affecting ciliogenesis. We recovered a mouse mutant with a mutation in the Mks1 gene (Mks1(del64-323)) that caused a 260-amino-acid deletion spanning nine amino acids in the B9 domain, a protein motif with unknown function conserved in two other basal body proteins. We showed that, in wild-type cells, Mks1 was localized to the mother centriole from which the cilium was generated. However, in mutant Mks1(del64-323) cells, Mks1 was not localized to the centriole, even though it maintained a punctate distribution. Resembling MKS patients, Mks1 mutants had craniofacial defects, polydactyly, congenital heart defects, polycystic kidneys and randomized left-right patterning. These defects reflected disturbance of functions subserved by motile and non-motile cilia. In the kidney, glomerular and tubule cysts were observed along with short cilia, and cilia were reduced in number to a near-complete loss. Underlying the left-right patterning defects were fewer and shorter nodal cilia, and analysis with fluorescent beads showed no directional flow at the embryonic node. In the cochlea, the stereocilia were mal-patterned, with the kinocilia being abnormally positioned. Together, these defects suggested disruption of planar cell polarity, which is known to regulate node, kidney and cochlea development. In addition, we also showed that Shh signaling was disrupted. Thus, in the neural tube, the floor plate was not specified posteriorly even as expression of the Shh mediator Gli2 increased. By contrast, the Shh signaling domain was expanded in the anterior neural tube and anterior limb bud, consistent with reduced Gli3-repressor (Gli3R) function. The latter probably accounted for the preaxial digit duplication exhibited by the Mks1(del64-323) mutants. Overall, these findings indicate that centriole localization of Mks1 is required for ciliogenesis of motile and

  20. Evolutionary implications of localization of the signaling scaffold protein parafusin to both cilia and the nucleus.

    PubMed

    Satir, Birgit Hegner; Wyroba, Elzbieta; Liu, Li; Lethan, Mette; Satir, Peter; Christensen, Søren Tvorup

    2015-02-01

    Parafusin (PFUS), a 63 kDa protein first discovered in the eukaryote Paramecium and known for its role in apicomplexan exocytosis, provides a model for the common origin of cellular systems employing scaffold proteins for targeting and signaling. PFUS is closely related to eubacterial rather than archeal phosphoglucomutases (PGM) - as we proved by comparison of their 88 sequences - but has no PGM activity. Immunofluorescence microscopy analysis with a PFUS-specific peptide antibody showed presence of this protein around the base region of primary cilia in a variety of mammalian cell types, including mouse embryonic (MEFs) and human foreskin fibroblasts (hFFs), human carcinoma stem cells (NT-2 cells), and human retinal pigment epithelial (RPE) cells. Further, PFUS localized to the nucleus of fibroblasts, and prominently to nucleoli of MEFs. Localization studies were confirmed by Western blot analysis, showing that the PFUS antibody specifically recognizes a single protein of ca. 63 kDa in both cytoplasmic and nuclear fractions. Finally, immunofluorescence microscopy analysis showed that PFUS localized to nuclei and cilia in Paramecium. These results support the suggestion that PFUS plays a role in signaling between nucleus and cilia, and that the cilium and the nucleus both evolved around the time of eukaryotic emergence. We hypothesize that near the beginnings of eukaryotic cell evolution, scaffold proteins such as PFUS arose as peripheral membrane protein identifiers for cytoplasmic membrane trafficking and were employed similarly during the subsequent evolution of exocytic, nuclear transport, and ciliogenic mechanisms. © 2014 International Federation for Cell Biology.

  1. Forces applied by cilia measured on explants from mucociliary tissue.

    PubMed

    Teff, Zvi; Priel, Zvi; Gheber, Levi A

    2007-03-01

    Forces applied by intact mucus-propelling cilia were measured for the first time that we know of using a combined atomic force microscopy (AFM) and electrooptic system. The AFM probe was dipped into a field of beating cilia and its time-dependent deflection was recorded as it was struck by the cilia while the electrooptic system simultaneously and colocally measured the frequency to ensure that no perturbation was induced by the AFM probe. Using cilia from frog esophagus, we measured forces of approximately 0.21 nN per cilium during the effective stroke. This value, together with the known internal structure of these cilia, leads to the conclusion that most dynein arms along the length of the axoneme contribute to the effective stroke of these cilia.

  2. IPMC cilia system for artificial muscle applications (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Hwang, Taeseon; Palmre, Viljar; Stalbaum, Tyler P.; Shen, Qi; Trabia, Sarah; Kim, Kwang Jin

    2016-04-01

    Artificial muscle (AM) technology is an excellent candidate for creating cilia-based structures for bio-inspired locomotion, maneuvering, and acoustic systems. We developed an AM based cilia fiber which are soft, flexible, easily shaped and low power consumption. The developed cilium has a diameter of around 200 µm and prepared through polymer injection technique. Nafion was used for base polymer for cilia and fabricated IPMCs via platinum electroless plating process. The prepared cilia were characterized by Fourier transform infrared spectroscopy, differential scanning calorimetry, and thermogravimetric analysis. The 2 point probe was conducted to measure electrode surface resistance of prepared IPMCs. We further characterized the cross-sectional morphology and studied the electromechanical performances (displacement and blocking force) of the prepared IPMC actuators. Also we created prototype mm-sized AM fiber cilia array (3x20) and tested the actuation of AM cilia fiber under external electric field.

  3. Primary Cilia Are Lost in Preinvasive and Invasive Prostate Cancer

    PubMed Central

    Hassounah, Nadia B.; Nagle, Ray; Saboda, Kathylynn; Roe, Denise J.; Dalkin, Bruce L.; McDermott, Kimberly M.

    2013-01-01

    Prostate cancer is the second most commonly diagnosed cancer in men worldwide. Little is known about the role of primary cilia in preinvasive and invasive prostate cancer. However, reduced cilia expression has been observed in human cancers including pancreatic cancer, renal cell carcinoma, breast cancer, cholangiocarcinoma, and melanoma. The aim of this study was to characterize primary cilia expression in preinvasive and invasive human prostate cancer, and to investigate the correlation between primary cilia and the Wnt signaling pathway. Human prostate tissues representative of stages of prostate cancer formation (normal prostate, prostatic intraepithelial neoplasia (PIN), and invasive prostate cancer (including perineural invasion)) were stained for ciliary proteins. The frequency of primary cilia was determined. A decrease in the percentage of ciliated cells in PIN, invasive cancer and perineural invasion lesions was observed when compared to normal. Cilia lengths were also measured to indirectly test functionality. Cilia were shorter in PIN, cancer, and perineural invasion lesions, suggesting dysfunction. Primary cilia have been shown to suppress the Wnt pathway. Increased Wnt signaling has been implicated in prostate cancer. Therefore, we investigated a correlation between loss of primary cilia and increased Wnt signaling in normal prostate and in preinvasive and invasive prostate cancer. To investigate Wnt signaling in our cohort, serial tissue sections were stained for β-catenin as a measure of Wnt signaling. Nuclear β-catenin was analyzed and Wnt signaling was found to be higher in un-ciliated cells in the normal prostate, PIN, a subset of invasive cancers, and perineural invasion. Our results suggest that cilia normally function to suppress the Wnt signaling pathway in epithelial cells and that cilia loss may play a role in increased Wnt signaling in some prostate cancers. These results suggest that cilia are dysfunctional in human prostate cancer, and

  4. Kinesin-2 motors transport IFT-particles, dyneins and tubulin subunits to the tips of Caenorhabditis elegans sensory cilia: relevance to vision research?

    PubMed

    Scholey, Jonathan M

    2012-12-15

    The sensory outer segments (OS) of vertebrate retinal photoreceptors, which detect photons of light, resemble the distal segments of Caenorhabditis elegans sensory cilia, which detect chemical ligands that influence the chemotactic movements of the animal. Based on fluorescence microscopy assays performed in sensory cilia of living, transgenic "wild type" and mutant C. elegans, combined with in vitro motility assays using purified motors, we have proposed that two types of kinesin-2 motor, heterotrimeric kinesin-II and homodimeric OSM-3, cooperate to build amphid and phasmid sensory cilia on chemosensory neurons. Specifically, we propose that these motors function together in a redundant manner to build the axoneme core (aka middle segments (MS)), whereas OSM-3 alone serves to build the distal segments (DS). Furthermore, our data suggest that these motors accomplish this by driving two sequential steps of anterograde transport of cargoes consisting of IFT-particles, retrograde dynein motors, and ciliary tubulin subunits, from the transition zone to the tips of the axonemal microtubules (MTs). Homologs of kinesin-II (KIF3) and OSM-3 (KIF17) are also proposed to contribute to the assembly of vertebrate photoreceptors, although how they do so is currently unclear. Here I review our work on kinesin-2 motors, intraflagellar transport (IFT) and cilium biogenesis in C. elegans sensory cilia, and comment on its possible relevance to current research on vertebrate photoreceptor cilia assembly and function. Copyright © 2012 Elsevier Ltd. All rights reserved.

  5. Mammalian Axoneme Central Pair Complex Proteins: Broader Roles Revealed by Gene Knockout Phenotypes

    PubMed Central

    Teves, Maria E.; Nagarkatti-Gude, David R.; Zhang, Zhibing; Strauss, Jerome F.

    2016-01-01

    The axoneme genes, their encoded proteins, their functions and the structures they form are largely conserved across species. Much of our knowledge of the function and structure of axoneme proteins in cilia and flagella is derived from studies on model organisms like the green algae, Chlamydomonas reinhardtii. The core structure of cilia and flagella is the axoneme, which in most motile cilia and flagella contains a 9 + 2 configuration of microtubules. The two central microtubules are the scaffold of the central pair complex (CPC). Mutations that disrupt CPC genes in Chlamydomonas and other model organisms result in defects in assembly, stability and function of the axoneme, leading to flagellar motility defects. However, targeted mutations generated in mice in the orthologous CPC genes have revealed significant differences in phenotypes of mutants compared to Chlamydomonas. Here we review observations that support the concept of cell-type specific roles for the CPC genes in mice, and an expanded repertoire of functions for the products of these genes in cilia, including non-motile cilia, and other microtubule-associated cellular functions. PMID:26785425

  6. The emerging face of primary cilia

    PubMed Central

    Zaghloul, Norann A.; Brugmann, Samantha A.

    2011-01-01

    Primary cilia are microtubule-based organelles that serve as hubs for the transduction of various developmental signaling pathways including Hedgehog, Wnt, FGF and PDGF. Ciliary dysfunction contributes to a range of disorders, collectively known as the ciliopathies. Recently, interest has grown in these syndromes, particularly among craniofacial biologists, as many known and putative ciliopathies have severe craniofacial defects. Herein we discuss the current understanding of ciliary biology and craniofacial development in an attempt to gain insight into the molecular etiology for craniofacial ciliopathies, and uncover a characteristic ciliopathic craniofacial gestalt. PMID:21305689

  7. ATR promotes cilia signalling: links to developmental impacts

    PubMed Central

    Stiff, Tom; Casar Tena, Teresa; O'Driscoll, Mark; Jeggo, Penny A.; Philipp, Melanie

    2016-01-01

    Mutations in ATR (ataxia telangiectasia and RAD3-related) cause Seckel syndrome (ATR-SS), a microcephalic primordial dwarfism disorder. Hitherto, the clinical manifestation of ATR deficiency has been attributed to its canonical role in DNA damage response signalling following replication fork stalling/collapse. Here, we show that ATR regulates cilia-dependent signalling in a manner that can be uncoupled from its function during replication. ATR-depleted or patient-derived ATR-SS cells form cilia of slightly reduced length but are dramatically impaired in cilia-dependent signalling functions, including growth factor and Sonic hedgehog signalling. To better understand the developmental impact of ATR loss of function, we also used zebrafish as a model. Zebrafish embryos depleted of Atr resembled ATR-SS morphology, showed a modest but statistically significant reduction in cilia length and other morphological features indicative of cilia dysfunction. Additionally, they displayed defects in left-right asymmetry including ambiguous expression of southpaw, incorrectly looped hearts and randomized localization of internal organs including the pancreas, features typically conferred by cilia dysfunction. Our findings reveal a novel role for ATR in cilia signalling distinct from its canonical function during replication and strengthen emerging links between cilia function and development. PMID:26908596

  8. Cilia and polycystic kidney disease, kith and kin.

    PubMed

    Huang, Liwei; Lipschutz, Joshua H

    2014-06-01

    In the past decade, cilia have been found to play important roles in renal cystogenesis. Many genes, such as PKD1 and PKD2 which, when mutated, cause autosomal dominant polycystic kidney disease (ADPKD), have been found to localize to primary cilia. The cilium functions as a sensor to transmit extracellular signals into the cell. Abnormal cilia structure and function are associated with the development of polyscystic kidney disease (PKD). Cilia assembly includes centriole migration to the apical surface of the cell, ciliary vesicle docking and fusion with the cell membrane at the intended site of cilium outgrowth, and microtubule growth from the basal body. This review summarizes the most recent advances in cilia and PKD research, with special emphasis on the mechanisms of cytoplasmic and intraciliary protein transport during ciliogenesis. Copyright © 2014 Wiley Periodicals, Inc.

  9. Cilia and Polycystic Kidney Disease, Kith and Kin

    PubMed Central

    Huang, Liwei; Lipschutz, Joshua H.

    2015-01-01

    In the past decade, cilia have been found to play important roles in renal cystogenesis. Many genes, such as PKD1 and PKD2 which, when mutated, cause autosomal dominant polycystic kidney disease (ADPKD), have been found to localize to primary cilia. The cilium functions as a sensor to transmit extracellular signals into the cell. Abnormal cilia structure and function are associated with the development of polyscystic kidney disease (PKD). Cilia assembly includes centriole migration to the apical surface of the cell, ciliary vesicle docking and fusion with the cell membrane at the intended site of cilium outgrowth, and microtubule growth from the basal body. This review summarizes the most recent advances in cilia and PKD research, with special emphasis on the mechanisms of cytoplasmic and intraciliary protein transport during ciliogenesis. PMID:24898006

  10. Proteomic Analysis of Bovine Axonemes Exposed to Acute Alcohol: Role of eNOS and HSP90 in Cilia Stimulation

    PubMed Central

    Simet, Samantha M.; Pavlik, Jacqueline A.; Sisson, Joseph H.

    2012-01-01

    Background Cilia are fingerlike motor-driven organelles, which propel inhaled particles and mucus from the lung and airways. We have previously shown that brief alcohol exposure stimulates ciliary motility through an endothelial nitric oxide (eNOS)-dependent pathway localized in the ciliary metabolon. However, the signaling molecules of the ciliary metabolon involved in alcohol-triggered cilia beat frequency (CBF) stimulation upstream of eNOS activation are unknown. Methods and Results We hypothesized that brief alcohol exposure alters threonine and serine phosphorylation of proteins involved in stimulating ciliary beat frequency. Two-dimensional electrophoresis indicated both increases and deceases in the serine and threonine phosphorylation states of several proteins. One of the proteins identified was heat shock protein 90 (HSP90), which undergoes increased threonine phosphorylation after brief alcohol exposure. Because HSP90 has been shown to associate with eNOS in lung tissue, we hypothesized that HSP90 is a key component in alcohol-triggered eNOS activation and that these two proteins co-localize within the ciliary metabolon. Immunofluorescence experiments demonstrate that eNOS and HSP90 co-localize within basal bodies of the ciliary metabolon and partially translocate to the axoneme upon brief alcohol exposure. Pretreatment with geldanamycin, which disrupts HSP90 chaperone functions, prevented eNOS-HSP90 association and prevented the translocation of eNOS from the ciliary metabolon to the axoneme. Functional cilia motility studies revealed that geldanamycin blocked alcohol-stimulated ciliary motility in bovine bronchial epithelial cells and mouse tracheal rings. Conclusions Based on the HSP90 localization with eNOS, alcohol activation of HSP90 phosphorylation, and geldanamycin’s ability inhibit HSP90-eNOS association, prevent eNOS translocation to the axoneme, and block alcohol-stimulated ciliary motility, we conclude that alcohol-induced cilia stimulation

  11. RFX7 is required for the formation of cilia in the neural tube.

    PubMed

    Manojlovic, Zarko; Earwood, Ryan; Kato, Akiko; Stefanovic, Branko; Kato, Yoichi

    2014-05-01

    Regulatory Factor X (RFX) transcription factors are important for development and are likely involved in the pathogenesis of serious human diseases including ciliopathies. While seven RFX genes have been identified in vertebrates and several RFX transcription factors have been reported to be regulators of ciliogenesis, the role of RFX7 in development including ciliogenesis is not known. Here we show that RFX7 in Xenopus laevis is expressed in the neural tube, eye, otic vesicles, and somites. Knockdown of RFX7 in Xenopus embryos resulted in a defect of ciliogenesis in the neural tube and failure of neural tube closure. RFX7 controlled the formation of cilia by regulating the expression of RFX4 gene, which has been reported to be required for ciliogenesis in the neural tube. Moreover, ectopic expression of Foxj1, which is a master regulator of motile cilia formation, suppressed the expression of RFX4 but not RFX7. Taken together, RFX7 plays an important role in the process of neural tube closure at the top of the molecular cascade which controls ciliogenesis in the neural tube.

  12. Biochemical Studies of Olfaction: Role of Cilia in Odorant Recognition

    PubMed Central

    Rhein, L. D.

    1983-01-01

    Chemoreception in vertebrates is beginning to be understood. Numerous anatomical, behavioral, and physiological studies are now available. Current research efforts are examining the molecular basis of chemoreception. Rainbow trout (Salmo gairdneri) have a functional olfactory system and are a suitable vertebrate model for studying odorant interactions with receptors. Using a biochemical approach, initial events of olfactory recognition were examined; the aim was to determine the location and specificity of odor receptors. Cilia occupy the distal region of the receptor neuron on the trout olfactory epithelium, and their membranes are the postulated locus of odorant receptor sites. A cilia preparation was isolated from the olfactory rosette. The preparation was characterized by quantifying biochemical markers for cilia, along with electron microscopy, all of which substantiated enrichment of cilia. Functional activity was assessed by quantifying binding of several radioactively labeled odorant amino acids. The odorants bound to the cilia in a manner similar to the sedimentable preparation previously isolated from t h e olfactory rosette of the same animal, thus verifying the presence of odor receptors in the cilia preparation. Evidence also confirmed a site TSA which binds L-threonine, L-serine, and L-alanine and a site L which binds L-lysine (and L-arginine). Binding of L-serine and D-alanine showed evidence for a single affinity site while the others showed two affinity sites. Separation of membrane fractions from the cilia preparation revealed that binding activity is associated with a very low density membrane fraction B. PMID:19295786

  13. Fetus Sound Stimulation: Cilia Memristor Effect of Signal Transduction

    PubMed Central

    Jankovic-Raznatovic, Svetlana; Dragojevic-Dikic, Svetlana; Rakic, Snezana; Nikolic, Branka; Plesinac, Snezana; Tasic, Lidija; Perisic, Zivko; Sovilj, Mirjana; Adamovic, Tatjana; Koruga, Djuro

    2014-01-01

    Background. This experimental study evaluates fetal middle cerebral artery (MCA) circulation after the defined prenatal acoustical stimulation (PAS) and the role of cilia in hearing and memory and could explain signal transduction and memory according to cilia optical-acoustical properties. Methods. PAS was performed twice on 119 no-risk term pregnancies. We analyzed fetal MCA circulation before, after first and second PAS. Results. Analysis of the Pulsatility index basic (PIB) and before PAS and Pulsatility index reactive after the first PAS (PIR 1) shows high statistical difference, representing high influence on the brain circulation. Analysis of PIB and Pulsatility index reactive after the second PAS (PIR 2) shows no statistical difference. Cilia as nanoscale structure possess magnetic flux linkage that depends on the amount of charge that has passed between two-terminal variable resistors of cilia. Microtubule resistance, as a function of the current through and voltage across the structure, leads to appearance of cilia memory with the “memristor” property. Conclusion. Acoustical and optical cilia properties play crucial role in hearing and memory processes. We suggest that fetuses are getting used to sound, developing a kind of memory patterns, considering acoustical and electromagnetically waves and involving cilia and microtubules and try to explain signal transduction. PMID:24719851

  14. Fetus sound stimulation: cilia memristor effect of signal transduction.

    PubMed

    Jankovic-Raznatovic, Svetlana; Dragojevic-Dikic, Svetlana; Rakic, Snezana; Nikolic, Branka; Plesinac, Snezana; Tasic, Lidija; Perisic, Zivko; Sovilj, Mirjana; Adamovic, Tatjana; Koruga, Djuro

    2014-01-01

    This experimental study evaluates fetal middle cerebral artery (MCA) circulation after the defined prenatal acoustical stimulation (PAS) and the role of cilia in hearing and memory and could explain signal transduction and memory according to cilia optical-acoustical properties. PAS was performed twice on 119 no-risk term pregnancies. We analyzed fetal MCA circulation before, after first and second PAS. Analysis of the Pulsatility index basic (PIB) and before PAS and Pulsatility index reactive after the first PAS (PIR 1) shows high statistical difference, representing high influence on the brain circulation. Analysis of PIB and Pulsatility index reactive after the second PAS (PIR 2) shows no statistical difference. Cilia as nanoscale structure possess magnetic flux linkage that depends on the amount of charge that has passed between two-terminal variable resistors of cilia. Microtubule resistance, as a function of the current through and voltage across the structure, leads to appearance of cilia memory with the "memristor" property. Acoustical and optical cilia properties play crucial role in hearing and memory processes. We suggest that fetuses are getting used to sound, developing a kind of memory patterns, considering acoustical and electromagnetically waves and involving cilia and microtubules and try to explain signal transduction.

  15. Micro-fluidic actuation using magnetic artificial cilia.

    PubMed

    Fahrni, Francis; Prins, Menno W J; van Ijzendoorn, Leo J

    2009-12-07

    We demonstrate advanced fluid manipulations using magnetic polymeric artificial cilia on the walls of a microfluidic channel. In nature, cilia are little hairs covering the surface of micro-organisms which enable them to manipulate a fluid on the micro-scale. The asymmetric movement of natural cilia is crucial to obtain a net fluid flow. We have developed a ferromagnetic polymer made from iron nanoparticles and polydimethylsiloxane, and describe a process that can structure the material into high aspect ratio lying artificial cilia with a length of 300 microm. These artificial cilia were actuated with a homogeneous rotating magnetic field (micro(0)H < 50 mT) generated with a compact external electromagnet. An asymmetric movement involving torsion could be created when the cilia were provided with a remanent magnetisation perpendicular to the plane of rotation of the magnetic field vector. The artificial cilia could be actuated in fluid up to a frequency of approximately 50 Hz. In an aqueous solution in a microfluidic chamber we were able to generate rotational as well as translational fluid movements with fluid velocities up to approximately 0.5 mm s(-1).

  16. Cystic kidney gene seahorse regulates cilia-mediated processes and Wnt pathways.

    PubMed

    Kishimoto, Norihito; Cao, Ying; Park, Alice; Sun, Zhaoxia

    2008-06-01

    Recently the cilium has emerged as an important sensory organelle for a wide range of cell types in vertebrates. However, the signaling cascade that links ciliary signals to cellular events remains poorly understood. Here, we show that the zebrafish cystic kidney gene seahorse is closely associated with ciliary functions: seahorse is required for establishing left-right asymmetry and for preventing kidney cyst formation; seahorse transcript is highly enriched in heavily ciliated tissues; and seahorse genetically interacts with the ciliary gene inversin. Yet seahorse is dispensable for cilia assembly or motility and the Seahorse protein is cytoplasmic. We provide evidence that Seahorse associates with Dishevelled. Finally, we show that seahorse constrains the canonical Wnt pathway and promotes the noncanonical Wnt pathway during gastrulation. Together, these data suggest that Seahorse may provide a link between ciliary signals and Wnt pathways.

  17. Emerging roles for renal primary cilia in epithelial repair.

    PubMed

    Deane, James A; Ricardo, Sharon D

    2012-01-01

    Primary cilia are microscopic sensory antennae that cells in many vertebrate tissues use to gather information about their environment. In the kidney, primary cilia sense urine flow and are essential for the maintenance of epithelial architecture. Defects of this organelle cause the cystic kidney disease characterized by epithelial abnormalities. These findings link primary cilia to the regulation of epithelial differentiation and proliferation, processes that must be precisely controlled during epithelial repair in the kidney. Here, we consider likely roles for primary cilium-based signaling during responses to renal injury and ensuing epithelial repair processes.

  18. Primary cilia and coordination of receptor tyrosine kinase (RTK) signalling

    PubMed Central

    Christensen, Søren T; Clement, Christian A; Satir, Peter; Pedersen, Lotte B

    2015-01-01

    Primary cilia are microtubule-based sensory organelles that coordinate signalling pathways in cell-cycle control, migration, differentiation and other cellular processes critical during development and for tissue homeostasis. Accordingly, defects in assembly or function of primary cilia lead to a plethora of developmental disorders and pathological conditions now known as ciliopathies. In this review, we summarize the current status of the role of primary cilia in coordinating receptor tyrosine kinase (RTK) signalling pathways. Further, we present potential mechanisms of signalling crosstalk and networking in the primary cilium and discuss how defects in ciliary RTK signalling are linked to human diseases and disorders. PMID:21956154

  19. CLEM Methods for Studying Primary Cilia.

    PubMed

    Macaluso, Frank P; Perumal, Geoffrey S; Kolstrup, Johan; Satir, Peter

    2016-01-01

    CLEM (correlated light and electron microscope) imaging is a highly useful technique for examining primary cilia. With CLEM, it is possible to determine the distribution of tagged proteins along the ciliary membrane and axoneme with high precision. Scanning electron microscopy (SEM) permits measurement of ciliary length and orientation in relation to nearby cellular structures in a 3D image; in optimal cases, this can be combined with superresolution microscopy of selected ciliary components as they enter or leave the cilium. This chapter discusses CLEM methods. In the method described in detail, samples are completely processed for sequential fluorescence and SEM observation. This method is ideal for robust antibody localization and minimizes image manipulation in correlating the fluorescent and SEM images. Alternative methods prepare samples for fluorescence imaging followed by processing for SEM then observation in the SEM. This method is ideal for optimal fluorescence imaging, particularly live cell imaging.

  20. A phenotypic screening platform to identify small molecule modulators of Chlamydomonas reinhardtii growth, motility and photosynthesis

    PubMed Central

    2012-01-01

    Chemical biology, the interfacial discipline of using small molecules as probes to investigate biology, is a powerful approach of developing specific, rapidly acting tools that can be applied across organisms. The single-celled alga Chlamydomonas reinhardtii is an excellent model system because of its photosynthetic ability, cilia-related motility and simple genetics. We report the results of an automated fitness screen of 5,445 small molecules and subsequent assays on motility/phototaxis and photosynthesis. Cheminformatic analysis revealed active core structures and was used to construct a naïve Bayes model that successfully predicts algal bioactive compounds. PMID:23158586

  1. Hydrodynamic interactions of cilia on a spherical body

    NASA Astrophysics Data System (ADS)

    Nasouri, Babak; Elfring, Gwynn J.

    2015-11-01

    The emergence of metachronal waves in ciliated microorganisms can arise solely from the hydrodynamic interactions between the cilia. For a chain of cilia attached to a flat ciliate, it was observed that fluid forces can lead the system to form a metachronal wave. However, several microorganisms such as paramecium and volvox possess a curved shaped ciliate body. To understand the effect of this geometry on the formation of metachronal waves, we evaluate the hydrodynamic interactions of cilia near a large spherical body. Using a minimal model, we show that for a chain of cilia around the sphere, the embedded periodicity in the geometry leads the system to synchronize. We also report an emergent wave-like behavior when an asymmetry is introduced to the system.

  2. Cilia in the CNS: the Quiet Organelle Claims Center Stage

    PubMed Central

    Louvi, Angeliki; Grove, Elizabeth A.

    2011-01-01

    Summary The primary cilium is a cellular organelle that is almost ubiquitous in eukaryotes, yet its functions in vertebrates have been slow to emerge. The last fifteen years have been marked by accelerating insight into the biology of primary cilia, arising from the synergy of three major lines of research. These research programs describe a specialized mode of protein trafficking in cilia, reveal that genetic disruptions of primary cilia cause complex human disease syndromes, and establish that Sonic hedgehog (Shh) signal transduction requires the primary cilium. New lines of research have branched off to investigate the role of primary cilia in neuronal signaling, adult neurogenesis, and brain tumor formation. We review a fast expanding literature to determine what we now know about the primary cilium in the developing and adult CNS, and what new directions should lead to further clarity. PMID:21435552

  3. Hydrodynamic interactions of cilia on a spherical body.

    PubMed

    Nasouri, Babak; Elfring, Gwynn J

    2016-03-01

    Microorganisms develop coordinated beating patterns on surfaces lined with cilia known as metachronal waves. For a chain of cilia attached to a flat ciliate, it has been shown that hydrodynamic interactions alone can lead the system to synchronize. However, several microorganisms possess a curve-shaped ciliate body and so to understand the effect of this geometry on the formation of metachronal waves, we evaluate the hydrodynamic interactions of cilia near a large spherical body. Using a minimal model, we show that for a chain of cilia around the sphere, the natural periodicity in the geometry leads the system to synchronize. We also report an emergent wavelike behavior when an asymmetry is introduced to the system.

  4. What we can learn from a tadpole about ciliopathies and airway diseases: Using systems biology in Xenopus to study cilia and mucociliary epithelia.

    PubMed

    Walentek, Peter; Quigley, Ian K

    2017-01-01

    Over the past years, the Xenopus embryo has emerged as an incredibly useful model organism for studying the formation and function of cilia and ciliated epithelia in vivo. This has led to a variety of findings elucidating the molecular mechanisms of ciliated cell specification, basal body biogenesis, cilia assembly, and ciliary motility. These findings also revealed the deep functional conservation of signaling, transcriptional, post-transcriptional, and protein networks employed in the formation and function of vertebrate ciliated cells. Therefore, Xenopus research can contribute crucial insights not only into developmental and cell biology, but also into the molecular mechanisms underlying cilia related diseases (ciliopathies) as well as diseases affecting the ciliated epithelium of the respiratory tract in humans (e.g., chronic lung diseases). Additionally, systems biology approaches including transcriptomics, genomics, and proteomics have been rapidly adapted for use in Xenopus, and broaden the applications for current and future translational biomedical research. This review aims to present the advantages of using Xenopus for cilia research, highlight some of the evolutionarily conserved key concepts and mechanisms of ciliated cell biology that were elucidated using the Xenopus model, and describe the potential for Xenopus research to address unresolved questions regarding the molecular mechanisms of ciliopathies and airway diseases. © 2017 Wiley Periodicals, Inc.

  5. Microscale imaging of cilia-driven fluid flow

    PubMed Central

    Huang, Brendan K.; Choma, Michael A.

    2015-01-01

    Cilia-driven fluid flow is important for multiple processes in the body, including respiratory mucus clearance, gamete transport in the oviduct, right-left patterning in the embryonic node, and cerebrospinal fluid circulation. Multiple imaging techniques have been applied towards quantifying ciliary flow. Here we review common velocimetry methods of quantifying fluid flow. We then discuss four important optical modalities, including light microscopy, epifluorescence, confocal microscopy, and optical coherence tomography, that have been used to investigate cilia-driven flow. PMID:25417211

  6. Tracing the origins of centrioles, cilia, and flagella

    PubMed Central

    Azimzadeh, Juliette; Pereira-Leal, José. B.

    2011-01-01

    Centrioles/basal bodies (CBBs) are microtubule-based cylindrical organelles that nucleate the formation of centrosomes, cilia, and flagella. CBBs, cilia, and flagella are ancestral structures; they are present in all major eukaryotic groups. Despite the conservation of their core structure, there is variability in their architecture, function, and biogenesis. Recent genomic and functional studies have provided insight into the evolution of the structure and function of these organelles. PMID:21788366

  7. Branchial Cilia and Sperm Flagella Recruit Distinct Axonemal Components

    PubMed Central

    Konno, Alu; Shiba, Kogiku; Cai, Chunhua; Inaba, Kazuo

    2015-01-01

    Eukaryotic cilia and flagella have highly conserved 9 + 2 structures. They are functionally diverged to play cell-type-specific roles even in a multicellular organism. Although their structural components are therefore believed to be common, few studies have investigated the molecular diversity of the protein components of the cilia and flagella in a single organism. Here we carried out a proteomic analysis and compared protein components between branchial cilia and sperm flagella in a marine invertebrate chordate, Ciona intestinalis. Distinct feature of protein recruitment in branchial cilia and sperm flagella has been clarified; (1) Isoforms of α- and β-tubulins as well as those of actins are distinctly used in branchial cilia or sperm flagella. (2) Structural components, such as dynein docking complex, tektins and an outer dense fiber protein, are used differently by the cilia and flagella. (3) Sperm flagella are specialized for the cAMP- and Ca2+-dependent regulation of outer arm dynein and for energy metabolism by glycolytic enzymes. Our present study clearly demonstrates that flagellar or ciliary proteins are properly recruited according to their function and stability, despite their apparent structural resemblance and conservation. PMID:25962172

  8. Left–Right Determination: Involvement of Molecular Motor KIF3, Cilia, and Nodal Flow

    PubMed Central

    Hirokawa, Nobutaka; Tanaka, Yosuke; Okada, Yasushi

    2009-01-01

    Mammalian left–right determination is a good example for how multiple cell biological processes coordinate in the formation of a basic body plan. The leftward movement of fluid at the ventral node, called nodal flow, is the central process in symmetry breaking on the left–right axis. Nodal flow is autonomously generated by the rotation of posteriorly tilted cilia that are built by transport via KIF3 motor on cells of the ventral node. How nodal flow is interpreted to create left–right asymmetry has been a matter of debate. Recent evidence suggests that the leftward movement of sheathed lipidic particles, called nodal vesicular parcels (NVPs), may result in the activation of the noncanonical hedgehog signaling pathway, an asymmetric elevation in intracellular Ca2+ and changes in gene expression. PMID:20066075

  9. Modelling the fluid mechanics of cilia and flagella in reproduction and development.

    PubMed

    Montenegro-Johnson, Thomas D; Smith, Andrew A; Smith, David J; Loghin, Daniel; Blake, John R

    2012-10-01

    Cilia and flagella are actively bending slender organelles, performing functions such as motility, feeding and embryonic symmetry breaking. We review the mechanics of viscous-dominated microscale flow, including time-reversal symmetry, drag anisotropy of slender bodies, and wall effects. We focus on the fundamental force singularity, higher-order multipoles, and the method of images, providing physical insight and forming a basis for computational approaches. Two biological problems are then considered in more detail: 1) left-right symmetry breaking flow in the node, a microscopic structure in developing vertebrate embryos, and 2) motility of microswimmers through non-Newtonian fluids. Our model of the embryonic node reveals how particle transport associated with morphogenesis is modulated by the gradual emergence of cilium posterior tilt. Our model of swimming makes use of force distributions within a body-conforming finite-element framework, allowing the solution of nonlinear inertialess Carreau flow. We find that a three-sphere model swimmer and a model sperm are similarly affected by shear-thinning; in both cases swimming due to a prescribed beat is enhanced by shear-thinning, with optimal Deborah number around 0.8. The sperm exhibits an almost perfect linear relationship between velocity and the logarithm of the ratio of zero to infinite shear viscosity, with shear-thickening hindering cell progress.

  10. Experimental investigation of the flow induced by artificial cilia.

    PubMed

    Hussong, J; Schorr, N; Belardi, J; Prucker, O; Rühe, J; Westerweel, J

    2011-06-21

    The fluid transport produced by rectangular shaped, magnetically actuated artificial cilia of 70 μm length and 20 μm width was determined by means of phase-locked Micro Particle Image Velocimetry (μPIV) measurements in a closed microfluidic chamber. The phase-averaged flow produced by the artificial cilia reached up to 130 μm s(-1) with an actuation cycle frequency of 10 Hz. Analysis of the measured flow data indicate that the present system is capable of achieving volume flow rates of V[combining dot above](cilia) = 14 ± 4 μl min(-1) in a micro channel of 0.5 × 5 mm(2) cross-sectional area when no back pressure is built up. This corresponds to an effective pressure gradient of 6 ± 1 Pa m(-1), which equals a pressure difference of 0.6 ± 0.1 mPa over a distance of 100 μm between two rows of cilia. These results were derived analytically from the measured velocity profile by treating the cilia as a thin boundary layer. While the cilia produce phase-averaged velocities of the order of O(10(2)μm s(-1)), time-resolved measurements showed that the flow field reverses two times during one actuation cycle inducing instantaneous velocities of up to approximately 2 mm s(-1). This shows that the flow field is dominated by fluid oscillations and flow rates are expected to increase if the beating motion of the cilia is further improved.

  11. TULP3 bridges the IFT-A complex and membrane phosphoinositides to promote trafficking of G protein-coupled receptors into primary cilia.

    PubMed

    Mukhopadhyay, Saikat; Wen, Xiaohui; Chih, Ben; Nelson, Christopher D; Lane, William S; Scales, Suzie J; Jackson, Peter K

    2010-10-01

    Primary cilia function as a sensory signaling compartment in processes ranging from mammalian Hedgehog signaling to neuronal control of obesity. Intraflagellar transport (IFT) is an ancient, conserved mechanism required to assemble cilia and for trafficking within cilia. The link between IFT, sensory signaling, and obesity is not clearly defined, but some novel monogenic obesity disorders may be linked to ciliary defects. The tubby mouse, which presents with adult-onset obesity, arises from mutation in the Tub gene. The tubby-like proteins comprise a related family of poorly understood proteins with roles in neural development and function. We find that specific Tubby family proteins, notably Tubby-like protein 3 (TULP3), bind to the IFT-A complex. IFT-A is linked to retrograde ciliary transport, but, surprisingly, we find that the IFT-A complex has a second role directing ciliary entry of TULP3. TULP3 and IFT-A, in turn, promote trafficking of a subset of G protein-coupled receptors (GPCRs), but not Smoothened, to cilia. Both IFT-A and membrane phosphoinositide-binding properties of TULP3 are required for ciliary GPCR localization. TULP3 and IFT-A proteins both negatively regulate Hedgehog signaling in the mouse embryo, and the TULP3-IFT-A interaction suggests how these proteins cooperate during neural tube patterning.

  12. Microscale flow propulsion through bioinspired and magnetically actuated artificial cilia.

    PubMed

    Chen, Chia-Yuan; Cheng, Ling-Ying; Hsu, Chun-Chieh; Mani, Karthick

    2015-05-01

    Recent advances in microscale flow propulsion through bioinspired artificial cilia provide a promising alternative for lab-on-a-chip applications. However, the ability of actuating artificial cilia to achieve a time-dependent local flow control with high accuracy together with the elegance of full integration into the biocompatible microfluidic platforms remains remote. Driven by this motive, the current work has constructed a series of artificial cilia inside a microchannel to facilitate the time-dependent flow propulsion through artificial cilia actuation with high-speed (>40 Hz) circular beating behavior. The generated flow was quantified using micro-particle image velocimetry and particle tracking with instantaneous net flow velocity of up to 10(1 ) μm/s. Induced flow patterns caused by the tilted conical motion of artificial cilia constitutes efficient fluid propulsion at microscale. This flow phenomenon was further measured and illustrated by examining the induced flow behavior across the depth of the microchannel to provide a global view of the underlying flow propulsion mechanism. The presented analytic paradigms and substantial flow evidence present novel insights into the area of flow manipulation at microscale.

  13. Peroxiredoxin 1 is involved in disassembly of flagella and cilia.

    PubMed

    Gong, Fanghua; Liu, Hongtao; Li, Jie; Xue, Lexun; Zhang, Mingzhi

    2014-02-14

    Cilia/flagella are evolutionarily conserved cellular organelles. In this study, we demonstrated that Dunaliella salina Peroxiredoxin 1 (DsPrdx1) localized to the flagella and basal bodies, and was involved in flagellar disassembly. The link between DsPrdx1 and flagella of Dunaliella salina (D. salina) encouraged us to explore the function of its human homologue, Homo sapiens Peroxiredoxin 1 (HsPrdx1) in development and physiology. Our results showed that HsPrdx1 was overexpressed, and cilia were lost in esophageal squamous cell carcinoma (ESCC) cells compared with the non-cancerous esophageal epithelial cells Het-1A. Furthermore, when HsPrdx1 was knocked down by short hairpin RNA (shRNA) lentivirus in ESCC cells, the phenotype of cilia lost can be reversed, and the expression levels of tumor suppressor genes LKB1 and p-AMPK were increased, and the activity of the oncogene Aurora A was inhibited compared with those in cells transfected with scrambe-shRNA lentivirus. These findings firstly showed that Prdx1 is involved in disassembly of flagella and cilia, and suggested that the abnormal expression of the cilia-related gene including Prdx1 may affect both ciliogenesis and cancernogenesis.

  14. Airway Epithelial Cell Cilia and Obstructive Lung Disease

    PubMed Central

    Yaghi, Asma; Dolovich, Myrna B.

    2016-01-01

    Airway epithelium is the first line of defense against exposure of the airway and lung to various inflammatory stimuli. Ciliary beating of airway epithelial cells constitutes an important part of the mucociliary transport apparatus. To be effective in transporting secretions out of the lung, the mucociliary transport apparatus must exhibit a cohesive beating of all ciliated epithelial cells that line the upper and lower respiratory tract. Cilia function can be modulated by exposures to endogenous and exogenous factors and by the viscosity of the mucus lining the epithelium. Cilia function is impaired in lung diseases such as COPD and asthma, and pharmacologic agents can modulate cilia function and mucus viscosity. Cilia beating is reduced in COPD, however, more research is needed to determine the structural-functional regulation of ciliary beating via all signaling pathways and how this might relate to the initiation or progression of obstructive lung diseases. Additionally, genotypes and how these can influence phenotypes and epithelial cell cilia function and structure should be taken into consideration in future investigations. PMID:27845721

  15. Microscale flow propulsion through bioinspired and magnetically actuated artificial cilia

    PubMed Central

    Chen, Chia-Yuan; Cheng, Ling-Ying; Hsu, Chun-Chieh; Mani, Karthick

    2015-01-01

    Recent advances in microscale flow propulsion through bioinspired artificial cilia provide a promising alternative for lab-on-a-chip applications. However, the ability of actuating artificial cilia to achieve a time-dependent local flow control with high accuracy together with the elegance of full integration into the biocompatible microfluidic platforms remains remote. Driven by this motive, the current work has constructed a series of artificial cilia inside a microchannel to facilitate the time-dependent flow propulsion through artificial cilia actuation with high-speed (>40 Hz) circular beating behavior. The generated flow was quantified using micro-particle image velocimetry and particle tracking with instantaneous net flow velocity of up to 101 μm/s. Induced flow patterns caused by the tilted conical motion of artificial cilia constitutes efficient fluid propulsion at microscale. This flow phenomenon was further measured and illustrated by examining the induced flow behavior across the depth of the microchannel to provide a global view of the underlying flow propulsion mechanism. The presented analytic paradigms and substantial flow evidence present novel insights into the area of flow manipulation at microscale. PMID:26045730

  16. Flagellar motility of the pathogenic spirochetes

    PubMed Central

    Wolgemuth, Charles W.

    2016-01-01

    Bacterial pathogens are often classified by their toxicity and invasiveness. The invasiveness of a given bacterium is determined by how capable the bacterium is at invading a broad range of tissues in its host. Of mammalian pathogens, some of the most invasive come from a group of bacteria known as the spirochetes, which cause diseases such as syphilis, Lyme disease, relapsing fever and leptospirosis. Most of the spirochetes are characterized by their distinct shapes and unique motility. They are long, thin bacteria that can be shaped like flat-waves, helices, or have more irregular morphologies. Like many other bacteria, the spirochetes use long, helical appendages known as flagella to move; however, the spirochetes enclose their flagella in the periplasm, the narrow space between the inner and outer membranes. Rotation of the flagella in the periplasm causes the entire cell body to rotate and/or undulate. These deformations of the bacterium produce the force that drives the motility of these organisms, and it is this unique motility that likely allows these bacteria to be highly invasive in mammals. This review will describe the current state of knowledge on the motility and biophysics of these organisms and provide evidence on how this knowledge can inform our understanding of spirochetal diseases. PMID:26481969

  17. Centriole distal appendages promote membrane docking, leading to cilia initiation

    PubMed Central

    Tanos, Barbara E.; Yang, Hui-Ju; Soni, Rajesh; Wang, Won-Jing; Macaluso, Frank P.; Asara, John M.; Tsou, Meng-Fu Bryan

    2013-01-01

    The distal appendages (DAPs) of centrioles have been proposed to anchor cilia to the plasma membrane, but their molecular composition, assembly, and exact function in ciliogenesis remain poorly understood. Using quantitative centrosome proteomics and superresolution microscopy, we identified five DAP components, including one previously described (CEP164), one partially characterized (CEP89 [ccdc123]), and three novel (CEP83 [ccdc41], SCLT1, and FBF1) DAP proteins. Analyses of DAP assembly revealed a hierarchy. CEP83 recruits both SCLT1 and CEP89 to centrioles. Subsequent recruitment of FBF1 and CEP164 is independent of CEP89 but mediated by SCLT1. All five DAP components are essential for ciliogenesis; loss of CEP83 specifically blocks centriole-to-membrane docking. Undocked centrioles fail to recruit TTBK2 or release CP110, the two earliest modifications found on centrioles prior to cilia assembly, revealing centriole-to-membrane docking as a temporal and spatial cue promoting cilia initiation. PMID:23348840

  18. Analysis of primary cilia in directional cell migration in fibroblasts.

    PubMed

    Christensen, Søren T; Veland, Iben R; Schwab, Albrecht; Cammer, Michael; Satir, Peter

    2013-01-01

    Early studies of migrating fibroblasts showed that primary cilia orient in front of the nucleus and point toward the leading edge. Recent work has shown that primary cilia coordinate a series of signaling pathways critical to fibroblast cell migration during development and in wound healing. In particular, platelet-derived growth factor receptor alpha (PDGFRα) is compartmentalized to the primary cilium to activate signaling pathways that regulate reorganization of the cytoskeleton required for lamellipodium formation and directional migration in the presence of a specific ligand gradient. We summarize selected methods in analyzing ciliary function in directional cell migration, including immunofluorescence microscopy, scratch assay, and chemotaxis assay by micropipette addition of PDGFRα ligands to cultures of fibroblasts. These methods should be useful not only in studying cell migration but also more generally in delineating response pathways in cells with primary cilia. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. Primary cilia and coordination of receptor tyrosine kinase (RTK) signalling.

    PubMed

    Christensen, Søren T; Clement, Christian A; Satir, Peter; Pedersen, Lotte B

    2012-01-01

    Primary cilia are microtubule-based sensory organelles that coordinate signalling pathways in cell-cycle control, migration, differentiation and other cellular processes critical during development and for tissue homeostasis. Accordingly, defects in assembly or function of primary cilia lead to a plethora of developmental disorders and pathological conditions now known as ciliopathies. In this review, we summarize the current status of the role of primary cilia in coordinating receptor tyrosine kinase (RTK) signalling pathways. Further, we present potential mechanisms of signalling crosstalk and networking in the primary cilium and discuss how defects in ciliary RTK signalling are linked to human diseases and disorders. Copyright © 2011 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  20. Fluid flow due to collective non-reciprocal motion of symmetrically-beating artificial cilia

    PubMed Central

    Khaderi, S. N.; den Toonder, J. M. J.; Onck, P. R.

    2012-01-01

    Using a magneto-mechanical solid-fluid numerical model for permanently magnetic artificial cilia, we show that the metachronal motion of symmetrically beating cilia establishes a net pressure gradient in the direction of the metachronal wave, which creates a unidirectional flow. The flow generated is characterised as a function of the cilia spacing, the length of the metachronal wave, and a dimensionless parameter that characterises the relative importance of the viscous forces over the elastic forces in the cilia. PMID:22662092

  1. Regulation of flagellar motility by the conserved flagellar protein CG34110/Ccdc135/FAP50

    PubMed Central

    Yang, Yong; Cochran, Deborah A.; Gargano, Mary D.; King, Iryna; Samhat, Nayef K.; Burger, Benjain P.; Sabourin, Katherine R.; Hou, Yuqing; Awata, Junya; Parry, David A.D.; Marshall, Wallace F.; Witman, George B.; Lu, Xiangyi

    2011-01-01

    Eukaryotic cilia and flagella are vital sensory and motile organelles. The calcium channel PKD2 mediates sensory perception on cilia and flagella, and defects in this can contribute to ciliopathic diseases. Signaling from Pkd2-dependent Ca2+ rise in the cilium to downstream effectors may require intermediary proteins that are largely unknown. To identify these proteins, we carried out genetic screens for mutations affecting Drosophila melanogaster sperm storage, a process mediated by Drosophila Pkd2. Here we show that a new mutation lost boys (lobo) encodes a conserved flagellar protein CG34110, which corresponds to vertebrate Ccdc135 (E = 6e-78) highly expressed in ciliated respiratory epithelia and sperm, and to FAP50 (E = 1e-28) in the Chlamydomonas reinhardtii flagellar proteome. CG34110 localizes along the fly sperm flagellum. FAP50 is tightly associated with the outer doublet microtubules of the axoneme and appears not to be a component of the central pair, radial spokes, dynein arms, or structures defined by the mbo waveform mutants. Phenotypic analyses indicate that both Pkd2 and lobo specifically affect sperm movement into the female storage receptacle. We hypothesize that the CG34110/Ccdc135/FAP50 family of conserved flagellar proteins functions within the axoneme to mediate Pkd2-dependent processes in the sperm flagellum and other motile cilia. PMID:21289096

  2. A TRPM4-dependent current in murine renal primary cilia

    PubMed Central

    Flannery, Richard J.; Kleene, Steven J.

    2015-01-01

    Defects in primary cilia lead to a variety of human diseases. One of these, polycystic kidney disease, can be caused by defects in a Ca2+-gated ion channel (TRPP2) found on the cilium. Other ciliary functions also contribute to cystogenesis, and defects in apical Ca2+ homeostasis have been implicated. By recording directly from the native cilia of mIMCD-3 cells, a murine cell line of renal epithelial origin, we have identified a second Ca2+-gated channel in the ciliary membrane: the transient receptor potential cation channel, subfamily M, member 4 (TRPM4). In excised primary cilia, TRPM4 was found to have a low sensitivity to Ca2+, with an EC50 of 646 μM at +100 mV. It was inhibited by MgATP and by 9-phenanthrol. The channel was not permeable to Ca2+ or Cl− and had a permeability ratio PK/PNa of 1.42. Reducing the expression of Trpm4 mRNA with short hairpin (sh) RNA reduced the TRPM4 current by 87% and shortened primary cilia by 43%. When phospholipase C was inhibited, the sensitivity to cytoplasmic Ca2+ greatly increased (EC50 = 26 μM at +100 mV), which is consistent with previous reports that phosphatidylinositol 4,5-bisphosphate (PIP2) modulates the channel. MgATP did not restore the channel to a preinactivation state, suggesting that the enzyme or substrate necessary for making PIP2 is not abundant in primary cilia of mIMCD-3 cells. The function of TRPM4 in renal primary cilia is not yet known, but it is likely to influence the apical Ca2+ dynamics of the cell, perhaps in tandem with TRPP2. PMID:26290373

  3. The Role of Cilia in the Pathogenesis of Cystic Kidney Disease

    PubMed Central

    Dell, Katherine M.

    2015-01-01

    Purpose of review Primary (immotile) cilia are specialized organelles present on most cell types. Almost all of proteins associated with a broad spectrum of human cystic kidney diseases have been localized to the region in or around the cilia. Abnormal cilia structure and/or function have been reported in animal models and human cystic kidneys. The goal of this review is to discuss current understanding of the mechanisms by which abnormal genes/proteins and cilia interact to potentially influence renal cystogenesis. Recent findings Novel direct recording of cilia calcium levels/channel activity suggest that cilia form a calcium-mediated signaling microenvironment separate from the cytoplasm, which could provide a mechanism for cilia-specific downstream signaling. Genetic-based studies confirm that cilia are not required for cystogenesis but modulate cystic kidney disease severity through a novel, undefined mechanism. Mechanisms by which both cilia-associated and non-cilia associated proteins can alter cilia structure/function have also been identified. Summary Considerable progress has been made in defining the mechanisms by which abnormal genes and proteins affect cilia structure and function. However, the exact mechanisms by which these interactions cause renal cyst formation and progression of cystic kidney disease are still unknown. PMID:25575298

  4. Primary cilia: a link between hormone signalling and endocrine-related cancers?

    PubMed

    O'Toole, Samuel M; Chapple, J Paul

    2016-10-15

    Primary cilia are sensory organelles that play a role as signalling hubs. Disruption of primary cilia structure and function is increasingly recognised in a range of cancers, with a growing body of evidence suggesting that ciliary disruption contributes to tumourigenesis. This review considers the role of primary cilia in the pathogenesis of endocrine-related cancers.

  5. The functional expression and motile properties of recombinant outer arm dynein from Tetrahymena.

    PubMed

    Edamatsu, Masaki

    2014-05-16

    Cilia and flagella are motile organelles that play various roles in eukaryotic cells. Ciliary movement is driven by axonemal dyneins (outer arm and inner arm dyneins) that bind to peripheral microtubule doublets. Elucidating the molecular mechanism of ciliary movement requires the genetic engineering of axonemal dyneins; however, no expression system for axonemal dyneins has been previously established. This study is the first to purify recombinant axonemal dynein with motile activity. In the ciliated protozoan Tetrahymena, recombinant outer arm dynein purified from ciliary extract was able to slide microtubules in a gliding assay. Furthermore, the recombinant dynein moved processively along microtubules in a single-molecule motility assay. This expression system will be useful for investigating the unique properties of diverse axonemal dyneins and will enable future molecular studies on ciliary movement. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Optimization of bio-inspired multi-segment IPMC cilia

    NASA Astrophysics Data System (ADS)

    Sareh, S.; Conn, A. T.; Rossiter, J. M.; Ieropoulos, I.; Walters, P.

    2010-04-01

    In nature, unidirectional fluid flows are often induced at micro-scales by cilia and related organelles. A controllable unidirectional flow is beneficial at these scales for a range of novel robotic and medical applications, whether the flow is used for propulsion (e.g. swimming robots) or mass transfer (e.g. prosthetic trachea). Ionic Polymer Metal Composites (IPMCs) are innovative smart materials that can be used directly as active propulsive surfaces rather than a traditional motor and propeller. IPMC actuators with two segmented electrodes that attempt to mimic the motion of cilia-like organelles have been realized. In this paper the optimization of these actuators towards producing unidirectional flows is described. A parametric study of the kinematic and hydrodynamic effect of modulating the drive signal has been conducted. As with eukaryotic cilia and flagella found in mammals, the segmented IPMC actuator can generate both flexural (asymmetric) and undulatory (symmetric) motions from the same physical structure. The motion is controlled by applying profiles of driving frequencies and phase differences. Kinematic analysis using a camera and laser displacement sensor has been used to measure and classify different motion types. The hydrodynamic forces produced by each motion type have been estimated using particle-tracking flow visualization. This allows drive signal profiles to be ranked in terms of fluid flow momentum transfer and directionality. Using the results of the parametric study, the IPMC motion is optimized towards producing unidirectional flow via repeatable cilia-inspired motion.

  7. DISC1 Regulates Primary Cilia That Display Specific Dopamine Receptors

    PubMed Central

    Marley, Aaron; von Zastrow, Mark

    2010-01-01

    Background Mutations in the DISC1 gene are strongly associated with major psychiatric syndromes such as schizophrenia. DISC1 encodes a cytoplasmic protein with many potential interaction partners, but its cellular functions remain poorly understood. We identified a role of DISC1 in the cell biology of primary cilia that display disease-relevant dopamine receptors. Methodology/Principal Findings A GFP-tagged DISC1 construct expressed in NIH3T3 cells and rat striatal neurons localized near the base of primary cilia. RNAi-mediated knockdown of endogenous DISC1 resulted in a marked reduction in the number of cells expressing a primary cilium. FLAG-tagged versions of the cloned human D1, D2 and D5 dopamine receptors concentrated highly on the ciliary surface, and this reflects a specific targeting mechanism specific because D3 and D4 receptors localized to the plasma membrane but were not concentrated on cilia. Conclusions/Significance These results identify a role of DISC1 in regulating the formation and/or maintenance of primary cilia, and establish subtype-specific targeting of dopamine receptors to the ciliary surface. Our findings provide new insight to receptor cell biology and suggest a relationship between DISC1 and neural dopamine signaling. PMID:20531939

  8. Model Cilia - Experiments with Biomimetic Actuable Structures and Surfaces

    NASA Astrophysics Data System (ADS)

    Lloyd Carroll, R.

    2005-03-01

    The use of cilia to drive fluid flow is a common motif in living organisms, and in the tissues of higher organisms. By understanding the ways that cilia function (or do not function), potential therapies to treat human diseases (such as cystic fibrosis) may be devised. The complex hydrodynamics of flow in beating ciliary tissues (such as lung epithelial tissues) are challenging to study in cultured tissues, suggesting the need for model systems that will mimic the morphology and beat patterns of living systems. To reach this goal, we have fabricated high aspect ratio cilia-like structures with dimensions similar to those of a lung epithelial cilium (0.2 to 2.0 μm diameter by ˜6 to 10 μm long). The structures and surfaces are composed of a magneto-elastomeric nanocomposite, allowing the actuation of artificial cilia by magnetic fields. We have studied the flexibility of the materials under conditions of flow (in microfluidics channels), and will present theoretical and experimental data from various efforts at actuation. We will discuss details of the fabrication of the ciliated structures and present results of mechanical characterization. The impact of this work on the understanding of fluid flow above ciliated cells and tissues and potential applications of such model systems will also be described.

  9. Ins and outs of GPCR signaling in primary cilia

    PubMed Central

    Schou, Kenneth Bødtker; Pedersen, Lotte Bang; Christensen, Søren Tvorup

    2015-01-01

    Primary cilia are specialized microtubule-based signaling organelles that convey extracellular signals into a cellular response in most vertebrate cell types. The physiological significance of primary cilia is underscored by the fact that defects in assembly or function of these organelles lead to a range of severe diseases and developmental disorders. In most cell types of the human body, signaling by primary cilia involves different G protein-coupled receptors (GPCRs), which transmit specific signals to the cell through G proteins to regulate diverse cellular and physiological events. Here, we provide an overview of GPCR signaling in primary cilia, with main focus on the rhodopsin-like (class A) and the smoothened/frizzled (class F) GPCRs. We describe how such receptors dynamically traffic into and out of the ciliary compartment and how they interact with other classes of ciliary GPCRs, such as class B receptors, to control ciliary function and various physiological and behavioral processes. Finally, we discuss future avenues for developing GPCR-targeted drug strategies for the treatment of ciliopathies. PMID:26297609

  10. Magnetically actuated artificial cilia for optimum mixing performance in microfluidics.

    PubMed

    Chen, Chia-Yuan; Chen, Chia-Yun; Lin, Cheng-Yi; Hu, Ya-Ting

    2013-07-21

    Contemporary lab-chip devices require efficient, high-performance mixing capability. A series of artificial cilia with embedded magnetic particles was fabricated to achieve precise flow manipulation through magnetically driven control. These fabricated structures were actuated in a homogeneous magnetic field generated by a built-in magnetic coil system for various beating cycles inside a microchannel. Three representative trajectories, namely, circular motion, back-and-forth oscillation, and a figure-of-eight pattern, of artificial cilia were designed and generated to mimic the motion of actual cilia. Homogeneous mixing of two highly viscous (>25 centipoise) dyed solutions by using the figure-of-eight trajectory achieved a mixing efficiency of approximately 86%. The underlying relationship between ciliated structures and the induced flow fields was further elucidated by performing a hydrodynamic analysis with micro-particle image velocimetry. In addition, a numerical modeling method which used a fluid structure interaction module was applied to provide quantitative 3D illustrations of induced flow patterns, including vortical structures and vortex core locations. The results reveal that both the magnitude and distribution of induced vortices primarily affect the mixing performance of two viscous flow streams. By using magnetically controlled artificial cilia along with the presented analytical paradigms, a new active flow mixing strategy was suggested to efficiently transport/agitate flows for microfluidics and biomedical applications.

  11. Renal Primary Cilia Lengthen after Acute Tubular Necrosis

    PubMed Central

    Verghese, Elizabeth; Ricardo, Sharon D.; Weidenfeld, Raphael; Zhuang, Junli; Hill, Prudence A.; Langham, Robyn G.

    2009-01-01

    Renal primary cilia are sensory antennas required for the maintenance of normal epithelial differentiation and proliferation in the kidney, but they also have a potential role in epithelial differentiation during renal injury and repair. In mice, tubular damage causes an increase in the length of renal cilia, which may modify their sensory sensitivity during repair. Here, we investigated whether the alteration of renal cilium length during renal injury is clinically relevant. Using biopsies of human renal transplants that suffered acute tubular necrosis during transplantation, we compared the length of renal primary cilia with renal function. Serial biopsies showed that acute tubular necrosis resulted in more than a doubling of cilium length throughout the nephron and collecting duct approximately 1 wk after injury. Allografts displayed a trend toward normalization of cilium length in later biopsies, and this correlated with functional recovery. A mouse model of renal ischemia-reperfusion confirmed the increase and subsequent regression of cilium length during renal repair, displaying complete normalization of cilium length within 6 wk of injury. These findings demonstrate that the length of renal cilia is a clinically relevant indicator of renal injury and repair. PMID:19608704

  12. A coating of passively oscillating flexible cilia to reduce drag

    NASA Astrophysics Data System (ADS)

    Revell, Alistair; Harwood, Adrian; O'Connor, Joseph; Sanchez, Jonathan; Favier, Julien

    2016-11-01

    We present results related to the reduction of wake drag by the coordinated action of a layer of passively oscillating flexible cilia. Inspired by the pop-up of bird feathers, this configuration is shown to self-adapt to the surrounding flow, leading to a stabilization of the wake, a reduction of the mean drag and of lift oscillations. The study is performed using Lattice Boltzmann method, coupled to a recent version of the immersed boundary method. We will present the physical analysis of the coupling between multiple beating cilia and an incoming fluid flow. The modal behaviour of the cilia dynamics will be discussed, as well as their effect on an archetype of unsteady separated boundary layer (first the oscillating channel flow and then the circular cylinder). In the latter case results demonstrate an optimal drag occurs for a particular stiffness, compared to the control case where the same cilia are fixed. It appears that the optimal results are due to a reconfiguration of the elastic coating according to the local vorticity of the flow, and a frequency lock-in, which leads to more stable wake and reduced drag. The structural parameters of the layer will be varied. Results from the PEL-SKIN project: funded by EU Grant #334954.

  13. Drosophila Sperm Motility in the Reproductive Tract1

    PubMed Central

    Yang, Yong; Lu, Xiangyi

    2011-01-01

    Motile cilia and flagella exhibit many waveforms as outputs of dynein activation sequences on the highly conserved axoneme. Motility change of sperm in the reproductive tract is difficult to study and remains an important area of investigation. Sperm typically execute a sinusoidal waveform. Increased viscosity in the medium induces somewhat unusual arc-line and helical waveforms in some sperm. However, whether the latter two waveforms occur in vivo is not known. Using green fluorescence protein imaging, we show that Drosophila sperm in the uterus move in circular foci via arc-line waves, predominantly in a tail-leading orientation. From the uterus, a small fraction of the sperm enters the seminal receptacle (SR) in parallel formations. After sperm storage and coincident with fertilization of the egg, the sperm exit the SR via head-leading helical waves. Consistent with the observed bidirectional movements, the sperm show the ability to propagate both base-to-tip and tip-to-base flagellar waves. Numerous studies have shown that sperm motility is regulated by intraflagellar calcium concentrations; in particular, the Pkd2 calcium channel has been shown to affect sperm storage. Our analyses here suggest that Pkd2 is required for the sperm to adopt the correct waveform and movement orientation during SR entry. A working model for the sperm's SR entry movement is proposed. PMID:21293028

  14. The role of cilia in the pathogenesis of cystic kidney disease.

    PubMed

    Dell, Katherine M

    2015-04-01

    Primary (immotile) cilia are specialized organelles present on most cell types. Almost all of proteins associated with a broad spectrum of human cystic kidney diseases have been localized to the region in or around the cilia. Abnormal cilia structure and function have both been reported in animal models and human cystic kidneys. The goal of this review is to discuss current understanding of the mechanisms by which abnormal genes/proteins and cilia interact to potentially influence renal cystogenesis. Novel direct recording of cilia calcium levels/channel activity suggests that cilia form a calcium-mediated signaling microenvironment separate from the cytoplasm, which could provide a mechanism for cilia-specific downstream signaling. Genetic-based studies confirm that cilia are not required for cystogenesis, but modulate cystic kidney disease severity through a novel, undefined mechanism. Mechanisms by which both cilia-associated and noncilia-associated proteins can alter cilia structure/function have also been identified. Considerable progress has been made in defining the mechanisms by which abnormal genes and proteins affect cilia structure and function. However, the exact mechanisms by which these interactions cause renal cyst formation and progression of cystic kidney disease are still unknown.

  15. Function and regulation of primary cilia and intraflagellar transport proteins in the skeleton

    PubMed Central

    Yuan, Xue; Serra, Rosa A.; Yang, Shuying

    2014-01-01

    Primary cilia are microtubule-based organelles that project from the cell surface to enable transduction of various developmental signaling pathways. The process of intraflagellar transport (IFT) is crucial for the building and maintenance of primary cilia. Ciliary dysfunction has been found in a range of disorders called ciliopathies, some of which display severe skeletal dysplasias. In recent years, interest has grown in uncovering the function of primary cilia/IFT proteins in bone development, mechanotransduction, and cellular regulation. We summarize recent advances in understanding the function of cilia and IFT proteins in the regulation of cell differentiation in osteoblasts, osteocytes, chondrocytes, and mesenchymal stem cells (MSCs). We also discuss the mechanosensory function of cilia and IFT proteins in bone cells, cilia orientation, and other functions of cilia in chondrocytes. PMID:24961486

  16. Cilia/Ift protein and motor -related bone diseases and mouse models.

    PubMed

    Yuan, Xue; Yang, Shuying

    2015-01-01

    Primary cilia are essential cellular organelles projecting from the cell surface to sense and transduce developmental signaling. They are tiny but have complicated structures containing microtubule (MT)-based internal structures (the axoneme) and mother centriole formed basal body. Intraflagellar transport (Ift) operated by Ift proteins and motors are indispensable for cilia formation and function. Mutations in Ift proteins or Ift motors cause various human diseases, some of which have severe bone defects. Over the last few decades, major advances have occurred in understanding the roles of these proteins and cilia in bone development and remodeling by examining cilia/Ift protein-related human diseases and establishing mouse transgenic models. In this review, we describe current advances in the understanding of the cilia/Ift structure and function. We further summarize cilia/Ift-related human diseases and current mouse models with an emphasis on bone-related phenotypes, cilia morphology, and signaling pathways.

  17. Cilia/Ift protein and motor-related bone diseases and mouse models

    PubMed Central

    Yuan, Xue; Yang, Shuying

    2015-01-01

    Primary cilia are essential cellular organelles projecting from the cell surface to sense and transduce developmental signaling. They are tiny but have complicated structures containing microtubule (MT)-based internal structures (the axoneme) and mother centriole formed basal body. Intraflagellar transport (Ift) operated by Ift proteins and motors are indispensable for cilia formation and function. Mutations in Ift proteins or Ift motors cause various human diseases, some of which have severe bone defects. Over the last few decades, major advances have occurred in understanding the roles of these proteins and cilia in bone development and remodeling by examining cilia/Ift protein-related human diseases and establishing mouse transgenic models. In this review, we describe current advances in the understanding of the cilia/Ift structure and function. We further summarize cilia/Ift-related human diseases and current mouse models with an emphasis on bone-related phenotypes, cilia morphology, and signaling pathways. PMID:25553465

  18. Function and regulation of primary cilia and intraflagellar transport proteins in the skeleton.

    PubMed

    Yuan, Xue; Serra, Rosa A; Yang, Shuying

    2015-01-01

    Primary cilia are microtubule-based organelles that project from the cell surface to enable transduction of various developmental signaling pathways. The process of intraflagellar transport (IFT) is crucial for the building and maintenance of primary cilia. Ciliary dysfunction has been found in a range of disorders called ciliopathies, some of which display severe skeletal dysplasias. In recent years, interest has grown in uncovering the function of primary cilia/IFT proteins in bone development, mechanotransduction, and cellular regulation. We summarize recent advances in understanding the function of cilia and IFT proteins in the regulation of cell differentiation in osteoblasts, osteocytes, chondrocytes, and mesenchymal stem cells (MSCs). We also discuss the mechanosensory function of cilia and IFT proteins in bone cells, cilia orientation, and other functions of cilia in chondrocytes.

  19. Chemically inducible diffusion trap at cilia reveals molecular sieve-like barrier.

    PubMed

    Lin, Yu-Chun; Niewiadomski, Pawel; Lin, Benjamin; Nakamura, Hideki; Phua, Siew Cheng; Jiao, John; Levchenko, Andre; Inoue, Takafumi; Rohatgi, Rajat; Inoue, Takanari

    2013-07-01

    Primary cilia function as specialized compartments for signal transduction. The stereotyped structure and signaling function of cilia inextricably depend on the selective segregation of molecules in cilia. However, the fundamental principles governing the access of soluble proteins to primary cilia remain unresolved. We developed a methodology termed 'chemically inducible diffusion trap at cilia' to visualize the diffusion process of a series of fluorescent proteins ranging in size from 3.2 nm to 7.9 nm into primary cilia. We found that the interior of the cilium was accessible to proteins as large as 7.9 nm. The kinetics of ciliary accumulation of this panel of proteins was exponentially limited by their Stokes radii. Quantitative modeling suggests that the diffusion barrier operates as a molecular sieve at the base of cilia. Our study presents a set of powerful, generally applicable tools for the quantitative monitoring of ciliary protein diffusion under both physiological and pathological conditions.

  20. Sperm Motility in Flow

    NASA Astrophysics Data System (ADS)

    Guasto, Jeffrey; Juarez, Gabriel; Stocker, Roman

    2012-11-01

    A wide variety of plants and animals reproduce sexually by releasing motile sperm that seek out a conspecific egg, for example in the reproductive tract for mammals or in the water column for externally fertilizing organisms. Sperm are aided in their quest by chemical cues, but must also contend with hydrodynamic forces, resulting from laminar flows in reproductive tracts or turbulence in aquatic habitats. To understand how velocity gradients affect motility, we subjected swimming sperm to a range of highly-controlled straining flows using a cross-flow microfluidic device. The motion of the cell body and flagellum were captured through high-speed video microscopy. The effects of flow on swimming are twofold. For moderate velocity gradients, flow simply advects and reorients cells, quenching their ability to cross streamlines. For high velocity gradients, fluid stresses hinder the internal bending of the flagellum, directly inhibiting motility. The transition between the two regimes is governed by the Sperm number, which compares the external viscous stresses with the internal elastic stresses. Ultimately, unraveling the role of flow in sperm motility will lead to a better understanding of population dynamics among aquatic organisms and infertility problems in humans.

  1. Mammalian pheromones.

    PubMed

    Liberles, Stephen D

    2014-01-01

    Mammalian pheromones control a myriad of innate social behaviors and acutely regulate hormone levels. Responses to pheromones are highly robust, reproducible, and stereotyped and likely involve developmentally predetermined neural circuits. Here, I review several facets of pheromone transduction in mammals, including (a) chemosensory receptors and signaling components of the main olfactory epithelium and vomeronasal organ involved in pheromone detection; (b) pheromone-activated neural circuits subject to sex-specific and state-dependent modulation; and (c) the striking chemical diversity of mammalian pheromones, which range from small, volatile molecules and sulfated steroids to large families of proteins. Finally, I review (d) molecular mechanisms underlying various behavioral and endocrine responses, including modulation of puberty and estrous; control of reproduction, aggression, suckling, and parental behaviors; individual recognition; and distinguishing of own species from predators, competitors, and prey. Deconstruction of pheromone transduction mechanisms provides a critical foundation for understanding how odor response pathways generate instinctive behaviors.

  2. Mammalian Pheromones

    PubMed Central

    Liberles, Stephen D.

    2015-01-01

    Mammalian pheromones control a myriad of innate social behaviors and acutely regulate hormone levels. Responses to pheromones are highly robust, reproducible, and stereotyped and likely involve developmentally predetermined neural circuits. Here, I review several facets of pheromone transduction in mammals, including (a) chemosensory receptors and signaling components of the main olfactory epithelium and vomeronasal organ involved in pheromone detection; (b) pheromone-activated neural circuits subject to sex-specific and state-dependent modulation; and (c) the striking chemical diversity of mammalian pheromones, which range from small, volatile molecules and sulfated steroids to large families of proteins. Finally, I review (d ) molecular mechanisms underlying various behavioral and endocrine responses, including modulation of puberty and estrous; control of reproduction, aggression, suckling, and parental behaviors; individual recognition; and distinguishing of own species from predators, competitors, and prey. Deconstruction of pheromone transduction mechanisms provides a critical foundation for understanding how odor response pathways generate instinctive behaviors. PMID:23988175

  3. Defining motility in the Staphylococci.

    PubMed

    Pollitt, Eric J G; Diggle, Stephen P

    2017-08-01

    The ability of bacteria to move is critical for their survival in diverse environments and multiple ways have evolved to achieve this. Two forms of motility have recently been described for Staphylococcus aureus, an organism previously considered to be non-motile. One form is called spreading, which is a type of sliding motility and the second form involves comet formation, which has many observable characteristics associated with gliding motility. Darting motility has also been observed in Staphylococcus epidermidis. This review describes how motility is defined and how we distinguish between passive and active motility. We discuss the characteristics of the various forms of Staphylococci motility, the molecular mechanisms involved and the potential future research directions.

  4. Hypomorphic CEP290/NPHP6 mutations result in anosmia caused by the selective loss of G proteins in cilia of olfactory sensory neurons

    PubMed Central

    McEwen, Dyke P.; Koenekoop, Robert K.; Khanna, Hemant; Jenkins, Paul M.; Lopez, Irma; Swaroop, Anand; Martens, Jeffrey R.

    2007-01-01

    Cilia regulate diverse functions such as motility, fluid balance, and sensory perception. The cilia of olfactory sensory neurons (OSNs) compartmentalize the signaling proteins necessary for odor detection; however, little is known regarding the mechanisms of protein sorting/entry into olfactory cilia. Nephrocystins are a family of ciliary proteins likely involved in cargo sorting during transport from the basal body to the ciliary axoneme. In humans, loss-of-function of the cilia–centrosomal protein CEP290/NPHP6 is associated with Joubert and Meckel syndromes, whereas hypomorphic mutations result in Leber congenital amaurosis (LCA), a form of early-onset retinal dystrophy. Here, we report that CEP290–LCA patients exhibit severely abnormal olfactory function. In a mouse model with hypomorphic mutations in CEP290 [retinal dystrophy-16 mice (rd16)], electro-olfactogram recordings revealed an anosmic phenotype analogous to that of CEP290–LCA patients. Despite the loss of olfactory function, cilia of OSNs remained intact in the rd16 mice. As in wild type, CEP290 localized to dendritic knobs of rd16 OSNs, where it was in complex with ciliary transport proteins and the olfactory G proteins Golf and Gγ13. Interestingly, we observed defective ciliary localization of Golf and Gγ13 but not of G protein-coupled odorant receptors or other components of the odorant signaling pathway in the rd16 OSNs. Our data implicate distinct mechanisms for ciliary transport of olfactory signaling proteins, with CEP290 being a key mediator involved in G protein trafficking. The assessment of olfactory function can, therefore, serve as a useful diagnostic tool for genetic screening of certain syndromic ciliary diseases. PMID:17898177

  5. Why motor proteins team up - Intraflagellar transport in C. elegans cilia.

    PubMed

    Mijalkovic, Jona; Prevo, Bram; Peterman, Erwin J G

    2016-01-01

    Inside the cell, vital processes such as cell division and intracellular transport are driven by the concerted action of different molecular motor proteins. In C. elegans chemosensory cilia, 2 kinesin-2 family motor proteins, kinesin-II and OSM-3, team up to drive intraflagellar transport (IFT) in the anterograde direction, from base to tip, whereas IFT dynein hitchhikes toward the tip and subsequently drives IFT in the opposite, retrograde direction, thereby recycling both kinesins. While it is evident that at least a retrograde and an anterograde motor are necessary to drive IFT, it has remained puzzling why 2 same-polarity kinesins are employed. Recently, we addressed this question by combining advanced genome-engineering tools with ultrasensitive, quantitative fluorescence microscopy to study IFT with single-molecule sensitivity.(1,2) Using this combination of approaches, we uncovered a differentiation in kinesin-2 function, in which the slower kinesin-II operates as an 'importer', loading IFT trains into the cilium before gradually handing them over to the faster OSM-3. OSM-3 subsequently acts as a long-range 'transporter', driving the IFT trains toward the tip. The two kinesin-2 motors combine their unique motility properties to achieve something neither motor can achieve on its own; that is to optimize the amount of cargo inside the cilium. In this commentary, we provide detailed insight into the rationale behind our research approach and comment on our recent findings. Moreover, we discuss the role of IFT dynein and provide an outlook on future studies.

  6. Modeling collective cell motility

    NASA Astrophysics Data System (ADS)

    Rappel, Wouter-Jan

    Eukaryotic cells often move in groups, a critical aspect of many biological and medical processes including wound healing, morphogenesis and cancer metastasis. Modeling can provide useful insights into the fundamental mechanisms of collective cell motility. Constructing models that incorporate the physical properties of the cells, however, is challenging. Here, I discuss our efforts to build a comprehensive cell motility model that includes cell membrane properties, cell-substrate interactions, cell polarity, and cell-cell interaction. The model will be applied to a variety of systems, including motion on micropatterned substrates and the migration of border cells in Drosophila. This work was supported by NIH Grant No. P01 GM078586 and NSF Grant No. 1068869.

  7. Motility of Mollicutes

    NASA Astrophysics Data System (ADS)

    Wolgemuth, Charles; Igoshin, Oleg; Oster, George

    2003-03-01

    Recent experiments show that the conformation of filament proteins play a role in the motility and morphology of many different types of bacteria. Conformational changes in the protein subunits may produce forces to drive propulsion and cell division. Here we present a molecular mechanism by which these forces can drive cell motion. Coupling of a biochemical cycle, such as ATP hydrolysis, to the dynamics of elastic filaments enable elastic filaments to propagate deformations that generate propulsive forces. We demonstrate this possibility for two classes of wall-less bacteria called mollicutes: the swimming of helical shaped Spiroplasma, and the gliding motility of Mycoplasma. Similar mechanisms may explain the locomotion of other prokaryotes, including the swimming of Synechococcus and the gliding of some myxobacteria.

  8. NCAM regulates cell motility.

    PubMed

    Prag, Søren; Lepekhin, Eugene A; Kolkova, Kateryna; Hartmann-Petersen, Rasmus; Kawa, Anna; Walmod, Peter S; Belman, Vadym; Gallagher, Helen C; Berezin, Vladimir; Bock, Elisabeth; Pedersen, Nina

    2002-01-15

    Cell migration is required during development of the nervous system. The regulatory mechanisms for this process, however, are poorly elucidated. We show here that expression of or exposure to the neural cell adhesion molecule (NCAM) strongly affected the motile behaviour of glioma cells independently of homophilic NCAM interactions. Expression of the transmembrane 140 kDa isoform of NCAM (NCAM-140) caused a significant reduction in cellular motility, probably through interference with factors regulating cellular attachment, as NCAM-140-expressing cells exhibited a decreased attachment to a fibronectin substratum compared with NCAM-negative cells. Ectopic expression of the cytoplasmic part of NCAM-140 also inhibited cell motility, presumably via the non-receptor tyrosine kinase p59(fyn) with which NCAM-140 interacts. Furthermore, we showed that the extracellular part of NCAM acted as a paracrine inhibitor of NCAM-negative cell locomotion through a heterophilic interaction with a cell-surface receptor. As we showed that the two N-terminal immunoglobulin modules of NCAM, which are known to bind to heparin, were responsible for this inhibition, we presume that this receptor is a heparan sulfate proteoglycan. A model for the inhibitory effect of NCAM is proposed, which involves competition between NCAM and extracellular components for the binding to membrane-associated heparan sulfate proteoglycan.

  9. Motility of Mycoplasma pneumoniae.

    PubMed Central

    Radestock, U; Bredt, W

    1977-01-01

    Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover. Images PMID:14925

  10. CILIA FORMATION IN THE ADULT CAT BRAIN AFTER PARGYLINE TREATMENT

    PubMed Central

    Milhaud, Monique; Pappas, George D.

    1968-01-01

    The brains of four adult cats treated with pargyline (a nonhydrazide monoaminoxidase inhibitor) were examined at both the light and electron microscopic levels. Formation of typical mature cilia with the 9 + 2 pattern was observed in neural cells in the following areas: habenula nuclei, interpeduncular nuclei, hippocampus, mammillary bodies, thalamus, and caudate nucleus. The most marked ciliation occurs in the habenula nuclei. In general, glial cells greatly predominate in the formation of cilia. It is not clear whether ciliation in the central nervous system is the direct result of pargyline or if it occurs indirectly as a result of inhibition of monoaminoxidase. These findings are compared with the serotonin effect on ciliation in the embryogenesis of lower forms. It is suggested that pharmacological stimulation of centriolar reproduction without subsequent mitosis may lead to ciliary formation. PMID:11905194

  11. Planar cell polarity effector gene Intu regulates cell fate-specific differentiation of keratinocytes through the primary cilia.

    PubMed

    Dai, D; Li, L; Huebner, A; Zeng, H; Guevara, E; Claypool, D J; Liu, A; Chen, J

    2013-01-01

    Genes involved in the planar cell polarity (PCP) signaling pathway are essential for a number of developmental processes in mammals, such as convergent extension and ciliogenesis. Tissue-specific PCP effector genes of the PCP signaling pathway are believed to mediate PCP signals in a tissue- and cell type-specific manner. However, how PCP signaling controls the morphogenesis of mammalian tissues remains unclear. In this study, we investigated the role of inturned (Intu), a tissue-specific PCP effector gene, during hair follicle formation in mice. Tissue-specific disruption of Intu in embryonic epidermis resulted in hair follicle morphogenesis arrest because of the failure of follicular keratinocyte to differentiate. Targeting Intu in the epidermis resulted in almost complete loss of primary cilia in epidermal and follicular keratinocytes, and a suppressed hedgehog signaling pathway. Surprisingly, the epidermal stratification and differentiation programs and barrier function were not affected. These results demonstrate that tissue-specific PCP effector genes of the PCP signaling pathway control the differentiation of keratinocytes through the primary cilia in a cell fate- and context-dependent manner, which may be critical in orchestrating the propagation and interpretation of polarity signals established by the core PCP components.

  12. Voltage-gated calcium channels of Paramecium cilia.

    PubMed

    Lodh, Sukanya; Yano, Junji; Valentine, Megan S; Van Houten, Judith L

    2016-10-01

    Paramecium cells swim by beating their cilia, and make turns by transiently reversing their power stroke. Reversal is caused by Ca(2+) entering the cilium through voltage-gated Ca(2+) (CaV) channels that are found exclusively in the cilia. As ciliary Ca(2+) levels return to normal, the cell pivots and swims forward in a new direction. Thus, the activation of the CaV channels causes cells to make a turn in their swimming paths. For 45 years, the physiological characteristics of the Paramecium ciliary CaV channels have been known, but the proteins were not identified until recently, when the P. tetraurelia ciliary membrane proteome was determined. Three CaVα1 subunits that were identified among the proteins were cloned and confirmed to be expressed in the cilia. We demonstrate using RNA interference that these channels function as the ciliary CaV channels that are responsible for the reversal of ciliary beating. Furthermore, we show that Pawn (pw) mutants of Paramecium that cannot swim backward for lack of CaV channel activity do not express any of the three CaV1 channels in their ciliary membrane, until they are rescued from the mutant phenotype by expression of the wild-type PW gene. These results reinforce the correlation of the three CaV channels with backward swimming through ciliary reversal. The PwB protein, found in endoplasmic reticulum fractions, co-immunoprecipitates with the CaV1c channel and perhaps functions in trafficking. The PwA protein does not appear to have an interaction with the channel proteins but affects their appearance in the cilia. © 2016. Published by The Company of Biologists Ltd.

  13. Voltage-gated calcium channels of Paramecium cilia

    PubMed Central

    Lodh, Sukanya; Valentine, Megan S.; Van Houten, Judith L.

    2016-01-01

    ABSTRACT Paramecium cells swim by beating their cilia, and make turns by transiently reversing their power stroke. Reversal is caused by Ca2+ entering the cilium through voltage-gated Ca2+ (CaV) channels that are found exclusively in the cilia. As ciliary Ca2+ levels return to normal, the cell pivots and swims forward in a new direction. Thus, the activation of the CaV channels causes cells to make a turn in their swimming paths. For 45 years, the physiological characteristics of the Paramecium ciliary CaV channels have been known, but the proteins were not identified until recently, when the P. tetraurelia ciliary membrane proteome was determined. Three CaVα1 subunits that were identified among the proteins were cloned and confirmed to be expressed in the cilia. We demonstrate using RNA interference that these channels function as the ciliary CaV channels that are responsible for the reversal of ciliary beating. Furthermore, we show that Pawn (pw) mutants of Paramecium that cannot swim backward for lack of CaV channel activity do not express any of the three CaV1 channels in their ciliary membrane, until they are rescued from the mutant phenotype by expression of the wild-type PW gene. These results reinforce the correlation of the three CaV channels with backward swimming through ciliary reversal. The PwB protein, found in endoplasmic reticulum fractions, co-immunoprecipitates with the CaV1c channel and perhaps functions in trafficking. The PwA protein does not appear to have an interaction with the channel proteins but affects their appearance in the cilia. PMID:27707864

  14. Artificial Muscle (AM) Cilia Array for Underwater Systems

    DTIC Science & Technology

    2016-12-15

    Composite (IPMC) 4 P.R. Bandyopadhyay and J.C. Hansen, ’’Breakup and then makeup: a predictive model of how cilia self-regulate hardness for posture...groups in the core layers of the composite which have not been metalized. Table 2 describes the initial cleaning process. The Nation was abraded with...This causes platinum to be deposited on the surface of the Nafion. Following this the resulting composite was removed and the resistances across the

  15. Artificial Muscle (AM) Cilia Array for Underwater Systems

    DTIC Science & Technology

    2016-12-15

    Metal Composite (IPMC) 4 P. R. Bandyopadhyay and J. C. Hansen, "Breakup and then makeup: a pred ictive model of how cilia self-regulate hardness for...sulfonyl end groups in the core layers of the composite which have not been metalized. Table 2 describes the initial cleaning process. The Nation was...Nation. Following this the resulting composite was removed and the resistances across the metalized surfaces were checked. For good conductivity

  16. Nature-inspired micro-fluidic manipulation using artificial cilia

    NASA Astrophysics Data System (ADS)

    den Toonder, Jaap; de Goede, Judith; Khatavkar, Vinayak; Anderson, Patrick

    2006-11-01

    One particular micro-fluidics manipulation mechanism ``designed'' by nature is that due to a covering of beating cilia over the external surface of micro-organisms (e.g. Paramecium). A cilium can be viewed as a small hair or flexible rod (in protozoa: typical length 10 μm and diameter 0.1 μm) which is attached to the surface. We have developed polymer micro-actuators, made with standard micro-technology processing, which respond to an applied electrical or magnetic field by changing their shape. The shape and size of the polymer actuators mimics that of cilia occurring in nature. We have shown experimentally that, indeed, our artificial cilia can induce significant flow velocities of at least 75 μm/s in a fluid with a viscosity of 10 mPas. In this paper we will give an overview of our activities in developing the polymer actuators and the corresponding technology, show experimental and numerical fluid flow results, and finally assess the feasibility of applying this new and attractive micro-fluidic actuation method in functional biosensors.

  17. Nanoscale Fluidics: Using magnetic nanorods as model cilia

    NASA Astrophysics Data System (ADS)

    Hao, Jing; Ben, Wilde; Jeremy, Cribb; Chris, Dwyer; Jay, Fisher; Kalpit, Desai; Leandra, Vicci; Russell, M. Taylor, II; Richard, Superfine

    2003-11-01

    The beating of cilia and flagella, slender cylinders 250 nanometers in diameter with lengths from 7 to 50 microns, is ubiquitous in biology. The fluid dynamics produced by the cilia or flagella motion is responsible for organism feeding, propulsion, for bacterial clearance in the lungs and for the right-left asymmetry in vertebrates. We are developing a model system for cilia beating through the use of magnetic nanorods. Using anodized aluminum oxide (AAO) membranes as templates, magnetic rods of permalloy with a diameter of 100 and 200 nm have been fabricated. We will describe the details of fabrication and characterization, and discuss methods used to study the hydrodynamic behavior of these nanorods in aqueous solutions under applied magnetic fields. Movies of these nanorods in oscillating 3-D magnetic fields generated by our 3-dimensional force microscopy (3DFM) clearly show bead motion in vortices around the nanorod. Deliberately transporting beads near the rods by laser trap, we can reproducibly study the hydrodynamic behavior around the nanorods and the influence of local rheological properties.

  18. A high-fat diet regulates gastrin and acid secretion through primary cilia.

    PubMed

    Saqui-Salces, Milena; Dowdle, William E; Reiter, Jeremy F; Merchant, Juanita L

    2012-08-01

    The role of primary cilia in the gastrointestinal tract has not been examined. Here we report the presence of primary cilia on gastric endocrine cells producing gastrin, ghrelin, and somatostatin (Sst), hormones regulated by food intake. During eating, cilia in the gastric antrum decreased, whereas gastric acid and circulating gastrin increased. Mice fed high-fat chow showed a delayed decrease in antral cilia, increased plasma gastrin, and gastric acidity. Mice fed high-fat chow for 3 wk showed lower cilia numbers and acid but higher gastrin levels than mice fed a standard diet, suggesting that fat affects gastric physiology. Ex vivo experiments showed that cilia in the corpus responded to acid and distension, whereas cilia in the antrum responded to food. To analyze the role of gastric cilia, we conditionally deleted the intraflagellar transport protein Ift88 (Ift88(-/fl)). In fed Ift88(-/fl) mice, gastrin levels were higher, and gastric acidity was lower. Moreover, gastrin and Sst gene expression did not change in response to food as in controls. At 8 mo, Ift88(-/fl) mice developed foveolar hyperplasia, hypergastrinemia, and hypochlorhydria associated with endocrine dysfunction. Our results show that components of food (fat) are sensed by antral cilia on endocrine cells, which modulates gastrin secretion and gastric acidity.

  19. A facile template-free approach to magnetodriven, multifunctional artificial cilia.

    PubMed

    Timonen, Jaakko V I; Johans, Christoffer; Kontturi, Kyösti; Walther, Andreas; Ikkala, Olli; Ras, Robin H A

    2010-08-01

    Flexible and magnetic artificial cilia were grown on various substrates by a facile bottom-up approach based on template-free magnetic assembly. The magnetic cilia formed spontaneously from a suspension of micrometer-sized ferromagnetic particles and elastomeric polymer in a liquid solvent when dried in an external magnetic field. The cilia mimics were mechanically stable even in the absence of an external magnetic field and a solvent due to the polymer, which acted as "glue" holding the particles together and connecting the cilia to the substrate. The length of the magnetic cilia was in the millimeter range, that is, two to three orders of magnitude times the length of typical biological cilia. The aspect ratio reached values over 100 and was tunable with the magnetic field gradient and the size of the ferromagnetic particles. The cilia mimics responded to an external magnetic field by reversibly bending along the field. The bending actuation was sufficiently powerful to allow two functions: to translate macroscopic nonmagnetic objects placed over the cilia mimics and to mix liquids of even high viscosity. The mechanical properties of the magnetic cilia could be easily tuned by changing the impregnating polymer. The particularly simple template-free construction and fixation on various surfaces suggest applications as an externally controllable surface.

  20. Dynamic Remodeling of Membrane Composition Drives Cell Cycle through Primary Cilia Excision.

    PubMed

    Phua, Siew Cheng; Chiba, Shuhei; Suzuki, Masako; Su, Emily; Roberson, Elle C; Pusapati, Ganesh V; Setou, Mitsutoshi; Rohatgi, Rajat; Reiter, Jeremy F; Ikegami, Koji; Inoue, Takanari

    2017-01-12

    The life cycle of a primary cilium begins in quiescence and ends prior to mitosis. In quiescent cells, the primary cilium insulates itself from contiguous dynamic membrane processes on the cell surface to function as a stable signaling apparatus. Here, we demonstrate that basal restriction of ciliary structure dynamics is established by the cilia-enriched phosphoinositide 5-phosphatase, Inpp5e. Growth induction displaces ciliary Inpp5e and accumulates phosphatidylinositol 4,5-bisphosphate in distal cilia. This change triggers otherwise-forbidden actin polymerization in primary cilia, which excises cilia tips in a process we call cilia decapitation. While cilia disassembly is traditionally thought to occur solely through resorption, we show that an acute loss of IFT-B through cilia decapitation precedes resorption. Finally, we propose that cilia decapitation induces mitogenic signaling and constitutes a molecular link between the cilia life cycle and cell-division cycle. This newly defined ciliary mechanism may find significance in cell proliferation control during normal development and cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. The motility of axonemal dynein is regulated by the tubulin code.

    PubMed

    Alper, Joshua D; Decker, Franziska; Agana, Bernice; Howard, Jonathon

    2014-12-16

    Microtubule diversity, arising from the utilization of different tubulin genes and from posttranslational modifications, regulates many cellular processes including cell division, neuronal differentiation and growth, and centriole assembly. In the case of cilia and flagella, multiple cell biological studies show that microtubule diversity is important for axonemal assembly and motility. However, it is not known whether microtubule diversity directly influences the activity of the axonemal dyneins, the motors that drive the beating of the axoneme, nor whether the effects on motility are indirect, perhaps through regulatory pathways upstream of the motors, such as the central pair, radial spokes, or dynein regulatory complex. To test whether microtubule diversity can directly regulate the activity of axonemal dyneins, we asked whether in vitro acetylation or deacetylation of lysine 40 (K40), a major posttranslational modification of α-tubulin, or whether proteolytic cleavage of the C-terminal tail (CTT) of α- and β-tubulin, the location of detyrosination, polyglutamylation, and polyglycylation modifications as well as most of the genetic diversity, can influence the activity of outer arm axonemal dynein in motility assays using purified proteins. By quantifying the motility with displacement-weighted velocity analysis and mathematically modeling the results, we found that K40 acetylation increases and CTTs decrease axonemal dynein motility. These results show that axonemal dynein directly deciphers the tubulin code, which has important implications for eukaryotic ciliary beat regulation.

  2. The Motility of Axonemal Dynein Is Regulated by the Tubulin Code

    PubMed Central

    Alper, Joshua D.; Decker, Franziska; Agana, Bernice; Howard, Jonathon

    2014-01-01

    Microtubule diversity, arising from the utilization of different tubulin genes and from posttranslational modifications, regulates many cellular processes including cell division, neuronal differentiation and growth, and centriole assembly. In the case of cilia and flagella, multiple cell biological studies show that microtubule diversity is important for axonemal assembly and motility. However, it is not known whether microtubule diversity directly influences the activity of the axonemal dyneins, the motors that drive the beating of the axoneme, nor whether the effects on motility are indirect, perhaps through regulatory pathways upstream of the motors, such as the central pair, radial spokes, or dynein regulatory complex. To test whether microtubule diversity can directly regulate the activity of axonemal dyneins, we asked whether in vitro acetylation or deacetylation of lysine 40 (K40), a major posttranslational modification of α-tubulin, or whether proteolytic cleavage of the C-terminal tail (CTT) of α- and β-tubulin, the location of detyrosination, polyglutamylation, and polyglycylation modifications as well as most of the genetic diversity, can influence the activity of outer arm axonemal dynein in motility assays using purified proteins. By quantifying the motility with displacement-weighted velocity analysis and mathematically modeling the results, we found that K40 acetylation increases and CTTs decrease axonemal dynein motility. These results show that axonemal dynein directly deciphers the tubulin code, which has important implications for eukaryotic ciliary beat regulation. PMID:25658008

  3. Morphogenesis of respiratory syncytial virus in human primary nasal ciliated epithelial cells occurs at surface membrane microdomains that are distinct from cilia.

    PubMed

    Jumat, Muhammad Raihan; Yan, Yan; Ravi, Laxmi Iyer; Wong, Puisan; Huong, Tra Nguyen; Li, Chunwei; Tan, Boon Huan; Wang, De Yun; Sugrue, Richard J

    2015-10-01

    The distribution of cilia and the respiratory syncytial virus (RSV) nucleocapsid (N) protein, fusion (F) protein, attachment (G) protein, and M2-1 protein in human ciliated nasal epithelial cells was examined at between 1 and 5 days post-infection (dpi). All virus structural proteins were localized at cell surface projections that were distinct from cilia. The F protein was also trafficked into the cilia, and while its presence increased as the infection proceeded, the N protein was not detected in the cilia at any time of infection. The presence of the F protein in the cilia correlated with cellular changes in the cilia and reduced cilia function. At 5dpi extensive cilia loss and further reduced cilia function was noted. These data suggested that although RSV morphogenesis occurs at non-cilia locations on ciliated nasal epithelial cells, RSV infection induces changes in the cilia body that leads to extensive cilia loss.

  4. MOTILE MARINE BACTERIA I.

    PubMed Central

    Leifson, Einar; Cosenza, B. J.; Murchelano, R.; Cleverdon, R. C.

    1964-01-01

    Leifson, Einar (Loyola University, Chicago, Ill.), B. J. Cosenza, R. Murchelano, and R. C. Cleverdon. Motile marine bacteria. I. Techniques, ecology, and general characteristics. J. Bacteriol. 87:652–666. 1964.—Aerobic, heterotrophic bacteria were isolated from the waters of the Long Island Sound, Narragansett Bay, Atlantic Ocean, and from the intestine of a variety of marine animals found along the shore of the Long Island Sound. A total of about 600 cultures of motile bacteria were studied morphologically and physiologically, with special emphasis on flagellar characteristics. The great majority of the bacteria isolated from the water were polar flagellate, nonfermentative, nonpigmented, and gramnegative. Most of these were straight, capsulated rods, but a considerable number were curved like vibrios. Yellow-pigmented isolates were often nonmotile, and the motile forms were most frequently subpolar flagellate. Several rosette-forming bacteria, including Caulobacter species, were isolated. Two typical spirilla and one flagellated coccus were found. Peritrichous flagellate bacteria, both gram-positive and gram-negative, were rare except in bottom mud. The normal intestinal flora of marine animals, such as fish and shellfish, consisted of polar flagellate, fermentative, non-pigmented, gram-negative, straight rods. Curved forms, like vibrios, were less common. Polar multitrichous flagellate forms were not uncommon and included all the luminescent types isolated. A considerable proportion of the polar monotrichous flagellate rods swarmed over the surface of agar media. When grown on solid media, all of these showed mixed polar and lateral flagellation; in liquid media, mainly polar flagellation was found. The ecology and general taxonomy of marine bacteria are discussed. Images PMID:14129669

  5. Actin-Based Motility of Intracellular Microbial Pathogens

    PubMed Central

    Goldberg, Marcia B.

    2001-01-01

    A diverse group of intracellular microorganisms, including Listeria monocytogenes, Shigella spp., Rickettsia spp., and vaccinia virus, utilize actin-based motility to move within and spread between mammalian host cells. These organisms have in common a pathogenic life cycle that involves a stage within the cytoplasm of mammalian host cells. Within the cytoplasm of host cells, these organisms activate components of the cellular actin assembly machinery to induce the formation of actin tails on the microbial surface. The assembly of these actin tails provides force that propels the organisms through the cell cytoplasm to the cell periphery or into adjacent cells. Each of these organisms utilizes preexisting mammalian pathways of actin rearrangement to induce its own actin-based motility. Particularly remarkable is that while all of these microbes use the same or overlapping pathways, each intercepts the pathway at a different step. In addition, the microbial molecules involved are each distinctly different from the others. Taken together, these observations suggest that each of these microbes separately and convergently evolved a mechanism to utilize the cellular actin assembly machinery. The current understanding of the molecular mechanisms of microbial actin-based motility is the subject of this review. PMID:11729265

  6. Symmetry-Breaking Motility

    NASA Astrophysics Data System (ADS)

    Lee, Allen; Lee, Ha Youn; Kardar, Mehran

    2005-09-01

    Locomotion of bacteria by actin polymerization and in vitro motion of spherical beads coated with a protein catalyzing polymerization are examples of active motility. Starting from a simple model of forces locally normal to the surface of a bead, we construct a phenomenological equation for its motion. The singularities at a continuous transition between moving and stationary beads are shown to be related to the symmetries of its shape. Universal features of the phase behavior are calculated analytically and confirmed by simulations. Fluctuations in velocity are shown to be generically non-Maxwellian and correlated to the shape of the bead.

  7. Cellular mechanics and motility

    NASA Astrophysics Data System (ADS)

    Hénon, Sylvie; Sykes, Cécile

    2015-10-01

    The term motility defines the movement of a living organism. One widely known example is the motility of sperm cells, or the one of flagellar bacteria. The propulsive element of such organisms is a cilium(or flagellum) that beats. Although cells in our tissues do not have a flagellum in general, they are still able to move, as we will discover in this chapter. In fact, in both cases of movement, with or without a flagellum, cell motility is due to a dynamic re-arrangement of polymers inside the cell. Let us first have a closer look at the propulsion mechanism in the case of a flagellum or a cilium, which is the best known, but also the simplest, and which will help us to define the hydrodynamic general conditions of cell movement. A flagellum is sustained by cellular polymers arranged in semi-flexible bundles and flagellar beating generates cell displacement. These polymers or filaments are part of the cellular skeleton, or "cytoskeleton", which is, in this case, external to the cellular main body of the organism. In fact, bacteria move in a hydrodynamic regime in which viscosity dominates over inertia. The system is thus in a hydrodynamic regime of low Reynolds number (Box 5.1), which is nearly exclusively the case in all cell movements. Bacteria and their propulsion mode by flagella beating are our unicellular ancestors 3.5 billion years ago. Since then, we have evolved to form pluricellular organisms. However, to keep the ability of displacement, to heal our wounds for example, our cells lost their flagellum, since it was not optimal in a dense cell environment: cells are too close to each other to leave enough space for the flagella to accomplish propulsion. The cytoskeleton thus developed inside the cell body to ensure cell shape changes and movement, and also mechanical strength within a tissue. The cytoskeleton of our cells, like the polymers or filaments that sustain the flagellum, is also composed of semi-flexible filaments arranged in bundles, and also in

  8. Spirochete motility and morpholgy

    NASA Astrophysics Data System (ADS)

    Charon, Nyles

    2004-03-01

    Spirochetes have a unique structure, and as a result their motility is different from that of other bacteria. These organisms can swim in a highly viscous, gel-like medium, such as that found in connective tissue, that inhibits the motility of most other bacteria. In spirochetes, the organelles for motility, the periplasmic flagella, reside inside the cell within the periplasmic space. A given periplasmic flagellum is attached only at one end of the cell, and depending on the species, may or may not overlap in the center of the cell. The number of periplasmic flagella varies from species to species. These structures have been shown to be directly involved in motility and function by rotating within the periplasmic space (1). The present talk focuses on the spirochete that causes Lyme disease, Borrelia burgdorferi. In many bacterial species, cell shape is usually dictated by the peptidoyglycan layer of the cell wall. In the first part of the talk, results will be presented that the morphology of B. burgdorferi is the result of a complex interaction between the cell cylinder and the internal periplasmic flagella resulting in a cell with a flat-wave morphology. Backward moving, propagating waves enable these bacteria to swim and translate in a given direction. Using targeted mutagenesis, we inactivated the gene encoding the major periplasmic flagellar filament protein FlaB. The resulting flaB mutants not only were non-motile, but were rod-shaped (2). Western blot analysis indicated that flaB was no longer synthesized, and electron microscopy revealed that the mutants were completely deficient in periplasmic flagella. Our results indicate that the periplasmic flagella of B. burgdorferi have a skeletal function. These organelles dynamically interact with the rod-shaped cell cylinder to enable the cell to swim, and to confer in part its flat-wave morphology The latter part of the talk concerns the basis for asymmetrical rotation of the periplasmic flagella of B

  9. GLUTs and mammalian sperm metabolism.

    PubMed

    Bucci, Diego; Rodriguez-Gil, Juan Enrique; Vallorani, Claudia; Spinaci, Marcella; Galeati, Giovanna; Tamanini, Carlo

    2011-01-01

    Mammalian cells use glucides as a substrate that can be catabolized through glycolitic pathways or oxidative phosphorylation, used as a source of reducing potential, or used for anabolic aims. An important role in supplying cells with energy is played by different membrane proteins that can actively (sodium-dependent glucose transporters) or passively (glucose transporters; GLUT) transport hexoses through the lipidic bilayer. In particular, GLUTs are a family of 13 proteins that facilitate the transport of sugars and have a peculiar distribution in different tissues as well as a particular affinity for substrates. These proteins are also present in mature sperm cells, which, in fact, need carriers for uptake energetic sources that are important for maintaining cell basic activity as well as specific functions, such as motility and fertilization ability. Likewise, several GLUTs have been studied in various mammalian species (man, bull, rat, mouse, boar, dog, stallion, and donkey) to point out both their actual presence or absence and their localization on plasma membrane. The aim of this work is to give an overall picture of the studies available on GLUTs in mammalian spermatozoa at this moment, pointing out the species peculiarity, the possible role of these proteins, and the potential future research on this item.

  10. Mammalian sleep

    NASA Astrophysics Data System (ADS)

    Staunton, Hugh

    2005-05-01

    This review examines the biological background to the development of ideas on rapid eye movement sleep (REM sleep), so-called paradoxical sleep (PS), and its relation to dreaming. Aspects of the phenomenon which are discussed include physiological changes and their anatomical location, the effects of total and selective sleep deprivation in the human and animal, and REM sleep behavior disorder, the latter with its clinical manifestations in the human. Although dreaming also occurs in other sleep phases (non-REM or NREM sleep), in the human, there is a contingent relation between REM sleep and dreaming. Thus, REM is taken as a marker for dreaming and as REM is distributed ubiquitously throughout the mammalian class, it is suggested that other mammals also dream. It is suggested that the overall function of REM sleep/dreaming is more important than the content of the individual dream; its function is to place the dreamer protagonist/observer on the topographical world. This has importance for the developing infant who needs to develop a sense of self and separateness from the world which it requires to navigate and from which it is separated for long periods in sleep. Dreaming may also serve to maintain a sense of ‘I’ness or “self” in the adult, in whom a fragility of this faculty is revealed in neurological disorders.

  11. Hearing in Drosophila Requires TilB, a Conserved Protein Associated With Ciliary Motility

    PubMed Central

    Kavlie, Ryan G.; Kernan, Maurice J.; Eberl, Daniel F.

    2010-01-01

    Cilia were present in the earliest eukaryotic ancestor and underlie many biological processes ranging from cell motility and propulsion of extracellular fluids to sensory physiology. We investigated the contribution of the touch insensitive larva B (tilB) gene to cilia function in Drosophila melanogaster. Mutants of tilB exhibit dysfunction in sperm flagella and ciliated dendrites of chordotonal organs that mediate hearing and larval touch sensitivity. Mutant sperm axonemes as well as sensory neuron dendrites of Johnston's organ, the fly's auditory organ, lack dynein arms. Through deficiency mapping and sequencing candidate genes, we identified tilB mutations in the annotated gene CG14620. A genomic CG14620 transgene rescued deafness and male sterility of tilB mutants. TilB is a 395-amino-acid protein with a conserved N-terminal leucine-rich repeat region at residues 16–164 and a coiled-coil domain at residues 171–191. A tilB-Gal4 transgene driving fluorescently tagged TilB proteins elicits cytoplasmic expression in embryonic chordotonal organs, in Johnston's organ, and in sperm flagella. TilB does not appear to affect tubulin polyglutamylation or polyglycylation. The phenotypes and expression of tilB indicate function in cilia construction or maintenance, but not in intraflagellar transport. This is also consistent with phylogenetic association of tilB homologs with presence of genes encoding axonemal dynein arm components. Further elucidation of tilB functional mechanisms will provide greater understanding of cilia function and will facilitate understanding ciliary diseases. PMID:20215474

  12. Mechanics of motility initiation and motility arrest in crawling cells

    NASA Astrophysics Data System (ADS)

    Recho, Pierre; Putelat, Thibaut; Truskinovsky, Lev

    2015-11-01

    Motility initiation in crawling cells requires transformation of a symmetric state into a polarized state. In contrast, motility arrest is associated with re-symmetrization of the internal configuration of a cell. Experiments on keratocytes suggest that polarization is triggered by the increased contractility of motor proteins but the conditions of re-symmetrization remain unknown. In this paper we show that if adhesion with the extra-cellular substrate is sufficiently low, the progressive intensification of motor-induced contraction may be responsible for both transitions: from static (symmetric) to motile (polarized) at a lower contractility threshold and from motile (polarized) back to static (symmetric) at a higher contractility threshold. Our model of lamellipodial cell motility is based on a 1D projection of the complex intra-cellular dynamics on the direction of locomotion. In the interest of analytical transparency we also neglect active protrusion and view adhesion as passive. Despite the unavoidable oversimplifications associated with these assumptions, the model reproduces quantitatively the motility initiation pattern in fish keratocytes and reveals a crucial role played in cell motility by the nonlocal feedback between the mechanics and the transport of active agents. A prediction of the model that a crawling cell can stop and re-symmetrize when contractility increases sufficiently far beyond the motility initiation threshold still awaits experimental verification.

  13. Spirochete periplasmic flagella and motility.

    PubMed

    Li, C; Motaleb, A; Sal, M; Goldstein, S F; Charon, N W

    2000-10-01

    Spirochetes have a unique structure, and as a result their motility is different from that of other bacteria. They also have a special attribute: spirochetes can swim in a highly viscous, gel-like medium, such as that found in connective tissue, that inhibits the motility of most other bacteria. In spirochetes, the organelles for motility, the periplasmic flagella, reside inside the cell within the periplasmic space. A given periplasmic flagellum is attached only at one end of the cell, and depending on the species, may or may not overlap in the center of the cell with those attached at the other end. The number of periplasmic flagella varies from species to species. These structures have been shown to be directly involved in spirochete motility, and they function by rotating within the periplasmic space. The mechanics of motility also vary among the spirochetes. In Leptospira, a motility model developed several years ago has been extensively tested, and the evidence supporting this model is convincing. Borrelia burgdorferi swims differently, and a model of its motility has been recently put forward. This model is based on analyzing the motion of swimming cells, high voltage electron microscopy of fixed cells, and mutant analysis. To better understand spirochete motility on a more molecular level, the proteins and genes involved in motility are being analyzed. Spirochete periplasmic flagellar filaments are among the most complex of bacterial flagella. They are composed of the FlaA sheath proteins, and in many species, multiple FlaB core proteins. Allelic exchange mutagenesis of the genes which encode these proteins is beginning to yield important information with respect to periplasmic flagellar structure and function. Although we are at an early stage with respect to analyzing the function, organization, and regulation of many of the genes involved in spirochete motility, unique aspects have already become evident. Future studies on spirochete motility should be

  14. Effect of Cilia Beat Frequency on Muco-ciliary Clearance

    PubMed Central

    Sedaghat, M.H.; Shahmardan, M.M.; Norouzi, M.; Heydari, M.

    2016-01-01

    Background: The airway surface liquid (ASL), which is a fluid layer coating the interior epithelial surface of the bronchi and bronchiolesis, plays an important defensive role against foreign particles and chemicals entering lungs. Objective: Numerical investigation has been employed to solve two-layer model consisting of mucus layer as a viscoelastic fluid and periciliary liquid layer as a Newtonian fluid to study the effects of cilia beat frequency (CBF) at various amounts of mucus properties on muco-ciliary transport problem. Methods: Hybrid finite difference-lattice Boltzmann-method (FB-LBM) has been used to solve the momentum equations and to simulate cilia forces, and also the PCL-mucus interface more accurately, immersed boundary method (IBM) has been employed. The main contribution of the current study is to use an Oldroyd-B model as the constitutive equation of mucus. Results: Our results show that increasing CBF and decreasing mucus viscosity ratio have great effects on mucus flow, but the effect of viscosity ratio is more significant. The results also illustrate that the relation between cilia beat frequency and mean mucus velocity is almost linear and it has similar behavior at different values of viscosity ratio. Conclusion: Numerical investigation based on hybrid IB-FD-LBM has been used to study the effect of CBF at various mounts of mucus viscosity ratio on the muco-ciliary clearance. The results showed that the effect of viscosity ratio on the muco-ciliary transport process is more significant compared with CBF. PMID:28144596

  15. Motility and microtubule depolymerization mechanisms of the Kinesin-8 motor, KIF19A

    PubMed Central

    Wang, Doudou; Nitta, Ryo; Morikawa, Manatsu; Yajima, Hiroaki; Inoue, Shigeyuki; Shigematsu, Hideki; Kikkawa, Masahide; Hirokawa, Nobutaka

    2016-01-01

    The kinesin-8 motor, KIF19A, accumulates at cilia tips and controls cilium length. Defective KIF19A leads to hydrocephalus and female infertility because of abnormally elongated cilia. Uniquely among kinesins, KIF19A possesses the dual functions of motility along ciliary microtubules and depolymerization of microtubules. To elucidate the molecular mechanisms of these functions we solved the crystal structure of its motor domain and determined its cryo-electron microscopy structure complexed with a microtubule. The features of KIF19A that enable its dual function are clustered on its microtubule-binding side. Unexpectedly, a destabilized switch II coordinates with a destabilized L8 to enable KIF19A to adjust to both straight and curved microtubule protofilaments. The basic clusters of L2 and L12 tether the microtubule. The long L2 with a characteristic acidic-hydrophobic-basic sequence effectively stabilizes the curved conformation of microtubule ends. Hence, KIF19A utilizes multiple strategies to accomplish the dual functions of motility and microtubule depolymerization by ATP hydrolysis. DOI: http://dx.doi.org/10.7554/eLife.18101.001 PMID:27690357

  16. Mechanism of olfactory masking in the sensory cilia

    PubMed Central

    Ishida, Hirohiko; Hikichi, Satoshi; Kurahashi, Takashi

    2009-01-01

    Olfactory masking has been used to erase the unpleasant sensation in human cultures for a long period of history. Here, we show a positive correlation between the human masking and the odorant suppression of the transduction current through the cyclic nucleotide–gated (CNG) and Ca2+-activated Cl− (Cl(Ca)) channels. Channels in the olfactory cilia were activated with the cytoplasmic photolysis of caged compounds, and their sensitiveness to odorant suppression was measured with the whole cell patch clamp. When 16 different types of chemicals were applied to cells, cyclic AMP (cAMP)-induced responses (a mixture of CNG and Cl(Ca) currents) were suppressed widely with these substances, but with different sensitivities. Using the same chemicals, in parallel, we measured human olfactory masking with 6-rate scoring tests and saw a correlation coefficient of 0.81 with the channel block. Ringer's solution that was just preexposed to the odorant-containing air affected the cAMP-induced current of the single cell, suggesting that odorant suppression occurs after the evaporation and air/water partition of the odorant chemicals at the olfactory mucus. To investigate the contribution of Cl(Ca), the current was exclusively activated by using the ultraviolet photolysis of caged Ca, DM-nitrophen. With chemical stimuli, it was confirmed that Cl(Ca) channels were less sensitive to the odorant suppression. It is interpreted, however, that in the natural odorant response the Cl(Ca) is affected by the reduction of Ca2+ influx through the CNG channels as a secondary effect. Because the signal transmission between CNG and Cl(Ca) channels includes nonlinear signal-boosting process, CNG channel blockage leads to an amplified reduction in the net current. In addition, we mapped the distribution of the Cl(Ca) channel in living olfactory single cilium using a submicron local [Ca2+]i elevation with the laser photolysis. Cl(Ca) channels are expressed broadly along the cilia. We conclude that

  17. Sychronization of flagella and cilia due to viscous interactions

    NASA Astrophysics Data System (ADS)

    Gagnon, David; Qian, Bian; Jiang, Hongyuan; Powers, Thomas; Breuer, Kenneth

    2009-11-01

    Motivated by the observed coordination of nearby beating cilia and rotating bacterial flagella, we use a scaled model experiment to show that hydrodynamic interactions can cause synchronization between rotating paddles driven at constant torque in a very viscous fluid. Systems with two and three paddles are explored, and interactions between symmetric and asymmetric paddles are tested. For two-paddle systems, synchronization is only observed when the shafts supporting the paddles have some flexibility, and the phase difference in the synchronized state depends on the symmetry of the paddles. Calculations using the method of regularized stokeslets and simple analytic theory match the experimental observations well.

  18. Relationship between Porcine Sperm Motility and Sperm Enzymatic Activity using Paper-based Devices

    NASA Astrophysics Data System (ADS)

    Matsuura, Koji; Huang, Han-Wei; Chen, Ming-Cheng; Chen, Yu; Cheng, Chao-Min

    2017-04-01

    Mammalian sperm motility has traditionally been analyzed to determine fertility using computer-assisted semen analysis (CASA) systems. To develop low-cost and robust male fertility diagnostics, we created a paper-based MTT assay and used it to estimate motile sperm concentration. When porcine sperm motility was inhibited using sperm enzyme inhibitors for sperm enzymes related to mitochondrial activity and glycolysis, we simultaneously recorded sperm motility and enzymatic reactivity using a portable motility analysis system (iSperm) and a paper-based MTT assay, respectively. When using our paper-based MTT-assay, we calculated the area mean value signal intensity (AMV) to evaluate enzymatic reactivity. Both sperm motility and AMV decreased following treatment with iodoacetamide (IODO) and 3-bromopyruvic acid (3BP), both of which are inhibitors of glycolytic enzymes including glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a correlation between recorded motility using iSperm and AMV from our paper-based assay (P < 0.05), suggesting that a sperm-related enzymatic reaction is involved in sperm motility. Under this protocol, MTT reduction was coupled with catalysis of GAPDH and was promoted by electron transfer from NADH. Based on this inhibitor study, sperm motility can be estimated using our paper-based MTT-assay.

  19. Relationship between Porcine Sperm Motility and Sperm Enzymatic Activity using Paper-based Devices

    PubMed Central

    Matsuura, Koji; Huang, Han-Wei; Chen, Ming-Cheng; Chen, Yu; Cheng, Chao-Min

    2017-01-01

    Mammalian sperm motility has traditionally been analyzed to determine fertility using computer-assisted semen analysis (CASA) systems. To develop low-cost and robust male fertility diagnostics, we created a paper-based MTT assay and used it to estimate motile sperm concentration. When porcine sperm motility was inhibited using sperm enzyme inhibitors for sperm enzymes related to mitochondrial activity and glycolysis, we simultaneously recorded sperm motility and enzymatic reactivity using a portable motility analysis system (iSperm) and a paper-based MTT assay, respectively. When using our paper-based MTT-assay, we calculated the area mean value signal intensity (AMV) to evaluate enzymatic reactivity. Both sperm motility and AMV decreased following treatment with iodoacetamide (IODO) and 3-bromopyruvic acid (3BP), both of which are inhibitors of glycolytic enzymes including glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a correlation between recorded motility using iSperm and AMV from our paper-based assay (P < 0.05), suggesting that a sperm-related enzymatic reaction is involved in sperm motility. Under this protocol, MTT reduction was coupled with catalysis of GAPDH and was promoted by electron transfer from NADH. Based on this inhibitor study, sperm motility can be estimated using our paper-based MTT-assay. PMID:28387379

  20. Relationship between Porcine Sperm Motility and Sperm Enzymatic Activity using Paper-based Devices.

    PubMed

    Matsuura, Koji; Huang, Han-Wei; Chen, Ming-Cheng; Chen, Yu; Cheng, Chao-Min

    2017-04-07

    Mammalian sperm motility has traditionally been analyzed to determine fertility using computer-assisted semen analysis (CASA) systems. To develop low-cost and robust male fertility diagnostics, we created a paper-based MTT assay and used it to estimate motile sperm concentration. When porcine sperm motility was inhibited using sperm enzyme inhibitors for sperm enzymes related to mitochondrial activity and glycolysis, we simultaneously recorded sperm motility and enzymatic reactivity using a portable motility analysis system (iSperm) and a paper-based MTT assay, respectively. When using our paper-based MTT-assay, we calculated the area mean value signal intensity (AMV) to evaluate enzymatic reactivity. Both sperm motility and AMV decreased following treatment with iodoacetamide (IODO) and 3-bromopyruvic acid (3BP), both of which are inhibitors of glycolytic enzymes including glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a correlation between recorded motility using iSperm and AMV from our paper-based assay (P < 0.05), suggesting that a sperm-related enzymatic reaction is involved in sperm motility. Under this protocol, MTT reduction was coupled with catalysis of GAPDH and was promoted by electron transfer from NADH. Based on this inhibitor study, sperm motility can be estimated using our paper-based MTT-assay.

  1. Functional characterization of putative cilia genes by high-content analysis

    PubMed Central

    Lai, Cary K.; Gupta, Nidhi; Wen, Xiaohui; Rangell, Linda; Chih, Ben; Peterson, Andrew S.; Bazan, J. Fernando; Li, Li; Scales, Suzie J.

    2011-01-01

    Cilia are microtubule-based protrusions from the cell surface that are involved in a number of essential signaling pathways, yet little is known about many of the proteins that regulate their structure and function. A number of putative cilia genes have been identified by proteomics and comparative sequence analyses, but functional data are lacking for the vast majority. We therefore monitored the effects in three cell lines of small interfering RNA (siRNA) knockdown of 40 of these genes by high-content analysis. We assayed cilia number, length, and transport of two different cargoes (membranous serotonin receptor 6-green fluorescent protein [HTR6-GFP] and the endogenous Hedgehog [Hh] pathway transcription factor Gli3) by immunofluorescence microscopy; and cilia function using a Gli-luciferase Hh signaling assay. Hh signaling was most sensitive to perturbations, with or without visible structural cilia defects. Validated hits include Ssa2 and mC21orf2 with ciliation defects; Ift46 with short cilia; Ptpdc1 and Iqub with elongated cilia; and Arl3, Nme7, and Ssna1 with distinct ciliary transport but not length defects. Our data confirm various ciliary roles for several ciliome proteins and show it is possible to uncouple ciliary cargo transport from cilia formation in vertebrates. PMID:21289087

  2. Primary cilia are present on human blood and bone marrow cells and mediate Hedgehog signaling.

    PubMed

    Singh, Mohan; Chaudhry, Parvesh; Merchant, Akil A

    2016-12-01

    Primary cilia are nonmotile, microtubule-based organelles that are present on the cellular membrane of all eukaryotic cells. Functional cilia are required for the response to developmental signaling pathways such as Hedgehog (Hh) and Wnt/β-catenin. Although the Hh pathway has been shown to be active in leukemia and other blood cancers, there have been no reports describing the presence of primary cilia in human blood or leukemia cells. In the present study, we show that nearly all human blood and bone marrow cells have primary cilia (97-99%). In contrast, primary cilia on AML cell lines (KG1, KG1a, and K562) were less frequent (10-36% of cells) and were often shorter and dysmorphic, with less well-defined basal bodies. Finally, we show that treatment of blood cells with the Hh pathway ligand Sonic Hedgehog (SHh) causes translocation of Smoothened (SMO) to the primary cilia and activation of Hh target genes, demonstrating that primary cilia in blood cells are functional and participate in Hh signaling. Loss of primary cilia on leukemia cells may have important implications for aberrant pathway activation and response to SMO inhibitors currently in clinical development. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

  3. Type 3 adenylyl cyclase: a key enzyme mediating the cAMP signaling in neuronal cilia

    PubMed Central

    Qiu, Liyan; LeBel, Robert P; Storm, Daniel R; Chen, Xuanmao

    2016-01-01

    Cilia are rigid, centriole-derived, microtubule-based organelles present in a majority of vertebrate cells including neurons. They are considered the cellular “antennae” attuned for detecting a range of extracellular signals including photons, odorants, morphogens, hormones and mechanical forces. The ciliary microenvironment is distinct from most actin-based subcellular structures such as microvilli or synapses. In the nervous system, there is no evidence that neuronal cilia process any synaptic structure. Apparently, the structural features of neuronal cilia do not allow them to harbor any synaptic connections. Nevertheless, a large number of G protein-coupled receptors (GPCRs) including odorant receptors, rhodopsin, Smoothened, and type 6 serotonin receptor are found in cilia, suggesting that these tiny processes largely depend on metabotropic receptors and their tuned signals to impact neuronal functions. The type 3 adenylyl cyclase (AC3), widely known as a cilia marker, is highly and predominantly expressed in olfactory sensory cilia and primary cilia throughout the brain. We discovered that ablation of AC3 in mice leads to pleiotropic phenotypes including anosmia, failure to detect mechanical stimulation of airflow, cognitive deficit, obesity, and depression-like behaviors. Multiple lines of human genetic evidence also demonstrate that AC3 is associated with obesity, major depressive disorder (MDD), sarcoidosis, and infertility, underscoring its functional importance. Here we review recent progress on AC3, a key enzyme mediating the cAMP signaling in neuronal cilia. PMID:27785336

  4. Loss of cilia suppresses cyst growth in genetic models of autosomal dominant polycystic kidney disease

    PubMed Central

    Ma, Ming; Tian, Xin; Igarashi, Peter; Pazour, Gregory J.; Somlo, Stefan

    2013-01-01

    Kidney cysts occur following inactivation of polycystins in otherwise intact cilia or following complete removal of cilia by inactivation of intraflagellar transport-related proteins. We investigated the mechanisms of cyst formation in these two distinct processes by combining conditional inactivation of polycystins with concomitant ablation of cilia in developing and adult kidney and liver. We found that loss of intact cilia suppresses cyst growth following inactivation of polycystins and that the severity of cystic disease was directly related to the length of time between the initial loss of the polycystin proteins and the subsequent involution of cilia. This cilia-dependent cyst growth was not explained by activation of the MAPK/ERK, mTOR or cAMP pathways and is likely to be distinct from the mechanism of cyst growth following complete loss of cilia. The data establish the existence of a novel pathway defined by polycystin-dependent inhibition and cilia-dependent activation that promotes rapid cyst growth. PMID:23892607

  5. Unilateral nephrectomy elongates primary cilia in the remaining kidney via reactive oxygen species

    PubMed Central

    Han, Sang Jun; Jang, Hee-Seong; Kim, Jee In; Lipschutz, Joshua H.; Park, Kwon Moo

    2016-01-01

    The length of primary cilia is associated with normal cell and organ function. In the kidney, the change of functional cilia length/mass is associated with various diseases such as ischemia/reperfusion injury, polycystic kidney disease, and congenital solitary kidney. Here, we investigate whether renal mass reduction affects primary cilia length and function. To induce renal mass reduction, mice were subjected to unilateral nephrectomy (UNx). UNx increased kidney weight and superoxide formation in the remaining kidney. Primary cilia were elongated in proximal tubule cells, collecting duct cells and parietal cells of the remaining kidney. Mn(III) Tetrakis (1-methyl-4-pyridyl) porphyrin (MnTMPyP), an antioxidant, reduced superoxide formation in UNx-mice and prevented the elongation of primary cilia. UNx increased the expression of phosphorylated ERK, p21, and exocyst complex members Sec8 and Sec10, in the remaining kidney, and these increases were prevented by MnTMPyP. In MDCK, a kidney tubular epithelial cell line, cells, low concentrations of H2O2 treatment elongated primary cilia. This H2O2-induced elongation of primary cilia was also prevented by MnTMPyP treatment. Taken together, these data demonstrate that kidney compensation, induced by a reduction of renal mass, results in primary cilia elongation, and this elongation is associated with an increased production of reactive oxygen species (ROS). PMID:26923764

  6. Unilateral nephrectomy elongates primary cilia in the remaining kidney via reactive oxygen species.

    PubMed

    Han, Sang Jun; Jang, Hee-Seong; Kim, Jee In; Lipschutz, Joshua H; Park, Kwon Moo

    2016-02-29

    The length of primary cilia is associated with normal cell and organ function. In the kidney, the change of functional cilia length/mass is associated with various diseases such as ischemia/reperfusion injury, polycystic kidney disease, and congenital solitary kidney. Here, we investigate whether renal mass reduction affects primary cilia length and function. To induce renal mass reduction, mice were subjected to unilateral nephrectomy (UNx). UNx increased kidney weight and superoxide formation in the remaining kidney. Primary cilia were elongated in proximal tubule cells, collecting duct cells and parietal cells of the remaining kidney. Mn(III) Tetrakis (1-methyl-4-pyridyl) porphyrin (MnTMPyP), an antioxidant, reduced superoxide formation in UNx-mice and prevented the elongation of primary cilia. UNx increased the expression of phosphorylated ERK, p21, and exocyst complex members Sec8 and Sec10, in the remaining kidney, and these increases were prevented by MnTMPyP. In MDCK, a kidney tubular epithelial cell line, cells, low concentrations of H2O2 treatment elongated primary cilia. This H2O2-induced elongation of primary cilia was also prevented by MnTMPyP treatment. Taken together, these data demonstrate that kidney compensation, induced by a reduction of renal mass, results in primary cilia elongation, and this elongation is associated with an increased production of reactive oxygen species (ROS).

  7. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    McMurray, R. J.; Wann, A. K. T.; Thompson, C. L.; Connelly, J. T.; Knight, M. M.

    2013-12-01

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation.

  8. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells.

    PubMed

    McMurray, R J; Wann, A K T; Thompson, C L; Connelly, J T; Knight, M M

    2013-12-18

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation.

  9. Structural and Functional Recovery of Sensory Cilia in C. elegans IFT Mutants upon Aging

    PubMed Central

    Kennedy, Julie; Brear, Andrea G.; Prahlad, Veena; Blacque, Oliver E.; Sengupta, Piali

    2016-01-01

    The majority of cilia are formed and maintained by the highly conserved process of intraflagellar transport (IFT). Mutations in IFT genes lead to ciliary structural defects and systemic disorders termed ciliopathies. Here we show that the severely truncated sensory cilia of hypomorphic IFT mutants in C. elegans transiently elongate during a discrete period of adult aging leading to markedly improved sensory behaviors. Age-dependent restoration of cilia morphology occurs in structurally diverse cilia types and requires IFT. We demonstrate that while DAF-16/FOXO is dispensable, the age-dependent suppression of cilia phenotypes in IFT mutants requires cell-autonomous functions of the HSF1 heat shock factor and the Hsp90 chaperone. Our results describe an unexpected role of early aging and protein quality control mechanisms in suppressing ciliary phenotypes of IFT mutants, and suggest possible strategies for targeting subsets of ciliopathies. PMID:27906968

  10. Structural and Functional Recovery of Sensory Cilia in C. elegans IFT Mutants upon Aging.

    PubMed

    Cornils, Astrid; Maurya, Ashish K; Tereshko, Lauren; Kennedy, Julie; Brear, Andrea G; Prahlad, Veena; Blacque, Oliver E; Sengupta, Piali

    2016-12-01

    The majority of cilia are formed and maintained by the highly conserved process of intraflagellar transport (IFT). Mutations in IFT genes lead to ciliary structural defects and systemic disorders termed ciliopathies. Here we show that the severely truncated sensory cilia of hypomorphic IFT mutants in C. elegans transiently elongate during a discrete period of adult aging leading to markedly improved sensory behaviors. Age-dependent restoration of cilia morphology occurs in structurally diverse cilia types and requires IFT. We demonstrate that while DAF-16/FOXO is dispensable, the age-dependent suppression of cilia phenotypes in IFT mutants requires cell-autonomous functions of the HSF1 heat shock factor and the Hsp90 chaperone. Our results describe an unexpected role of early aging and protein quality control mechanisms in suppressing ciliary phenotypes of IFT mutants, and suggest possible strategies for targeting subsets of ciliopathies.

  11. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells

    PubMed Central

    McMurray, R. J.; Wann, A. K. T.; Thompson, C. L.; Connelly, J. T.; Knight, M. M.

    2013-01-01

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation. PMID:24346024

  12. Morphogenesis of respiratory syncytial virus in human primary nasal ciliated epithelial cells occurs at surface membrane microdomains that are distinct from cilia

    SciTech Connect

    Jumat, Muhammad Raihan; Yan, Yan; Ravi, Laxmi Iyer; Wong, Puisan; Huong, Tra Nguyen; Li, Chunwei; Tan, Boon Huan; Wang, De Yun; Sugrue, Richard J.

    2015-10-15

    The distribution of cilia and the respiratory syncytial virus (RSV) nucleocapsid (N) protein, fusion (F) protein, attachment (G) protein, and M2-1 protein in human ciliated nasal epithelial cells was examined at between 1 and 5 days post-infection (dpi). All virus structural proteins were localized at cell surface projections that were distinct from cilia. The F protein was also trafficked into the cilia, and while its presence increased as the infection proceeded, the N protein was not detected in the cilia at any time of infection. The presence of the F protein in the cilia correlated with cellular changes in the cilia and reduced cilia function. At 5 dpi extensive cilia loss and further reduced cilia function was noted. These data suggested that although RSV morphogenesis occurs at non-cilia locations on ciliated nasal epithelial cells, RSV infection induces changes in the cilia body that leads to extensive cilia loss. - Highlights: • Respiratory syncytial virus (RSV) infects nasal ciliated epithelial cells. • Virus morphogenesis occurs within filamentous projections distinct from cilia. • The RSV N protein was not detected in the cilia at any time during infection. • Trafficking of the F protein into the cilia occurred early in infection. • Presence of the F protein in cilia correlated with impaired cilia function.

  13. Why motor proteins team up - Intraflagellar transport in C. elegans cilia

    PubMed Central

    Mijalkovic, Jona; Prevo, Bram; Peterman, Erwin J. G.

    2016-01-01

    ABSTRACT Inside the cell, vital processes such as cell division and intracellular transport are driven by the concerted action of different molecular motor proteins. In C. elegans chemosensory cilia, 2 kinesin-2 family motor proteins, kinesin-II and OSM-3, team up to drive intraflagellar transport (IFT) in the anterograde direction, from base to tip, whereas IFT dynein hitchhikes toward the tip and subsequently drives IFT in the opposite, retrograde direction, thereby recycling both kinesins. While it is evident that at least a retrograde and an anterograde motor are necessary to drive IFT, it has remained puzzling why 2 same-polarity kinesins are employed. Recently, we addressed this question by combining advanced genome-engineering tools with ultrasensitive, quantitative fluorescence microscopy to study IFT with single-molecule sensitivity.1,2 Using this combination of approaches, we uncovered a differentiation in kinesin-2 function, in which the slower kinesin-II operates as an ‘importer’, loading IFT trains into the cilium before gradually handing them over to the faster OSM-3. OSM-3 subsequently acts as a long-range ‘transporter’, driving the IFT trains toward the tip. The two kinesin-2 motors combine their unique motility properties to achieve something neither motor can achieve on its own; that is to optimize the amount of cargo inside the cilium. In this commentary, we provide detailed insight into the rationale behind our research approach and comment on our recent findings. Moreover, we discuss the role of IFT dynein and provide an outlook on future studies. PMID:27384150

  14. 5-HT6 receptor blockade regulates primary cilia morphology in striatal neurons.

    PubMed

    Brodsky, Matthew; Lesiak, Adam J; Croicu, Alex; Cohenca, Nathalie; Sullivan, Jane M; Neumaier, John F

    2017-04-01

    The 5-HT6 receptor has been implicated in a variety of cognitive processes including habitual behaviors, learning, and memory. It is found almost exclusively in the brain, is expressed abundantly in striatum, and localizes to neuronal primary cilia. Primary cilia are antenna-like, sensory organelles found on most neurons that receive both chemical and mechanical signals from other cells and the surrounding environment; however, the effect of 5-HT6 receptor function on cellular morphology has not been examined. We confirmed that 5-HT6 receptors were localized to primary cilia in wild-type (WT) but not 5-HT6 knockout (5-HT6KO) in both native mouse brain tissue and primary cultured striatal neurons then used primary neurons cultured from WT or 5-HT6KO mice to study the function of these receptors. Selective 5-HT6 antagonists reduced cilia length in neurons cultured from wild-type mice in a concentration and time-dependent manner without altering dendrites, but had no effect on cilia length in 5-HT6KO cultured neurons. Varying the expression levels of heterologously expressed 5-HT6 receptors affected the fidelity of ciliary localization in both WT and 5-HT6KO neurons; overexpression lead to increasing amounts of 5-HT6 localization outside of the cilia but did not alter cilia morphology. Introducing discrete mutations into the third cytoplasmic loop of the 5-HT6 receptor greatly reduced, but did not entirely eliminate, trafficking of the 5-HT6 receptor to primary cilia. These data suggest that blocking 5-HT6 receptor activity reduces the length of primary cilia and that mechanisms that regulate trafficking of 5-HT6 receptors to cilia are more complex than previously thought. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Molecular complexes that direct rhodopsin transport to primary cilia

    PubMed Central

    Wang, Jing; Deretic, Dusanka

    2013-01-01

    Rhodopsin is a key molecular constituent of photoreceptor cells, yet understanding of how it regulates photoreceptor membrane trafficking and biogenesis of light-sensing organelles, the rod outer segments (ROS) is only beginning to emerge. Recently identified sequence of well-orchestrated molecular interactions of rhodopsin with the functional networks of Arf and Rab GTPases at multiple stages of intracellular targeting fits well into the complex framework of the biogenesis and maintenance of primary cilia, of which the ROS is one example. This review will discuss the latest progress in dissecting the molecular complexes that coordinate rhodopsin incorporation into ciliary-targeted carriers with the recruitment and activation of membrane tethering complexes and regulators of fusion with the periciliary plasma membrane. In addition to revealing the fundamental principals of ciliary membrane renewal, recent advances also provide molecular insight into the ways by which disruptions of the exquisitely orchestrated interactions lead to cilia dysfunction and result in human retinal dystrophies and syndromic diseases that affect multiple organs, including the eyes. PMID:24135424

  16. IFT46 plays an essential role in cilia development

    PubMed Central

    Lee, Mi-Sun; Hwang, Kyu-Seok; Oh, Hyun-Woo; Ji-Ae, Kim; Kim, Hyun-Taek; Cho, Hyun-Soo; Lee, Jeong-Ju; Ko, Je Yeong; Choi, Jung-Hwa; Jeong, Yun-Mi; You, Kwan-Hee; Kim, Joon; Park, Doo-Sang; Nam, Ki-Hoan; Aizawa, Shinichi; Kiyonari, Hiroshi; Shioi, Go; Park, Jong-Hoon; Zhou, Weibin; Kim, Nam-Soon; Kim, Cheol-Hee

    2015-01-01

    Cilia are microtubule-based structures that project into the extracellular space. Ciliary defects are associated with several human diseases, including polycystic kidney disease, primary ciliary dyskinesia, left-right axis patterning, hydrocephalus and retinal degeneration. However, the genetic and cellular biological control of ciliogenesis remains poorly understood. The IFT46 is one of the highly conserved intraflagellar transport complex B proteins. In zebrafish, ift46 is expressed in various ciliated tissues such as Kupffer’s vesicle, pronephric ducts, ears and spinal cord. We show that ift46 is localized to the basal body. Knockdown of ift46 gene results in multiple phenotypes associated with various ciliopathies including kidney cysts, pericardial edema and ventral axis curvature. In ift46 morphants, cilia in kidney and spinal canal are shortened and abnormal. Similar ciliary defects are observed in otic vesicles, lateral line hair cells, olfactory pits, but not in Kupffer’s vesicle. To explore the functions of Ift46 during mouse development, we have generated Ift46 knock-out mice. The Ift46 mutants have developmental defects in brain, neural tube and heart. In particular Ift46(−/−) homozygotes displays randomization of the embryo heart looping, which is a hallmark of defective left-right (L/R) axis patterning. Taken together, our results demonstrated that IFT46 has an essential role in vertebrate ciliary development. PMID:25722189

  17. The BBSome controls IFT assembly and turnaround in cilia.

    PubMed

    Wei, Qing; Zhang, Yuxia; Li, Yujie; Zhang, Qing; Ling, Kun; Hu, Jinghua

    2012-09-01

    The bidirectional movement of intraflagellar transport (IFT) particles, which are composed of motors, IFT-A and IFT-B subcomplexes, and cargoes, is required for the biogenesis and signalling of cilia(1,2). A successful IFT cycle depends on the proper assembly of the massive IFT particle at the ciliary base and its turnaround from anterograde to retrograde transport at the ciliary tip. However, how IFT assembly and turnaround are regulated in vivo remains elusive. From a whole-genome mutagenesis screen in Caenorhabditis elegans, we identified two hypomorphic mutations in dyf-2 and bbs-1 as the only mutants showing normal anterograde IFT transport but defective IFT turnaround at the ciliary tip. Further analyses revealed that the BBSome (refs 3, 4), a group of conserved proteins affected in human Bardet-Biedl syndrome(5) (BBS), assembles IFT complexes at the ciliary base, then binds to the anterograde IFT particle in a DYF-2- (an orthologue of human WDR19) and BBS-1-dependent manner, and lastly reaches the ciliary tip to regulate proper IFT recycling. Our results identify the BBSome as the key player regulating IFT assembly and turnaround in cilia.

  18. Alcohol stimulates ciliary motility of isolated airway axonemes through a nitric oxide, cyclase, and cyclic nucleotide-dependent kinase mechanism.

    PubMed

    Sisson, Joseph H; Pavlik, Jacqueline A; Wyatt, Todd A

    2009-04-01

    Lung mucociliary clearance provides the first line of defense from lung infections and is impaired in individuals who consume heavy amounts of alcohol. Previous studies have demonstrated that this alcohol-induced ciliary dysfunction occurs through impairment of nitric oxide (NO) and cyclic nucleotide-dependent kinase-signaling pathways in lung airway ciliated epithelial cells. Recent studies have established that all key elements of this alcohol-driven signaling pathway co-localize to the apical surface of the ciliated cells with the basal bodies. These findings led us to hypothesize that alcohol activates the cilia stimulation pathway at the organelle level. To test this hypothesis we performed experiments exposing isolated demembranated cilia (isolated axonemes) to alcohol and studied the effect of alcohol-stimulated ciliary motility on the pathways involved with isolated axoneme activation. Isolated demembranated cilia were prepared from bovine trachea and activated with adenosine triphosphate. Ciliary beat frequency, NO production, adenylyl and guanylyl cyclase activities, cAMP- and cGMP-dependent kinase activities were measured following exposure to biologically relevant concentrations of alcohol. Alcohol rapidly stimulated axoneme beating 40% above baseline at very low concentrations of alcohol (1 to 10 mM). This activation was specific to ethanol, required the synthesis of NO, the activation of soluble adenylyl cyclase (sAC), and the activation of both cAMP- and cGMP-dependent kinases (PKA and PKG), all of which were present in the isolated organelle preparation. Alcohol rapidly and sequentially activates the eNOS-->NO-->GC-->cGMP-->PKG and sAC-->cAMP--> PKA dual signaling pathways in isolated airway axonemes. These findings indicate a direct effect of alcohol on airway cilia organelle function and fully recapitulate the alcohol-driven activation of cilia known to exist in vivo and in intact lung ciliated cells in vitro following brief moderate alcohol

  19. Rimonabant, Gastrointestinal Motility and Obesity

    PubMed Central

    Sun, Yan; Chen, Jiande

    2012-01-01

    Background: Obesity and overweight affect more than half of the US population and are associated with a number of diseases. Rimonabant, a cannabinoid receptor 1 blocker in the endocannabinoid (EC) system, was indicated in Europe for the treatment of obesity and overweight patients with associated risk factors but withdrawn on Jan, 2009 because of side effects. Many studies have reported the effects of rimonabant on gastrointestinal (GI) motility and food intake. The aims of this review are: to review the relationship of EC system with GI motility and food intake;to review the studies of rimonabant on GI motility, food intake and obesity;and to report the tolerance and side effects of rimonabant. Methods: the literature (Pubmed database) was searched using keywords: rimonabant, obesity and GI motility. Results: GI motility is related with appetite, food intake and nutrients absorption. The EC system inhibits GI motility, reduces emesis and increases food intake; Rimonabant accelerates gastric emptying and intestinal transition but decreases energy metabolism and food intake. There is rapid onset of tolerance to the prokinetic effect of rimonabant. The main side effects of rimonabant are depression and GI symptoms. Conclusions: Rimonabant has significant effects on energy metabolism and food intake, probably mediated via its effects on GI motility. PMID:23449551

  20. Cholangiocyte cilia express TRPV4 and detect changes in luminal tonicity inducing bicarbonate secretion

    PubMed Central

    Gradilone, Sergio A.; Masyuk, Anatoliy I.; Splinter, Patrick L.; Banales, Jesus M.; Huang, Bing Q.; Tietz, Pamela S.; Masyuk, Tatyana V.; LaRusso, Nicholas F.

    2007-01-01

    Cholangiocytes, epithelial cells lining the biliary tree, have primary cilia extending from their apical membrane into the ductal lumen. Although important in disease, cilia also play a vital role in normal cellular functions. We reported that cholangiocyte cilia are sensory organelles responding to mechanical stimuli (i.e., luminal fluid flow) by alterations in intracellular Ca2+ and cAMP. Because cholangiocyte cilia are also ideally positioned to detect changes in composition and tonicity of bile, we hypothesized that cilia also function as osmosensors. TRPV4, a Ca2+-permeable ion channel, has been implicated in signal transduction of osmotic stimuli. Using purified rat cholangiocytes and perfused intrahepatic bile duct units (IBDUs), we found that TRPV4 is expressed on cholangiocyte cilia, and that hypotonicity induces an increase in intracellular Ca2+ in a TRPV4-, ciliary-, and extracellular calcium-dependent manner. The osmosensation of luminal tonicity by ciliary TRPV4 induces bicarbonate secretion, the main determinant of ductal bile formation, by a mechanism involving apical ATP release. Furthermore, the activation of TRPV4 in vivo, by its specific agonist, 4αPDD, induces an increase in bile flow as well as ATP release and bicarbonate secretion. Our results suggest that cholangiocyte primary cilia play an important role in ductal bile formation by acting as osmosensors. PMID:18024594

  1. Cholangiocyte cilia express TRPV4 and detect changes in luminal tonicity inducing bicarbonate secretion.

    PubMed

    Gradilone, Sergio A; Masyuk, Anatoliy I; Splinter, Patrick L; Banales, Jesus M; Huang, Bing Q; Tietz, Pamela S; Masyuk, Tatyana V; Larusso, Nicholas F

    2007-11-27

    Cholangiocytes, epithelial cells lining the biliary tree, have primary cilia extending from their apical membrane into the ductal lumen. Although important in disease, cilia also play a vital role in normal cellular functions. We reported that cholangiocyte cilia are sensory organelles responding to mechanical stimuli (i.e., luminal fluid flow) by alterations in intracellular Ca(2+) and cAMP. Because cholangiocyte cilia are also ideally positioned to detect changes in composition and tonicity of bile, we hypothesized that cilia also function as osmosensors. TRPV4, a Ca(2+)-permeable ion channel, has been implicated in signal transduction of osmotic stimuli. Using purified rat cholangiocytes and perfused intrahepatic bile duct units (IBDUs), we found that TRPV4 is expressed on cholangiocyte cilia, and that hypotonicity induces an increase in intracellular Ca(2+) in a TRPV4-, ciliary-, and extracellular calcium-dependent manner. The osmosensation of luminal tonicity by ciliary TRPV4 induces bicarbonate secretion, the main determinant of ductal bile formation, by a mechanism involving apical ATP release. Furthermore, the activation of TRPV4 in vivo, by its specific agonist, 4alphaPDD, induces an increase in bile flow as well as ATP release and bicarbonate secretion. Our results suggest that cholangiocyte primary cilia play an important role in ductal bile formation by acting as osmosensors.

  2. Role of cilia in normal pancreas function and in diseased states.

    PubMed

    diIorio, Philip; Rittenhouse, Ann R; Bortell, Rita; Jurczyk, Agata

    2014-06-01

    Primary cilia play an essential role in modulating signaling cascades that shape cellular responses to environmental cues to maintain proper tissue development. Mutations in primary cilium proteins have been linked to several rare developmental disorders, collectively known as ciliopathies. Together with other disorders associated with dysfunctional cilia/centrosomes, affected individuals have increased risk of developing metabolic syndrome, neurologic disorders, and diabetes. In pancreatic tissues, cilia are found exclusively in islet and ductal cells where they play an essential role in pancreatic tissue organization. Their absence or disorganization leads to pancreatic duct abnormalities, acinar cell loss, polarity defects, and dysregulated insulin secretion. Cilia in pancreatic tissues are hubs for cellular signaling. Many signaling components, such as Hh, Notch, and Wnt, localize to pancreatic primary cilia and are necessary for proper development of pancreatic epithelium and β-cell morphogenesis. Receptors for neuroendocrine hormones, such as Somatostatin Receptor 3, also localize to the cilium and may play a more direct role in controlling insulin secretion due to somatostatin's inhibitory function. Finally, unique calcium signaling, which is at the heart of β-cell function, also occurs in primary cilia. Whereas voltage-gated calcium channels trigger insulin secretion and serve a variety of homeostatic functions in β-cells, transient receptor potential channels regulate calcium levels within the cilium that may serve as a feedback mechanism, regulating insulin secretion. This review article summarizes our current understanding of the role of primary cilia in normal pancreas function and in the diseased state.

  3. Cilia Internal Mechanism and Metachronal Coordination as the Result of Hydrodynamical Coupling

    NASA Astrophysics Data System (ADS)

    Gueron, Shay; Levit-Gurevich, Konstantin; Liron, Nadav; Blum, Jacob J.

    1997-06-01

    We present a simple but realistic model for the internal bend-generating mechanism of cilia, using parameters obtained from the analysis of data of the beat of a single cilium, and incorporate it into a recently developed dynamical model. Comparing the results to experimental data for two-dimensional beats, we demonstrate that the model captures the essential features of the motion, including many properties that are not built in explicitly. The beat pattern and frequency change in response to increased viscosity and the presence of neighboring cilia in a realistic fashion. Using the model, we are able to investigate multicilia configurations such as rows of cilia and two-dimensional arrays of cilia. When two adjacent model cilia start beating at different phase, they synchronize within two cycles, as observed in experiments in which two flagella beating out of phase are brought close together. Examination of various multicilia configurations shows that metachronal patterns (i.e., beats with a constant phase difference between neighboring cilia) evolve autonomously. This provides modeling evidence in support of the conjecture that metachronism may occur as a self-organized phenomenon due to hydrodynamical interactions between the cilia.

  4. The Par-PrkC Polarity Complex Is Required for Cilia Growth in Zebrafish Photoreceptors

    PubMed Central

    Krock, Bryan L.; Perkins, Brian D.

    2014-01-01

    Specification and development of the apical membrane in epithelial cells requires the function of polarity proteins, including Pard3 and an atypical protein kinase C (PrkC). Many epithelial cells possess microtubule-based organelles, known as cilia, that project from their apical surface and the membrane surrounding the cilium is contiguous with the apical cell membrane. Although cilia formation in cultured cells required Pard3, the in vivo requirement for Pard3 in cilia development remains unknown. The vertebrate photoreceptor outer segment represents a highly specialized cilia structure in which to identify factors necessary for apical and ciliary membrane formation. Pard3 and PrkC localized to distinct domains within vertebrate photoreceptors. Using partial morpholino knockdown, photo-morpholinos, and pharmacological approaches, the function of Pard3 and PrkC were found to be required for the formation of both the apical and ciliary membrane of vertebrate photoreceptors. Inhibition of Pard3 or PrkC activity significantly reduced the size of photoreceptor outer segments and resulted in mislocalization of rhodopsin. Suppression of Pard3 or PrkC also led to a reduction in cilia size and cilia number in Kupffer’s Vesicle, which resulted in left-right asymmetry defects. Thus, the Par-PrkC complex functions in cilia formation in vivo and this likely reflects a general role in specifying non-ciliary and ciliary compartments of the apical domain. PMID:25144710

  5. INPP5E regulates phosphoinositide-dependent cilia transition zone function.

    PubMed

    Dyson, Jennifer M; Conduit, Sarah E; Feeney, Sandra J; Hakim, Sandra; DiTommaso, Tia; Fulcher, Alex J; Sriratana, Absorn; Ramm, Georg; Horan, Kristy A; Gurung, Rajendra; Wicking, Carol; Smyth, Ian; Mitchell, Christina A

    2017-01-02

    Human ciliopathies, including Joubert syndrome (JBTS), arise from cilia dysfunction. The inositol polyphosphate 5-phosphatase INPP5E localizes to cilia and is mutated in JBTS. Murine Inpp5e ablation is embryonically lethal and recapitulates JBTS, including neural tube defects and polydactyly; however, the underlying defects in cilia signaling and the function of INPP5E at cilia are still emerging. We report Inpp5e(-/-) embryos exhibit aberrant Hedgehog-dependent patterning with reduced Hedgehog signaling. Using mouse genetics, we show increasing Hedgehog signaling via Smoothened M2 expression rescues some Inpp5e(-/-) ciliopathy phenotypes and "normalizes" Hedgehog signaling. INPP5E's phosphoinositide substrates PI(4,5)P2 and PI(3,4,5)P3 accumulated at the transition zone (TZ) in Hedgehog-stimulated Inpp5e(-/-) cells, which was associated with reduced recruitment of TZ scaffolding proteins and reduced Smoothened levels at cilia. Expression of wild-type, but not 5-phosphatase-dead, INPP5E restored TZ molecular organization and Smoothened accumulation at cilia. Therefore, we identify INPP5E as an essential point of convergence between Hedgehog and phosphoinositide signaling at cilia that maintains TZ function and Hedgehog-dependent embryonic development.

  6. Histone deacetylase 6–mediated selective autophagy regulates COPD-associated cilia dysfunction

    PubMed Central

    Lam, Hilaire C.; Cloonan, Suzanne M.; Bhashyam, Abhiram R.; Haspel, Jeffery A.; Singh, Anju; Sathirapongsasuti, J. Fah; Cervo, Morgan; Yao, Hongwei; Chung, Anna L.; Mizumura, Kenji; An, Chang Hyeok; Shan, Bin; Franks, Jonathan M.; Haley, Kathleen J.; Owen, Caroline A.; Tesfaigzi, Yohannes; Washko, George R.; Quackenbush, John; Silverman, Edwin K.; Rahman, Irfan; Kim, Hong Pyo; Mahmood, Ashfaq; Biswal, Shyam S.; Ryter, Stefan W.; Choi, Augustine M.K.

    2013-01-01

    Chronic obstructive pulmonary disease (COPD) involves aberrant airway inflammatory responses to cigarette smoke (CS) that are associated with epithelial cell dysfunction, cilia shortening, and mucociliary clearance disruption. Exposure to CS reduced cilia length and induced autophagy in vivo and in differentiated mouse tracheal epithelial cells (MTECs). Autophagy-impaired (Becn1+/– or Map1lc3B–/–) mice and MTECs resisted CS-induced cilia shortening. Furthermore, CS increased the autophagic turnover of ciliary proteins, indicating that autophagy may regulate cilia homeostasis. We identified cytosolic deacetylase HDAC6 as a critical regulator of autophagy-mediated cilia shortening during CS exposure. Mice bearing an X chromosome deletion of Hdac6 (Hdac6–/Y) and MTECs from these mice had reduced autophagy and were protected from CS-induced cilia shortening. Autophagy-impaired Becn1–/–, Map1lc3B–/–, and Hdac6–/Y mice or mice injected with an HDAC6 inhibitor were protected from CS-induced mucociliary clearance (MCC) disruption. MCC was preserved in mice given the chemical chaperone 4-phenylbutyric acid, but was disrupted in mice lacking the transcription factor NRF2, suggesting that oxidative stress and altered proteostasis contribute to the disruption of MCC. Analysis of human COPD specimens revealed epigenetic deregulation of HDAC6 by hypomethylation and increased protein expression in the airways. We conclude that an autophagy-dependent pathway regulates cilia length during CS exposure and has potential as a therapeutic target for COPD. PMID:24200693

  7. Trichoplein and Aurora A block aberrant primary cilia assembly in proliferating cells.

    PubMed

    Inoko, Akihito; Matsuyama, Makoto; Goto, Hidemasa; Ohmuro-Matsuyama, Yuki; Hayashi, Yuko; Enomoto, Masato; Ibi, Miho; Urano, Takeshi; Yonemura, Shigenobu; Kiyono, Tohru; Izawa, Ichiro; Inagaki, Masaki

    2012-04-30

    The primary cilium is an antenna-like organelle that modulates differentiation, sensory functions, and signal transduction. After cilia are disassembled at the G0/G1 transition, formation of cilia is strictly inhibited in proliferating cells. However, the mechanisms of this inhibition are unknown. In this paper, we show that trichoplein disappeared from the basal body in quiescent cells, whereas it localized to mother and daughter centrioles in proliferating cells. Exogenous expression of trichoplein inhibited primary cilia assembly in serum-starved cells, whereas ribonucleic acid interference-mediated depletion induced primary cilia assembly upon cultivation with serum. Trichoplein controlled Aurora A (AurA) activation at the centrioles predominantly in G1 phase. In vitro analyses confirmed that trichoplein bound and activated AurA directly. Using trichoplein mutants, we demonstrate that the suppression of primary cilia assembly by trichoplein required its ability not only to localize to centrioles but also to bind and activate AurA. Trichoplein or AurA knockdown also induced G0/G1 arrest, but this phenotype was reversed when cilia formation was prevented by simultaneous knockdown of IFT-20. These data suggest that the trichoplein-AurA pathway is required for G1 progression through a key role in the continuous suppression of primary cilia assembly.

  8. Specific localization of scallop gill epithelial calmodulin in cilia.

    PubMed

    Stommel, E W; Stephens, R E; Masure, H R; Head, J F

    1982-03-01

    Calmodulin has been isolated and characterized from the gill of the bay scallop aequipecten irradians. Quantitative electrophoretic analysis of epithelial cell fractions show most of the calmodulin to be localized in the cilia, specifically in the detergent- solubilized membrane-matrix fraction. Calmodulin represents 2.2 +/- 0.3 percent of the membrane-matrix protein or 0.41 +/- 0.5 percent of the total ciliary protein. Its concentration is at least 10(-4) M if distributed uniformly within the matrix. Extraction in the presence of calcium suggests that the calmodulin is not bound to the axoneme proper. The ciliary protein is identified as a calmodulin on the basis of its calcium- dependent binding to a fluphenazine-sepharose affinity column and its comigration with bovine brain calmodulin on alkaline-urea and SDS polyacrylamide gels in both the presence and absence of calcium. Scallop ciliary calmodulin activates bovine brain phosphodiesterase to the same extent as bovine brain and chicken gizzard calmodulins. Containing trimethyllysine and lacking cysteine and tryptophan, the amino acid composition of gill calmodulin is typical of known calmodulins, except that it is relatively high in serine and low in methionine. Its composition is less acidic than other calmodulins, in agreement with an observed isoelectric point approximately 0.2 units higher than that of bovine brain. Comparative tryptic peptide mapping of scallop gill ciliary and bovine brain calmodulins indicates coincidence of over 75 percent of the major peptides, but at least two major peptides in each show no near-equivalency. Preliminary results using ATP-reactivated gill cell models show no effect of calcium at micromolar levels on ciliary beat or directionality of the lateral cilia, the cilia which constitute the vast majority of those isolated. However, ciliary arrest will occur at calcium levels more than 150 muM. Because calmodulin usually functions in the micromolar range, its role in this system

  9. Effects of cochlear loading on the motility of active outer hair cells

    PubMed Central

    Ó Maoiléidigh, Dáibhid; Hudspeth, A. J.

    2013-01-01

    Outer hair cells (OHCs) power the amplification of sound-induced vibrations in the mammalian inner ear through an active process that involves hair-bundle motility and somatic motility. It is unclear, though, how either mechanism can be effective at high frequencies, especially when OHCs are mechanically loaded by other structures in the cochlea. We address this issue by developing a model of an active OHC on the basis of observations from isolated cells, then we use the model to predict the response of an active OHC in the intact cochlea. We find that active hair-bundle motility amplifies the receptor potential that drives somatic motility. Inertial loading of a hair bundle by the tectorial membrane reduces the bundle’s reactive load, allowing the OHC’s active motility to influence the motion of the cochlear partition. The system exhibits enhanced sensitivity and tuning only when it operates near a dynamical instability, a Hopf bifurcation. This analysis clarifies the roles of cochlear structures and shows how the two mechanisms of motility function synergistically to create the cochlear amplifier. The results suggest that somatic motility evolved to enhance a preexisting amplifier based on active hair-bundle motility, thus allowing mammals to hear high-frequency sounds. PMID:23509256

  10. Gastrointestinal Motility Disorders in Children

    PubMed Central

    Ambartsumyan, Lusine

    2014-01-01

    The most common and challenging gastrointestinal motility disorders in children include gastroesophageal reflux disease (GERD), esophageal achalasia, gastroparesis, chronic intestinal pseudo-obstruction, and constipation. GERD is the most common gastrointestinal motility disorder affecting children and is diagnosed clinically and treated primarily with acid secretion blockade. Esophageal achalasia, a less common disorder in the pediatric patient population, is characterized by dysphagia and treated with pneumatic balloon dilation and/or esophagomyotomy. Gastroparesis and chronic intestinal pseudo-obstruction are poorly characterized in children and are associated with significant morbidity. Constipation is among the most common complaints in children and is associated with significant morbidity as well as poor quality of life. Data on epidemiology and outcomes, clinical trials, and evaluation of new diagnostic techniques are needed to better diagnose and treat gastrointestinal motility disorders in children. We present a review of the conditions and challenges related to these common gastrointestinal motility disorders in children. PMID:24799835

  11. Shape determination in motile cells

    NASA Astrophysics Data System (ADS)

    Mogilner, Alex

    2010-03-01

    Flat, simple shaped, rapidly gliding fish keratocyte cell is the model system of choice to study cell motility. The cell motile appendage, lamellipod, has a characteristic bent-rectangular shape. Recent experiments showed that the lamellipodial geometry is tightly correlated with cell speed and with actin dynamics. These quantitative data combined with computational modeling suggest that a model for robust actin treadmill inside the 'unstretchable membrane bag'. According to this model, a force balance between membrane tension and growing and pushing actin network distributed unevenly along the cell periphery can explain the cell shape and motility. However, when adhesion of the cell to the surface weakens, the actin dynamics become less regular, and myosin-powered contraction starts playing crucial role in stabilizing the cell shape. I will illustrate how the combination of theoretical and experimental approaches helped to unravel the keratocyte motile behavior.

  12. Surface motility of Myxococcus Xanthus

    NASA Astrophysics Data System (ADS)

    Gibiansky, Maxsim; Hu, William; Jin, Fan; Zhao, Kun; Shi, Wenyuan; Wong, Gerard

    2011-03-01

    We examine the surface motility of Myxococcus Xanthus, a bacterium species found in soil that exhibits a broad range of self-organizing behavior, including predatory ``swarms'' and survival-enhancing ``fruiting bodies.'' To quantify the effects of exopolysaccharides (EPS) on surface adhesion and motility, we use modified versions of particle tracking algorithms from colloid physics to analyze bacterial trajectories, and compare the wild type (WT) strain to EPS knockout and EPS overproducer strains. We find that EPS deficiency leads to an increase in the number of ``standing'' bacteria oriented normal to the surface, attached by one end with minimal motility. EPS overproduction, by contrast, suppresses this phenotype. A detailed investigation of the influence of EPS on Myxococcus social motility will be presented.

  13. The fine structure of the cilia from ctenophore swimming-plates.

    PubMed

    AFZELIUS, B A

    1961-02-01

    The ctenophore swimming-plate has been examined with the electron microscope. It has been recognized as an association of long cilia in tight hexagonal packing. One of the directions of the hexagonal packing is parallel to the long edge of the swimming-plate and is perpendicular to the direction of the ciliary beat. All the cilia in the swimming-plate are identically oriented. The effective beat in the movement of the swimming-plate is directed towards the aboral pole of the animal, and this is also the side of the unpaired peripheral filament in all the cilia. The direction of the ciliary beat is fixed in relation to the position of the filaments of the cilia. The swimming-plate cilium differs from other types of cilia and flagella in having a filament arrangement that can be described as 9 + 3 as opposed to the conventional 9 + 2 pattern. The central filaments appear in a group of two "tubular" filaments and an associated compact filament. The compact filament might have a supporting function. It has been called "midfilament." Two of the peripheral nine filaments (Fig. 1, Nos. 3 and 8) are joined to the ciliary membrane by means of slender lamellae, which divide the cilium into two unequal compartments. These lamellae have been called "compartmenting lamellae." Some observations of the arrangement of the compartmenting lamelae indicate that they function by cementing the cilia together in lateral rows. The cilia of the rows meet at a short distance from each other, leaving a gap of 30 A only. The meeting points are close to the termini of the compartmenting ridges. An electron-dense substance is sometimes seen bridging the gap. Some irregularities are noted with regard to the arrangement of the compartmenting lamellae particularly at the peripheral rows of cilia. In many cilia in these rows there are small vesicles beneath the ciliary membrane.

  14. Elenoside increases intestinal motility

    PubMed Central

    Navarro, E; Alonso, SJ; Navarro, R; Trujillo, J; Jorge, E

    2006-01-01

    AIM: To study the effects of elenoside, an arylnaph-thalene lignan from Justicia hyssopifolia, on gastro-intestinal motility in vivo and in vitro in rats. METHODS: Routine in vivo experimental assessments were catharsis index, water percentage of boluses, intestinal transit, and codeine antagonism. The groups included were vehicle control (propylene glycol-ethanol-plant oil-tween 80), elenoside (i.p. 25 and 50 mg/kg), cisapride (i.p. 10 mg/kg), and codeine phosphate (intragastric route, 50 mg/kg). In vitro approaches used isolated rat intestinal tissues (duodenum, jejunum, and ileum). The effects of elenoside at concentrations of 3.2 x 10-4, 6.4 x 10-4 and 1.2 x 10-3 mol/L, and cisapride at 10-6 mol/L were investigated. RESULTS: Elenoside in vivo produced an increase in the catharsis index and water percentage of boluses and in the percentage of distance traveled by a suspension of activated charcoal. Codeine phosphate antagonized the effect of 25 mg/kg of elenoside. In vitro, elenoside in duodenum, jejunum and ileum produced an initial decrease in the contraction force followed by an increase. Elenoside resulted in decreased intestinal frequency in duodenum, jejunum, and ileum. The in vitro and in vivo effects of elenoside were similar to those produced by cisapride. CONCLUSION: Elenoside is a lignan with an action similar to that of purgative and prokinetics drugs. Elenoside, could be an alternative to cisapride in treatment of gastrointestinal diseases as well as a preventive therapy for the undesirable gastrointestinal effects produced by opioids used for mild to moderate pain. PMID:17131476

  15. G-protein-coupled receptors, Hedgehog signaling and primary cilia.

    PubMed

    Mukhopadhyay, Saikat; Rohatgi, Rajat

    2014-09-01

    The Hedgehog (Hh) pathway has become an important model to study the cell biology of primary cilia, and reciprocally, the study of ciliary processes provides an opportunity to solve longstanding mysteries in the mechanism of vertebrate Hh signal transduction. The cilium is emerging as an unique compartment for G-protein-coupled receptor (GPCR) signaling in many systems. Two members of the GPCR family, Smoothened and Gpr161, play important roles in the Hh pathway. We review the current understanding of how these proteins may function to regulate Hh signaling and also highlight some of the critical unanswered questions being tackled by the field. Uncovering GPCR-regulated mechanisms important in Hh signaling may provide therapeutic strategies against the Hh pathway that plays important roles in development, regeneration and cancer.

  16. Bardet-Biedl syndrome: Is it only cilia dysfunction?

    PubMed

    Novas, Rossina; Cardenas-Rodriguez, Magdalena; Irigoín, Florencia; Badano, Jose L

    2015-11-14

    Bardet-Biedl syndrome (BBS) is a genetically heterogeneous, pleiotropic disorder, characterized by both congenital and late onset defects. From the analysis of the mutational burden in patients to the functional characterization of the BBS proteins, this syndrome has become a model for both understanding oligogenic patterns of inheritance and the biology of a particular cellular organelle: the primary cilium. Here we briefly review the genetics of BBS to then focus on the function of the BBS proteins, not only in the context of the cilium but also highlighting potential extra-ciliary roles that could be relevant to the etiology of the disorder. Finally, we provide an overview of how the study of this rare syndrome has contributed to the understanding of cilia biology and how this knowledge has informed on the cellular basis of different clinical manifestations that characterize BBS and the ciliopathies.

  17. Microfabrication of IPMC cilia for bio-inspired flow sensing

    NASA Astrophysics Data System (ADS)

    Lei, Hong; Li, Wen; Tan, Xiaobo

    2012-04-01

    As the primary flow sensing organ for fishes, the lateral line system plays a critical role in fish behavior. Analogous to its biological counterpart, an artificial lateral line system, consisting of arrays of micro flow sensors, is expected to be instrumental in the navigation and control of underwater robots. In this paper we investigate the microfabrication of ionic polymer-metal composite (IPMC) cilia for the purpose of flow sensing. While existing macro- and microfabrication methods for IPMCs have predominantly focused on planar structures, we propose a device where micro IPMC beams stand upright on a substrate to effectively interact with the flow. Challenges in the casting of 3D Nafion structure and selective formation of electrodes are discussed, and potential solutions for addressing these challenges are presented together with preliminary microfabrication results.

  18. [Establishment of osteoblast primary cilia model removed by chloral hyrate].

    PubMed

    Ma, Xiao-ni; Shi, Wen-gui; Xie, Yan-fang; Ma, Hui-ping; Ge, Bao-feng; Zhen, Ping; Chen, Ke-ming

    2015-06-01

    To establish osteoblast model, primary cilla model was removed by chloral hyrate, observe effects of osteoblast primary cilla moved on enhancing ALP staining and calcified nodules staining in electromagnetic field. Three 3-day-old male SD rats weighed between 6 and 9 g were killed, cranial osteoblast was drawed and adherencing cultured respectively. Cells were subcultured and randomly divided into 4 groups until reach to fusion states. The four groups included chloral hydrate non-involved group (control group), 2 mM, 4 mM and 8 mM chloral hydrate group, and cultured in 37 °C, 5% CO2 incubator for 72 h. Morphology of primary cilla was observed by laser confocal scanning microscope, and incidence of osteoblast primary cilia was analyzed by Image-Pro Plus 6.0 software. Cells in the correct concentration group which can removed cillia most effectively were selected and divided into 3 groups, including control group (C), Electromagnetic fields group (EMFs), and EMFs with 4 mM chloral hydrate group. DMEM nutrient solution contained 10%FBS were added into three groups and cultured for 9 days and formation of ALP were observed by histochemical staining of alkaline phosphatase. After 12 days' cultivation, formation of mineralization nodes was observed by alizarin red staining. Compared with control group and 2mM chloral hydrate group,4 mM chloral hydrate group could effectively remove osteoblast primary cilla (P<0.01). Removal of osteoblast primary cilla could weaken the formation of ALP and mineralization nodes in osteoblast in EMFS. Compared with EMFs group, the area of ALP and mineralization nodes in EMFs with 4 mM chloral hydrate group were decreased obviously (P<0.01). 4mM chloral hydrate could effectively remove osteoblast primary cilia. Primary cilla participate in EMFs promoting formation of ALP and mineralization nodes in osteoblast and provide new ideas for exploring mechanism of EMFs promoting osteoblast maturation and mineralization.

  19. Intestinal cell kinase, a protein associated with endocrine-cerebro-osteodysplasia syndrome, is a key regulator of cilia length and Hedgehog signaling.

    PubMed

    Moon, Heejung; Song, Jieun; Shin, Jeong-Oh; Lee, Hankyu; Kim, Hong-Kyung; Eggenschwiller, Jonathan T; Bok, Jinwoong; Ko, Hyuk Wan

    2014-06-10

    Endocrine-cerebro-osteodysplasia (ECO) syndrome is a recessive genetic disorder associated with multiple congenital defects in endocrine, cerebral, and skeletal systems that is caused by a missense mutation in the mitogen-activated protein kinase-like intestinal cell kinase (ICK) gene. In algae and invertebrates, ICK homologs are involved in flagellar formation and ciliogenesis, respectively. However, it is not clear whether this role of ICK is conserved in mammals and how a lack of functional ICK results in the characteristic phenotypes of human ECO syndrome. Here, we generated Ick knockout mice to elucidate the precise role of ICK in mammalian development and to examine the pathological mechanisms of ECO syndrome. Ick null mouse embryos displayed cleft palate, hydrocephalus, polydactyly, and delayed skeletal development, closely resembling ECO syndrome phenotypes. In cultured cells, down-regulation of Ick or overexpression of kinase-dead or ECO syndrome mutant ICK resulted in an elongation of primary cilia and abnormal Sonic hedgehog (Shh) signaling. Wild-type ICK proteins were generally localized in the proximal region of cilia near the basal bodies, whereas kinase-dead ICK mutant proteins accumulated in the distal part of bulged ciliary tips. Consistent with these observations in cultured cells, Ick knockout mouse embryos displayed elongated cilia and reduced Shh signaling during limb digit patterning. Taken together, these results indicate that ICK plays a crucial role in controlling ciliary length and that ciliary defects caused by a lack of functional ICK leads to abnormal Shh signaling, resulting in congenital disorders such as ECO syndrome.

  20. Construction of (001) facets exposed ZnO nanosheets on magnetically driven cilia film for highly active photocatalysis

    NASA Astrophysics Data System (ADS)

    Peng, Fengping; Zhou, Qiang; Lu, Chunhua; Ni, Yaru; Kou, Jiahui; Xu, Zhongzi

    2017-02-01

    ZnO nanosheet arrays with exposed (001) facets have been constructed onto a biomimetic inner-motile film, using a seed-mediated hydrothermal growth technology without adding capping agents. The growth of ZnO nanoparticles along the [001] direction is impeded because of a physical steric hindrance, and therefore (001) planes are left behind as the dominant crystal facets. In comparison to ZnO nanorod arrays film, the photocatalytic activity of the actuated (001) facets exposed ZnO nanosheet arrays film is dramatically improved to approximately 2.48 times. Moreover, when it is subjected to a rotational magnetic field, the ZnO nanosheet arrays film is driven to mimic ciliary motion like nature beating cilia, which can boost the interior mass transfer and help to promote release of active sites for improving the photocatalytic activity. As a consequence of the exposed (001) high active facets, the singular ability of microfluidic manipulation has greater effect on ZnO nanosheet arrays films. The enhancement of photocatalytic activity of the actuated ZnO nanosheet arrays film is much more than that of ZnO nanorod arrays film.

  1. Tubulin glycylases are required for primary cilia, control of cell proliferation and tumor development in colon

    PubMed Central

    Rocha, Cecilia; Papon, Laura; Cacheux, Wulfran; Marques Sousa, Patricia; Lascano, Valeria; Tort, Olivia; Giordano, Tiziana; Vacher, Sophie; Lemmers, Benedicte; Mariani, Pascale; Meseure, Didier; Medema, Jan Paul; Bièche, Ivan; Hahne, Michael; Janke, Carsten

    2014-01-01

    TTLL3 and TTLL8 are tubulin glycine ligases catalyzing posttranslational glycylation of microtubules. We show here for the first time that these enzymes are required for robust formation of primary cilia. We further discover the existence of primary cilia in colon and demonstrate that TTLL3 is the only glycylase in this organ. As a consequence, colon epithelium shows a reduced number of primary cilia accompanied by an increased rate of cell division in TTLL3-knockout mice. Strikingly, higher proliferation is compensated by faster tissue turnover in normal colon. In a mouse model for tumorigenesis, lack of TTLL3 strongly promotes tumor development. We further demonstrate that decreased levels of TTLL3 expression are linked to the development of human colorectal carcinomas. Thus, we have uncovered a novel role for tubulin glycylation in primary cilia maintenance, which controls cell proliferation of colon epithelial cells and plays an essential role in colon cancer development. PMID:25180231

  2. Sensory signaling-dependent remodeling of olfactory cilia architecture in C. elegans.

    PubMed

    Mukhopadhyay, Saikat; Lu, Yun; Shaham, Shai; Sengupta, Piali

    2008-05-01

    Nonmotile primary cilia are sensory organelles composed of a microtubular axoneme and a surrounding membrane sheath that houses signaling molecules. Optimal cellular function requires the precise regulation of axoneme assembly, membrane biogenesis, and signaling protein targeting and localization via as yet poorly understood mechanisms. Here, we show that sensory signaling is required to maintain the architecture of the specialized AWB olfactory neuron cilia in C. elegans. Decreased sensory signaling results in alteration of axoneme length and expansion of a membraneous structure, thereby altering the topological distribution of a subset of ciliary transmembrane signaling molecules. Signaling-regulated alteration of ciliary structures can be bypassed by modulation of intracellular cGMP or calcium levels and requires kinesin-II-driven intraflagellar transport (IFT), as well as BBS- and RAB8-related proteins. Our results suggest that compensatory mechanisms in response to altered levels of sensory activity modulate AWB cilia architecture, revealing remarkable plasticity in the regulation of cilia structure.

  3. A role for central spindle proteins in cilia structure and function

    PubMed Central

    Smith, Katherine R.; Kieserman, Esther K.; Wang, Peggy I.; Basten, Sander G.; Giles, Rachel H.; Marcotte, Edward M.; Wallingford, John B.

    2013-01-01

    Cytokinesis and ciliogenesis are fundamental cellular processes that require strict coordination of microtubule organization and directed membrane trafficking. These processes have been intensely studied, but there has been little indication that regulatory machinery might be extensively shared between them. Here, we show that several central spindle/midbody proteins (PRC1, MKLP-1, INCENP, centriolin) also localize in specific patterns at the basal body complex in vertebrate ciliated epithelial cells. Moreover, bioinformatic comparisons of midbody and cilia proteomes reveal a highly significant degree of overlap. Finally, we used temperature-sensitive alleles of PRC1/spd-1 and MKLP-1/zen-4 in C. elegans to assess ciliary functions while bypassing these proteins' early role in cell division. These mutants displayed defects in both cilia function and cilia morphology. Together, these data suggest the conserved re-use of a surprisingly large number of proteins in the cytokinetic apparatus and in cilia. PMID:21246755

  4. Synchronization and Collective Dynamics of Flagella and Cilia as Hydrodynamically Coupled Oscillators

    NASA Astrophysics Data System (ADS)

    Uchida, Nariya; Golestanian, Ramin; Bennett, Rachel R.

    2017-10-01

    Cooperative motion of flagella and cilia faciliates swimming of microorganisms and material transport in the body of multicellular organisms. Using minimal models, we address the roles of hydrodynamic interaction in synchronization and collective dynamics of flagella and cilia. Collective synchronization of bacterial flagella is studied with a model of bacterial carpets. Cilia and eukaryotic flagella are characterized by periodic modulation of their driving forces, which produces various patterns of two-body synchronization and metachronal waves. Long-range nature of the interaction introduces novel features in the dynamics of these model systems. The flagella of a swimmer synchronize also by a viscous drag force mediated through the swimmer's body. Recent advance in experimental studies of the collective dynamics of flagella, cilia and related artificial systems are summarized.

  5. An Experimental and Computational Analysis of Primary Cilia Deflection Under Fluid Flow

    PubMed Central

    Downs, Matthew E.; Nguyen, An M.; Herzog, Florian A.; Hoey, David A.; Jacobs, Christopher R.

    2013-01-01

    In this work we have developed a novel model of the deflection of primary cilia experiencing fluid flow accounting for phenomena not previously considered. Specifically, we developed a large rotation formulation that accounts for rotation at the base of the cilium, the initial shape of the cilium and fluid drag at high deflection angles. We utilized this model to analyze full three dimensional datasets of primary cilia deflecting under fluid flow acquired with high-speed confocal microscopy. We found a wide variety of previously unreported bending shapes and behaviors. We also analyzed post-flow relaxation patterns. Results from our combined experimental and theoretical approach suggest that the average flexural rigidity of primary cilia might be higher than previously reported (Schwartz et al. 1997). In addition our findings indicate the mechanics of primary cilia are richly varied and mechanisms may exist to alter their mechanical behavior. PMID:22452422

  6. Cilia assembly: a role for F-actin in IFT recruitment.

    PubMed

    Quarmby, Lynne

    2014-09-08

    Ciliary growth rates are limited by the availability of precursors at the growing tip. A new paper reveals that the early rapid growth of nascent cilia is supported by F-actin-facilitated delivery of IFT proteins to basal bodies.

  7. Roles for primary cilia in gonadotrophin-releasing hormone neurones in the mouse.

    PubMed

    Shin, J; Prescott, M; Mair, J; Campbell, R E

    2014-01-01

    During embryonic development, gonadotrophin-releasing hormone (GnRH) neurones make an extraordinary migration out of the nose and into the brain where, in adulthood, they drive the pituitary regulation of gonadal function and fertility. Primary cilia are antennae-like, immotile organelles that project from the surface of nearly all cells, including GnRH neurones. Links between defects in primary cilia and a variety of human pathologies have been discovered that suggest a role for primary cilia in embryogenesis and reproductive function. The present study aimed to investigate whether GnRH neurone primary cilia are critical for their embryonic migration and the adult control of fertility. To achieve this, we used a Cre-loxP strategy to selectively disrupt primary cilia by deleting Kif3a, an intraflagellar transport protein family member essential for primary cilia assembly and function, specifically in GnRH neurones. Confocal analysis revealed that, in Kif3a(fl/fl) (WT-Kif3a) controls, all GnRH neurones possessed primary cilia, whereas, in GnRH-Cre(+/-) ;Kif3a(fl/fl) (GnRH-Kif3aKO) mice, 60% of GnRH neurones lacked any evidence of primary cilia and the remaining 40% possessed only stunted primary cilia (< 2 μm). Despite abolishing normal primary cilia assembly in GnRH neurones from embryogenesis, adult GnRH neurone distribution and reproductive function was remarkably normal. The total number of GnRH neurones was the same in GnRH-Kif3aKO and WT-Kif3a controls; however, a significant increase (25%) was identified in the number of GnRH neurones sampled through the midpoint of the rostral pre-optic area in GnRH-Kif3aKO mice (P < 0.05). The time to vaginal opening was not different in GnRH-Kif3aKO mice, although they displayed significantly advanced first oestrus (P < 0.05), and oestrous cycle length was increased (P < 0.05). However, females displayed normal basal levels of luteinising hormone, responded normally to oestrogen-induced negative- and positive

  8. CB-08KIF3A IS ESSENTIAL FOR CILIOGENESIS, CILIA FUNCTION AND PROMOTES GLIOBLASTOMA PROGRESSION

    PubMed Central

    Hoang-Minh, Lan; Deleyrolle, Loic; Ugartemendia, George; Breunig, Joshua; Semple-Rowland, Susan; Reynolds, Brent; Sarkisian, Matthew

    2014-01-01

    Despite recent findings that cilia transduce diverse signaling pathways affecting cell proliferation, migration and survival, little is known about the influence of cilia or cilia-associated proteins in glioblastoma multiforme (GBM). We recently showed that primary cilia project from subsets of cells in GBM patient biopsies and derived cell lines. To determine if cilia contribute to GBM growth, we blocked ciliogenesis using a lentivirus expressing a dominant negative form of KIF3A, an essential ciliogenesis protein. We generated stable GBM cell lines (L0 and S3; representing different molecular subclasses) whereby dnKIF3A+ cells exhibited virtual complete loss of cilia compared to controls (confirmed by immunostaining and EM). Canonically, secreted Sonic hedgehog (SHH) ligand binds and activates receptor signaling cascades (e.g., smoothened (SMO)) within cilia to promote normal cell proliferation and tumor cell growth in specific developmental and pathological contexts, respectively. To examine the role of SHH in GBM proliferation, we exposed control and dnKIF3A+ L0 and S3 cells to saline or recombinant SHH. We found the number of L0 control cells significantly increased after SHH compared to saline, an effect blocked by pretreatment with cyclopamine (SMO inhibitor). However, SHH did not increase the number of L0 dnKIF3A+ cells. Interestingly, SHH exposure had no effect on S3 control cell numbers, despite observations that SHH signaling components (SMO and Gli3) were recruited to their cilia in response to SHH. This suggests GBM cilia are SHH-responsive but the downstream consequences of ciliary signaling may differ between cell lines. Notably, mice intracranially xenografted with L0 cells expressing dnKIF3A survived significantly longer than mice receiving control cells, and retained the loss of cilia phenotype in the tumors. Collectively, these data suggest KIF3A promotes GBM tumor progression, but the extent to which the effects are mediated by cilia and the

  9. Type 3 Adenylyl Cyclase and Somatostatin Receptor 3 Expression Persists in Aged Rat Neocortical and Hippocampal Neuronal Cilia

    PubMed Central

    Guadiana, Sarah M.; Parker, Alexander K.; Filho, Gileno F.; Sequeira, Ashton; Semple-Rowland, Susan; Shaw, Gerry; Mandel, Ronald J.; Foster, Thomas C.; Kumar, Ashok; Sarkisian, Matthew R.

    2016-01-01

    The primary cilia of forebrain neurons assemble around birth and become enriched with neuromodulatory receptors. Our understanding of the permanence of these structures and their associated signaling pathways in the aging brain is poor, but they are worthy of investigation because disruptions in neuronal cilia signaling have been implicated in changes in learning and memory, depression-like symptoms, and sleep anomalies. Here, we asked whether neurons in aged forebrain retain primary cilia and whether the staining characteristics of aged cilia for type 3 adenylyl cyclase (ACIII), somatostatin receptor 3 (SSTR3), and pericentrin resemble those of cilia in younger forebrain. To test this, we analyzed immunostained sections of forebrain tissues taken from young and aged male Fischer 344 (F344) and F344 × Brown Norway (F344 × BN) rats. Analyses of ACIII and SSTR3 in young and aged cortices of both strains of rats revealed that the staining patterns in the neocortex and hippocampus were comparable. Virtually every NeuN positive cell examined possessed an ACIII positive cilium. The lengths of ACIII positive cilia in neocortex were similar between young and aged for both strains, whereas in F344 × BN hippocampus, the cilia lengths increased with age in CA1 and CA3, but not in dentate gyrus (DG). Additionally, the percentages of ACIII positive cilia that were also SSTR3 positive did not differ between young and aged tissues in either strain. We also found that pericentrin, a protein that localizes to the basal bodies of neuronal cilia and functions in primary cilia assembly, persisted in aged cortical neurons of both rat strains. Collectively, our data show that neurons in aged rat forebrain possess primary cilia and that these cilia, like those present in younger brain, continue to localize ACIII, SSTR3, and pericentrin. Further studies will be required to determine if the function and signaling pathways regulated by cilia are similar in aged compared to young brain

  10. Biophysics and biofluid dynamics of primary cilia: evidence for and against the flow-sensing function.

    PubMed

    Nag, Subhra; Resnick, Andrew

    2017-09-01

    Primary cilia have been called "the forgotten organelle" for over 20 yr. As cilia now have their own journal and several books devoted to their study, perhaps it is time to reconsider the moniker "forgotten organelle." In fact, during the drafting of this review, 12 relevant publications have been issued; we therefore apologize in advance for any relevant work we inadvertently omitted. What purpose is yet another ciliary review? The primary goal of this review is to specifically examine the evidence for and against the hypothesized flow-sensing function of primary cilia expressed by differentiated epithelia within a kidney tubule, bringing together differing disciplines and their respective conceptual and experimental approaches. We will show that understanding the biophysics/biomechanics of primary cilia provides essential information for understanding any potential role of ciliary function in disease. We will summarize experimental and mathematical models used to characterize renal fluid flow and incident force on primary cilia and to characterize the mechanical response of cilia to an externally applied force and discuss possible ciliary-mediated cell signaling pathways triggered by flow. Throughout, we stress the importance of separating the effects of fluid shear and stretch from the action of hydrodynamic drag. Copyright © 2017 the American Physiological Society.

  11. Zonal variation in primary cilia elongation correlates with localized biomechanical degradation in stress deprived tendon

    PubMed Central

    Rowson, Daniel; Screen, Hazel R.C.

    2016-01-01

    ABSTRACT Tenocytes express primary cilia, which elongate when tendon is maintained in the absence of biomechanical load. Previous work indicates differences in the morphology and metabolism of the tenocytes in the tendon fascicular matrix (FM) and the inter‐fascicular matrix (IFM). This study tests the hypothesis that primary cilia in these two regions respond differently to stress deprivation and that this is associated with differences in the biomechanical degradation of the extracellular matrix. Rat tail tendon fascicles were examined over a 7‐day period of either stress deprivation or static load. Seven days of stress deprivation induced cilia elongation in both regions. However, elongation was greater in the IFM compared to the FM. Stress deprivation also induced a loss of biomechanical integrity, primarily in the IFM. Static loading reduced both the biomechanical degradation and cilia elongation. The different responses to stress deprivation in the two tendon regions are likely to be important for the aetiology of tendinopathy. Furthermore, these data suggest that primary cilia elongate in response to biomechanical degradation rather than simply the removal of load. This response to degradation is likely to have important consequences for cilia signalling in tendon and as well as in other connective tissues. © 2016 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 34:2146–2153, 2016. PMID:26969839

  12. Hippocampal and Cortical Primary Cilia Are Required for Aversive Memory in Mice

    PubMed Central

    Yazdi, S. M. Zaki R.; McNair, Andrew D.; Kippe, Jordyn M.; Croyle, Mandy J.; Kraft, Timothy W.; Yoder, Bradley K.

    2014-01-01

    It has been known for decades that neurons throughout the brain possess solitary, immotile, microtubule based appendages called primary cilia. Only recently have studies tried to address the functions of these cilia and our current understanding remains poor. To determine if neuronal cilia have a role in behavior we specifically disrupted ciliogenesis in the cortex and hippocampus of mice through conditional deletion of the Intraflagellar Transport 88 (Ift88) gene. The effects on learning and memory were analyzed using both Morris Water Maze and fear conditioning paradigms. In comparison to wild type controls, cilia mutants displayed deficits in aversive learning and memory and novel object recognition. Furthermore, hippocampal neurons from mutants displayed an altered paired-pulse response, suggesting that loss of IFT88 can alter synaptic properties. A variety of other behavioral tests showed no significant differences between conditional cilia mutants and controls. This type of conditional allele approach could be used to distinguish which behavioral features of ciliopathies arise due to defects in neural development and which result from altered cell physiology. Ultimately, this could lead to an improved understanding of the basis for the cognitive deficits associated with human cilia disorders such as Bardet-Biedl syndrome, and possibly more common ailments including depression and schizophrenia. PMID:25184295

  13. Zonal variation in primary cilia elongation correlates with localized biomechanical degradation in stress deprived tendon.

    PubMed

    Rowson, Daniel; Knight, Martin M; Screen, Hazel R C

    2016-12-01

    Tenocytes express primary cilia, which elongate when tendon is maintained in the absence of biomechanical load. Previous work indicates differences in the morphology and metabolism of the tenocytes in the tendon fascicular matrix (FM) and the inter-fascicular matrix (IFM). This study tests the hypothesis that primary cilia in these two regions respond differently to stress deprivation and that this is associated with differences in the biomechanical degradation of the extracellular matrix. Rat tail tendon fascicles were examined over a 7-day period of either stress deprivation or static load. Seven days of stress deprivation induced cilia elongation in both regions. However, elongation was greater in the IFM compared to the FM. Stress deprivation also induced a loss of biomechanical integrity, primarily in the IFM. Static loading reduced both the biomechanical degradation and cilia elongation. The different responses to stress deprivation in the two tendon regions are likely to be important for the aetiology of tendinopathy. Furthermore, these data suggest that primary cilia elongate in response to biomechanical degradation rather than simply the removal of load. This response to degradation is likely to have important consequences for cilia signalling in tendon and as well as in other connective tissues. © 2016 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 34:2146-2153, 2016.

  14. Directed Fluid Flow Produced by Arrays of Magnetically Actuated Core-Shell Biomimetic Cilia

    NASA Astrophysics Data System (ADS)

    Fiser, B. L.; Shields, A. R.; Evans, B. A.; Superfine, R.

    2010-03-01

    We have developed a novel core-shell microstructure that we use to fabricate arrays of flexible, magnetically actuated biomimetic cilia. Our biomimetic cilia mimic the size and beat shape of biological cilia in order to replicate the transport of fluid driven by cilia in many biological systems including the determination of left-right asymmetry in the vertebrate embryonic nodal plate and mucociliary clearance in the lung. Our core-shell structures consist of a flexible poly(dimethylsiloxane) (PDMS) core surrounded by a shell of nickel approximately forty nanometers thick; by using a core-shell structure, we can tune the mechanical and magnetic properties independently. We present the fabrication process and the long-range transport that occurs above the beating biomimetic cilia tips and will report on progress toward biomimetic cilia induced flow in viscoelastic fluids similar to mucus in the human airway. These flows may have applications in photonics and microfluidics, and our structures may be further useful as sensors or actuators in microelectromechanical systems.

  15. Centrosomal protein CP110 controls maturation of the mother centriole during cilia biogenesis

    PubMed Central

    Yadav, Sharda Prasad; Sharma, Neel Kamal; Liu, Chunqiao; Dong, Lijin; Li, Tiansen; Swaroop, Anand

    2016-01-01

    ABSTRACT Defects in cilia centrosomal genes cause pleiotropic clinical phenotypes, collectively called ciliopathies. Cilia biogenesis is initiated by the interaction of positive and negative regulators. Centriolar coiled coil protein 110 (CP110) caps the distal end of the mother centriole and is known to act as a suppressor to control the timing of ciliogenesis. Here, we demonstrate that CP110 promotes cilia formation in vivo, in contrast to findings in cultured cells. Cp110−/− mice die shortly after birth owing to organogenesis defects as in ciliopathies. Shh signaling is impaired in null embryos and primary cilia are reduced in multiple tissues. We show that CP110 is required for anchoring of basal bodies to the membrane during cilia formation. CP110 loss resulted in an abnormal distribution of core components of subdistal appendages (SDAs) and of recycling endosomes, which may be associated with premature extension of axonemal microtubules. Our data implicate CP110 in SDA assembly and ciliary vesicle docking, two requisite early steps in cilia formation. We suggest that CP110 has unique context-dependent functions, acting as both a suppressor and a promoter of ciliogenesis. PMID:26965371

  16. Effect of Fluid Viscosity on the Cilia-Generated Flow on a Mouse Tracheal Lumen.

    PubMed

    Kikuchi, Kenji; Haga, Tomofumi; Numayama-Tsuruta, Keiko; Ueno, Hironori; Ishikawa, Takuji

    2017-04-01

    Mucous flow in a tracheal lumen is generated by the beat motion of ciliated cells to provide a clearance function by discharging harmful dust particles and viruses. Due to its physiological importance, the cilia-generated flow and the rheological properties of mucus have been investigated intensively. The effects of viscosity on the cilia-generated flow, however, have not been fully clarified. In this study, we measured bulk background velocity of ciliary flow using a micro particle tracking velocimetry method under various viscosity conditions in mice. The results showed that the flow velocity decreased as the increase with viscosity of ambient fluid. Moreover, no previous study has clarified the pump power generated by cilia, which provides important information with regard to understanding the molecular motor properties of cilia. Measurements of both the ciliary flow and the ciliary motion were conducted to determine the cilia pump power. Our results indicated that the cilia pump during the effective stroke did not drive the ciliary flow efficiently under high viscosity conditions; these findings are necessary to resolve the clearance function.

  17. Phosphorylation by casein kinase 2 induces PACS-1 binding of nephrocystin and targeting to cilia

    PubMed Central

    Schermer, Bernhard; Höpker, Katja; Omran, Heymut; Ghenoiu, Cristina; Fliegauf, Manfred; Fekete, Andrea; Horvath, Judit; Köttgen, Michael; Hackl, Matthias; Zschiedrich, Stefan; Huber, Tobias B; Kramer-Zucker, Albrecht; Zentgraf, Hanswalter; Blaukat, Andree; Walz, Gerd; Benzing, Thomas

    2005-01-01

    Mutations in proteins localized to cilia and basal bodies have been implicated in a growing number of human diseases. Access of these proteins to the ciliary compartment requires targeting to the base of the cilia. However, the mechanisms involved in transport of cilia proteins to this transitional zone are elusive. Here we show that nephrocystin, a ciliary protein mutated in the most prevalent form of cystic kidney disease in childhood, is expressed in respiratory epithelial cells and accumulates at the base of cilia, overlapping with markers of the basal body area and the transition zone. Nephrocystin interacts with the phosphofurin acidic cluster sorting protein (PACS)-1. Casein kinase 2 (CK2)-mediated phosphorylation of three critical serine residues within a cluster of acidic amino acids in nephrocystin mediates PACS-1 binding, and is essential for colocalization of nephrocystin with PACS-1 at the base of cilia. Inhibition of CK2 activity abrogates this interaction and results in the loss of correct nephrocystin targeting. These data suggest that CK2-dependent transport processes represent a novel pathway of targeting proteins to the cilia. PMID:16308564

  18. Human embryonic stem cells in culture possess primary cilia with hedgehog signaling machinery

    PubMed Central

    Kiprilov, Enko N.; Awan, Aashir; Desprat, Romain; Velho, Michelle; Clement, Christian A.; Byskov, Anne Grete; Andersen, Claus Y.; Satir, Peter; Bouhassira, Eric E.; Christensen, Søren T.; Hirsch, Rhoda Elison

    2008-01-01

    Human embryonic stem cells (hESCs) are potential therapeutic tools and models of human development. With a growing interest in primary cilia in signal transduction pathways that are crucial for embryological development and tissue differentiation and interest in mechanisms regulating human hESC differentiation, demonstrating the existence of primary cilia and the localization of signaling components in undifferentiated hESCs establishes a mechanistic basis for the regulation of hESC differentiation. Using electron microscopy (EM), immunofluorescence, and confocal microscopies, we show that primary cilia are present in three undifferentiated hESC lines. EM reveals the characteristic 9 + 0 axoneme. The number and length of cilia increase after serum starvation. Important components of the hedgehog (Hh) pathway, including smoothened, patched 1 (Ptc1), and Gli1 and 2, are present in the cilia. Stimulation of the pathway results in the concerted movement of Ptc1 out of, and smoothened into, the primary cilium as well as up-regulation of GLI1 and PTC1. These findings show that hESCs contain primary cilia associated with working Hh machinery. PMID:18332216

  19. Human embryonic stem cells in culture possess primary cilia with hedgehog signaling machinery.

    PubMed

    Kiprilov, Enko N; Awan, Aashir; Desprat, Romain; Velho, Michelle; Clement, Christian A; Byskov, Anne Grete; Andersen, Claus Y; Satir, Peter; Bouhassira, Eric E; Christensen, Søren T; Hirsch, Rhoda Elison

    2008-03-10

    Human embryonic stem cells (hESCs) are potential therapeutic tools and models of human development. With a growing interest in primary cilia in signal transduction pathways that are crucial for embryological development and tissue differentiation and interest in mechanisms regulating human hESC differentiation, demonstrating the existence of primary cilia and the localization of signaling components in undifferentiated hESCs establishes a mechanistic basis for the regulation of hESC differentiation. Using electron microscopy (EM), immunofluorescence, and confocal microscopies, we show that primary cilia are present in three undifferentiated hESC lines. EM reveals the characteristic 9 + 0 axoneme. The number and length of cilia increase after serum starvation. Important components of the hedgehog (Hh) pathway, including smoothened, patched 1 (Ptc1), and Gli1 and 2, are present in the cilia. Stimulation of the pathway results in the concerted movement of Ptc1 out of, and smoothened into, the primary cilium as well as up-regulation of GLI1 and PTC1. These findings show that hESCs contain primary cilia associated with working Hh machinery.

  20. Spatial organization of cilia tufts governs airways mucus transport: Application to severe asthma

    NASA Astrophysics Data System (ADS)

    Khelloufi, Mustapha Kamel; Gras, Delphine; Chanez, Pascal; Viallat, Annie

    2014-11-01

    We study the coupling between both density and spatial repartition of beating cilia tufts, and the coordinated transport of mucus in an in-vitro epithelial model. We use a fully differentiated model epithelium in air liquid interface (ALI) obtained from endo-bronchial biopsies from healthy subjects and patients with asthma. The asthma phenotype is known to persist in the model. Mucus transport is characterized by the trajectories and velocities of microscopic beads incorporated in the mucus layer. When the beating cilia tufts density is higher than dc = 11/100 × 100 μm2 a spherical spiral coordinated mucus transport is observed over the whole ALI chamber (radius = 6 mm). Below dc, local mucus coordinated transport is observed on small circular domains on the epithelium surface. We reveal that the radii of these domains scale with the beating cilia tufts density with a power 3.7. Surprisingly, this power law is independent on cilia beat frequency, concentration and rheological properties of mucus for healthy subject and patient with asthma. The rotating or linear mucus transport is related to dispersion of the cilia tufts on the epithelium surface. We show that impaired mucus transport observed in severe asthma model epithelia is due to a drastic lack and dysfunction of cilia tufts. The author acknowledges the support of the French Agence Nationale de la Recherche (ANR) under reference ANR-13-BSV5-0015-01.

  1. A role for primary cilia in aortic valve development and disease.

    PubMed

    Toomer, Katelynn A; Fulmer, Diana; Guo, Lilong; Drohan, Alex; Peterson, Neal; Swanson, Paige; Brooks, Brittany; Mukherjee, Rupak; Body, Simon; Lipschutz, Joshua H; Wessels, Andy; Norris, Russell A

    2017-08-01

    Bicuspid aortic valve (BAV) disease is the most common congenital heart defect, affecting 0.5-1.2% of the population and causing significant morbidity and mortality. Only a few genes have been identified in pedigrees, and no single gene model explains BAV inheritance, thus supporting a complex genetic network of interacting genes. However, patients with rare syndromic diseases that stem from alterations in the structure and function of primary cilia ("ciliopathies") exhibit BAV as a frequent cardiovascular finding, suggesting primary cilia may factor broadly in disease etiology. Our data are the first to demonstrate that primary cilia are expressed on aortic valve mesenchymal cells during embryonic development and are lost as these cells differentiate into collagen-secreting fibroblastic-like cells. The function of primary cilia was tested by genetically ablating the critical ciliogenic gene Ift88. Loss of Ift88 resulted in abrogation of primary cilia and increased fibrogenic extracellular matrix (ECM) production. Consequentially, stratification of ECM boundaries normally present in the aortic valve were lost and a highly penetrant BAV phenotype was evident at birth. Our data support cilia as a novel cellular mechanism for restraining ECM production during aortic valve development and broadly implicate these structures in the etiology of BAV disease in humans. Developmental Dynamics 246:625-634, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  2. Esophageal motility disorders: medical therapy.

    PubMed

    Lacy, Brian E; Weiser, Kirsten

    2008-01-01

    Symptoms of chest pain and dysphagia are common in the adult population. Most patients initially undergo an evaluation to exclude anatomic causes (ie, esophagitis, stricture) and cardiovascular disease as the etiology of these symptoms. Patients with persistent symptoms may then be referred for specialized testing of the esophagus, including esophageal manometry. Disorders of esophageal motility, which include achalasia, diffuse esophageal spasm, nutcracker esophagus, hypertensive lower esophageal sphincter, and ineffective motility are often identified in these patients. Unfortunately, the etiology of these disorders has not been well characterized and the treatment has not been standardized. This review will briefly discuss the impact, etiology, and diagnosis of esophageal motility disorders, and then focus on the medical management of these disorders using evidence from well-designed, prospective studies, where available.

  3. The mechanism of self-organized beating of cilia

    NASA Astrophysics Data System (ADS)

    Vidyadharan, Jyothish Sulochana

    The internal structure and physical properties of cilia are well known. The relevant hydrodynamics is also well known. But the mechanism behind the coordinated activity of the dynein molecular motors is not known. Based on experimental observations, it has been concluded that this mechanism cannot be due to control from the cell body. The possible mechanism has to be self-organized and the trigger for motor activation/deactivation has to be something related to the geometry of the ciliary axoneme. This thesis critically evaluates the most widely currently cited models and suggests an alternative model for how cilia beat. From the literature we obtained wave forms of ciliary beating at different instants in the beat cycle. These instants were digitized and interpolated. From this data, we were able to calculate the hydrodynamic force distribution (external force distribution) on the cilia and the translational and rotational velocities of the cell body. Once the hydrodynamic force distribution was obtained, we calculated the internal force distribution in the cilium using an equation we derived. Once this was known, we were able to calculate parameters of the ciliary axoneme such as the dynamic stiffness. The stiffness is the ratio of the first Fourier modes of the internal force distribution and the relative sliding between the doublet microtubules that form the axoneme. We found that the first mode was the dominant one and is the one we used for calculations. We were also able to calculate the energy involved in formation and propagation of the wave that produces the ciliary beating. We discovered that the dynamic stiffness varies along the length of a cilium. We determined that in the central region of the cilium, the stiffness is almost purely imaginary which means that the sliding velocity follows the internal force generation in that region rather than sliding. We also found that in Fourier space, the flexural rigidity (kappa=EI where E is Young's modulus and

  4. Kit signaling is required for development of coordinated motility patterns in zebrafish gastrointestinal tract.

    PubMed

    Rich, Adam; Gordon, Scott; Brown, Chris; Gibbons, Simon J; Schaefer, Katherine; Hennig, Grant; Farrugia, Gianrico

    2013-06-01

    Interstitial cells of Cajal (ICC) provide a pacemaker signal for coordinated motility patterns in the mammalian gastrointestinal (GI) tract. Kit signaling is required for development and maintenance of ICC, and these cells can be identified by Kit-like immunoreactivity. The zebrafish GI tract has two distinct ICC networks similar to mammals, suggesting a similar role in the generation of GI motility; however, a functional role for Kit-positive cells in zebrafish has not been determined. Analysis of GI motility in intact zebrafish larvae was performed during development and after disruption of Kit signaling. Development of coordinated motility patterns occurred after 5 days post-fertilization (dpf) and correlated with appearance of Kit-positive cells. Disruptions of Kit signaling using the Kit antagonist imatinib mesylate, and in Sparse, a null kita mutant, also disrupted development of coordinated motility patterns. These data suggest that Kit signaling is necessary for development of coordinated motility patterns and that Kit-positive cells in zebrafish are necessary for coordinated motility patterns.

  5. No Correlates for Somatic Motility in Freeze-Fractured Hair-Cell Membranes of Lizards and Birds

    NASA Astrophysics Data System (ADS)

    Köppl, C.; Forge, A.; Manley, G. A.

    2003-02-01

    It is not known whether active processes in mammals and non-mammals are due to the same underlying mechanism. To address this, we studied the size and density of particles in hair-cell membranes in mammals, in a lizard, the Tokay gecko, and in a bird, the barn owl. We surmised that if the prominent particles described in mammalian outer-hair-cell membranes are responsible for cochlear motility, a similar occurrence in non-mammalian hair cells would argue for similar mechanisms. Particle densities differed, however, substantially from those of mammals, suggesting that non-mammals have no membrane-based motility.

  6. High-Frequency Power Gain in the Mammalian Cochlea

    NASA Astrophysics Data System (ADS)

    Maoiléidigh, Dáibhid Ó.; Hudspeth, A. J.

    2011-11-01

    Amplification in the mammalian inner ear is thought to result from a nonlinear active process known as the cochlear amplifier. Although there is much evidence that outer hair cells (OHCs) play a central role in the cochlear amplifier, the mechanism of amplification remains uncertain. In non-mammalian ears hair bundles can perform mechanical work and account for the active process in vitro, yet in the mammalian cochlea membrane-based electromotility is required for amplification in vivo. A key issue is how OHCs conduct mechanical power amplification at high frequencies. We present a physical model of a segment of the mammalian cochlea that can amplify the power of external signals. In this representation both electromotility and active hair-bundle motility are required for mechanical power gain at high frequencies. We demonstrate how the endocochlear potential, the OHC resting potential, Ca2+ gradients, and ATP-fueled myosin motors serve as the energy sources underlying mechanical power gain in the cochlear amplifier.

  7. The presence of primary cilia in cancer cells does not predict responsiveness to modulation of smoothened activity.

    PubMed

    Spann, Ashley L; Yuan, Kun; Goliwas, Kayla F; Steg, Adam D; Kaushik, Devanshu D; Kwon, Yeon-Jin; Frost, Andra R

    2015-07-01

    Primary cilia are microtubule-based organelles that regulate smoothened-dependent activation of the GLI transcription factors in canonical hedgehog signaling. In many cancers, primary cilia are markedly decreased or absent. The lack of primary cilia may inhibit or alter canonical hedgehog signaling and, thereby, interfere in the cellular responsiveness to modulators of smoothened activity. Clinical trials of smoothened antagonists for cancer treatment have shown the best response in basal cell carcinomas, with limited response in other solid tumors. To determine whether the presence or absence of primary cilia in cancer cells will predict their responsiveness to modulation of smoothened activity, we compared the ability of an agonist and/or inhibitor of smoothened (SAG and SANT1, respectively) to modulate GLI-mediated transcription, as measured by GLI1 mRNA level or GLI-luciferase reporter activity, in non-cancer cells with primary cilia (ovarian surface epithelial cells and breast fibroblasts), in cancer cells that cannot assemble primary cilia (MCF7, MDA-MB-231 cell lines), and in cancer cells with primary cilia (SKOV3, PANC1 cell lines). As expected, SAG and SANT1 resulted in appropriate modulation of GLI transcriptional activity in ciliated non-cancer cells, and failed to modulate GLI transcriptional activity in cancer cells without primary cilia. However, there was also no modulation of GLI transcriptional activity in either ciliated cancer cell line. SAG treatment of SKOV3 induced localization of smoothened to primary cilia, as assessed by immunofluorescence, even though there was no increase in GLI transcriptional activity, suggesting a defect in activation of SMO in the primary cilia or in steps later in the hedgehog pathway. In contrast to SKOV3, SAG treatment of PANC1 did not cause the localization of smoothened to primary cilia. Our data demonstrate that the presence of primary cilia in the cancer epithelial cells lines tested does not indicate their

  8. Target molecules of calmodulin on microtubules of Tetrahymena cilia

    SciTech Connect

    Hirano-Ohnishi, Junko; Watanabe, Yoshio )

    1988-09-01

    In the course of an attempt to isolate the calmodulin-binding proteins (CaMBPs) from cilia of Tetrahymena, it was found that some CaMBPs tend to interact with axonemal microtubules. The present study demonstrates this interaction by cosedimentation experiments using in vitro polymerized Tetrahymena axonemal microtubules and Tetrahymena CaMBPs purified from axonemes by calmodulin affinity column chromatography. Analysis by the ({sup 125}I)calmodulin overlay method showed that at least three CaMBPs (M{sub r} 69, 45, and 37 kDa) cosediment with microtubules. Furthermore, without any addition of exogenous CaMBPs, microtubules purified after three cycles of temperature-dependent polymerization and depolymerization included the above CaMBPs and additional CaMBPs which could not cosediment with microtubules. From the results, the authors have classified these microtubule-associated CaMBPs into two groups: (i) CaMBPs which interact with microtubules only during polymerization, and (ii) CaMBPs which interact not only with microtubules during polymerization, but also with polymerized microtubules. These results suggest that the microtubule-associated CaMBPs, especially those of the latter group, are located on the surface of ciliary microtubules, and may become the target molecules of calmodulin at Ca{sup 2+}-triggered ciliary reversal.

  9. Primary cilia mechanics affects cell mechanosensation: A computational study.

    PubMed

    Khayyeri, Hanifeh; Barreto, Sara; Lacroix, Damien

    2015-08-21

    Primary cilia (PC) are mechanical cell structures linked to the cytoskeleton and are central to how cells sense biomechanical signals from their environment. However, it is unclear exactly how PC mechanics influences cell mechanosensation. In this study we investigate how the PC mechanical characteristics are involved in the mechanotransduction process whereby cilium deflection under fluid flow induces strains on the internal cell components that regulate the cell׳s mechanosensitive response. Our investigation employs a computational approach in which a finite element model of a cell consisting of a nucleus, cytoplasm, cortex, microtubules, actin bundles and a primary cilium was used together with a finite element representation of a flow chamber. Fluid-structure interaction analysis was performed by simulating perfusion flow of 1mm/s on the cell model. Simulations of cells with different PC mechanical characteristics, showed that the length and the stiffness of PC are responsible for the transmission of mechanical stimuli to the cytoskeleton. Fluid flow deflects the cilium, with the highest strains found at the base of the PC and in the cytoplasm. The PC deflection created further strains on the cell nucleus but did not influence microtubules and actin bundles significantly. Our results indicate that PC deflection under fluid flow stimulation transmits mechanical strain primarily to other essential organelles in the cytoplasm, such as the Golgi complex, that regulate cells' mechanoresponse. The simulations further suggest that cell mechanosensitivity can be altered by targeting PC length and rigidity.

  10. Primary Cilia Integrate Hedgehog and Wnt Signaling during Tooth Development

    PubMed Central

    Liu, B.; Chen, S.; Cheng, D.; Jing, W.; Helms, J.A.

    2014-01-01

    Many ciliopathies have clinical features that include tooth malformations but how these defects come about is not clear. Here we show that genetic deletion of the motor protein Kif3a in dental mesenchyme results in an arrest in odontogenesis. Incisors are completely missing, and molars are enlarged in Wnt1Cre+Kif3afl/fl embryos. Although amelogenesis and dentinogenesis initiate in the molar tooth bud, both processes terminate prematurely. We demonstrate that loss of Kif3a in dental mesenchyme results in loss of Hedgehog signaling and gain of Wnt signaling in this same tissue. The defective dental mesenchyme then aberrantly signals to the dental epithelia, which prompts an up-regulation in the Hedgehog and Wnt responses in the epithelia and leads to multiple attempts at invagination and an expanded enamel organ. Thus, the primary cilium integrates Hedgehog and Wnt signaling between dental epithelia and mesenchyme, and this cilia-dependent integration is required for proper tooth development. PMID:24659776

  11. Primary cilia integrate hedgehog and Wnt signaling during tooth development.

    PubMed

    Liu, B; Chen, S; Cheng, D; Jing, W; Helms, J A

    2014-05-01

    Many ciliopathies have clinical features that include tooth malformations but how these defects come about is not clear. Here we show that genetic deletion of the motor protein Kif3a in dental mesenchyme results in an arrest in odontogenesis. Incisors are completely missing, and molars are enlarged in Wnt1(Cre+)Kif3a(fl/fl) embryos. Although amelogenesis and dentinogenesis initiate in the molar tooth bud, both processes terminate prematurely. We demonstrate that loss of Kif3a in dental mesenchyme results in loss of Hedgehog signaling and gain of Wnt signaling in this same tissue. The defective dental mesenchyme then aberrantly signals to the dental epithelia, which prompts an up-regulation in the Hedgehog and Wnt responses in the epithelia and leads to multiple attempts at invagination and an expanded enamel organ. Thus, the primary cilium integrates Hedgehog and Wnt signaling between dental epithelia and mesenchyme, and this cilia-dependent integration is required for proper tooth development.

  12. Motility of Electric Cable Bacteria.

    PubMed

    Bjerg, Jesper Tataru; Damgaard, Lars Riis; Holm, Simon Agner; Schramm, Andreas; Nielsen, Lars Peter

    2016-07-01

    Cable bacteria are filamentous bacteria that electrically couple sulfide oxidation and oxygen reduction at centimeter distances, and observations in sediment environments have suggested that they are motile. By time-lapse microscopy, we found that cable bacteria used gliding motility on surfaces with a highly variable speed of 0.5 ± 0.3 μm s(-1) (mean ± standard deviation) and time between reversals of 155 ± 108 s. They frequently moved forward in loops, and formation of twisted loops revealed helical rotation of the filaments. Cable bacteria responded to chemical gradients in their environment, and around the oxic-anoxic interface, they curled and piled up, with straight parts connecting back to the source of sulfide. Thus, it appears that motility serves the cable bacteria in establishing and keeping optimal connections between their distant electron donor and acceptors in a dynamic sediment environment. This study reports on the motility of cable bacteria, capable of transmitting electrons over centimeter distances. It gives us a new insight into their behavior in sediments and explains previously puzzling findings. Cable bacteria greatly influence their environment, and this article adds significantly to the body of knowledge about this organism. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Brow motility in mitochondrial myopathy.

    PubMed

    de Castro, Flávia Augusta Attié; Cruz, Antonio Augusto V; Sobreira, Cláudia Ferreira da Rosa

    2010-01-01

    To quantify the range of brow excursion in patients with mitochondrial myopathy and chronic progressive external ophthalmoplegia (CPEO). Comparative case series. Digital image processing techniques were used to quantify the upper eyelid resting position, brow excursion, and monocular eye movements (ductions) in 19 patients with mitochondrial myopathy and CPEO and in 27 healthy control subjects. All patients with CPEO had ptosis ranging from 0.6 to 8 mm. For most patients, eye motility limitation was symmetrical. Elevation was the most affected eye movement. Patient's brow motility was on average 56.7% of the motility seen in the control group, and did not correlate with age or eye motility in any direction. Seventy-six percent of the brows displayed more than 2 mm of excursion. In patients with CPEO, the occipitofrontalis muscle is less affected than the extraocular muscles. Most patients display a useful degree of brow excursion that theoretically can be used to clear the visual axis after a conservative brow suspension.

  14. Motility of Electric Cable Bacteria

    PubMed Central

    Damgaard, Lars Riis; Holm, Simon Agner; Schramm, Andreas; Nielsen, Lars Peter

    2016-01-01

    ABSTRACT Cable bacteria are filamentous bacteria that electrically couple sulfide oxidation and oxygen reduction at centimeter distances, and observations in sediment environments have suggested that they are motile. By time-lapse microscopy, we found that cable bacteria used gliding motility on surfaces with a highly variable speed of 0.5 ± 0.3 μm s−1 (mean ± standard deviation) and time between reversals of 155 ± 108 s. They frequently moved forward in loops, and formation of twisted loops revealed helical rotation of the filaments. Cable bacteria responded to chemical gradients in their environment, and around the oxic-anoxic interface, they curled and piled up, with straight parts connecting back to the source of sulfide. Thus, it appears that motility serves the cable bacteria in establishing and keeping optimal connections between their distant electron donor and acceptors in a dynamic sediment environment. IMPORTANCE This study reports on the motility of cable bacteria, capable of transmitting electrons over centimeter distances. It gives us a new insight into their behavior in sediments and explains previously puzzling findings. Cable bacteria greatly influence their environment, and this article adds significantly to the body of knowledge about this organism. PMID:27084019

  15. IFT88 plays a cilia- and PCP-independent role in controlling oriented cell divisions during vertebrate embryonic development.

    PubMed

    Borovina, Antonia; Ciruna, Brian

    2013-10-17

    The role for cilia in establishing planar cell polarity (PCP) is contentious. Although knockdown of genes known to function in ciliogenesis has been reported to cause PCP-related morphogenesis defects in zebrafish, genetic mutations affecting intraflagellar transport (IFT) do not show PCP phenotypes despite the requirement for IFT in cilia formation. This discrepancy has been attributed to off-target effects of antisense morpholino oligonucleotide (MO) injection, confounding maternal effects in zygotic mutant embryos, or an inability to distinguish between cilia-dependent versus cilia-independent protein functions. To determine the role of cilia in PCP, we generated maternal + zygotic IFT88 (MZift88) mutant zebrafish embryos, which never form cilia. We clearly demonstrate that cilia are not required to establish PCP. Rather, IFT88 plays a cilia-independent role in controlling oriented cell divisions at gastrulation and neurulation. Our results have important implications for the interpretation of cilia gene function in normal development and in disease. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  16. DNAH11 Localization in the Proximal Region of Respiratory Cilia Defines Distinct Outer Dynein Arm Complexes.

    PubMed

    Dougherty, Gerard W; Loges, Niki T; Klinkenbusch, Judith A; Olbrich, Heike; Pennekamp, Petra; Menchen, Tabea; Raidt, Johanna; Wallmeier, Julia; Werner, Claudius; Westermann, Cordula; Ruckert, Christian; Mirra, Virginia; Hjeij, Rim; Memari, Yasin; Durbin, Richard; Kolb-Kokocinski, Anja; Praveen, Kavita; Kashef, Mohammad A; Kashef, Sara; Eghtedari, Fardin; Häffner, Karsten; Valmari, Pekka; Baktai, György; Aviram, Micha; Bentur, Lea; Amirav, Israel; Davis, Erica E; Katsanis, Nicholas; Brueckner, Martina; Shaposhnykov, Artem; Pigino, Gaia; Dworniczak, Bernd; Omran, Heymut

    2016-08-01

    Primary ciliary dyskinesia (PCD) is a recessively inherited disease that leads to chronic respiratory disorders owing to impaired mucociliary clearance. Conventional transmission electron microscopy (TEM) is a diagnostic standard to identify ultrastructural defects in respiratory cilia but is not useful in approximately 30% of PCD cases, which have normal ciliary ultrastructure. DNAH11 mutations are a common cause of PCD with normal ciliary ultrastructure and hyperkinetic ciliary beating, but its pathophysiology remains poorly understood. We therefore characterized DNAH11 in human respiratory cilia by immunofluorescence microscopy (IFM) in the context of PCD. We used whole-exome and targeted next-generation sequence analysis as well as Sanger sequencing to identify and confirm eight novel loss-of-function DNAH11 mutations. We designed and validated a monoclonal antibody specific to DNAH11 and performed high-resolution IFM of both control and PCD-affected human respiratory cells, as well as samples from green fluorescent protein (GFP)-left-right dynein mice, to determine the ciliary localization of DNAH11. IFM analysis demonstrated native DNAH11 localization in only the proximal region of wild-type human respiratory cilia and loss of DNAH11 in individuals with PCD with certain loss-of-function DNAH11 mutations. GFP-left-right dynein mice confirmed proximal DNAH11 localization in tracheal cilia. DNAH11 retained proximal localization in respiratory cilia of individuals with PCD with distinct ultrastructural defects, such as the absence of outer dynein arms (ODAs). TEM tomography detected a partial reduction of ODAs in DNAH11-deficient cilia. DNAH11 mutations result in a subtle ODA defect in only the proximal region of respiratory cilia, which is detectable by IFM and TEM tomography.

  17. Primary cilia expression in bone marrow in response to mechanical stimulation in explant bioreactor culture.

    PubMed

    Coughlin, T R; Schiavi, J; Alyssa Varsanik, M; Voisin, M; Birmingham, E; Haugh, M G; McNamara, L M; Niebur, G L

    2016-07-19

    Bone marrow contains a multitude of mechanically sensitive cells that may participate in mechanotransduction. Primary cilia are sensory organelles expressed on mesenchymal stem cells (MSCs), osteoblasts, osteocytes, and other cell types that sense fluid flow in monolayer culture. In marrow, cilia could similarly facilitate the sensation of relative motion between adjacent cells or interstitial fluid. The goal of this study was to determine the response of cilia to mechanical stimulation of the marrow. Bioreactors were used to supply trabecular bone explants with low magnitude mechanical stimulation (LMMS) of 0.3 ×g at 30 Hz for 1 h/d, 5 d/week, inducing shear stresses in the marrow. Four groups were studied: unstimulated (UNSTIM), stimulated (LMMS), and with and without chloral hydrate (UNSTIM+CH and LMMS+CH, respectively), which was used to disrupt cilia. After 19 days of culture, immunohistochemistry for acetylated α-tubulin revealed that more cells expressed cilia in culture compared to in vivo controls. Stimulation decreased the number of cells expressing cilia in untreated explants, but not in CH-treated explants. MSCs represented a greater fraction of marrow cells in the untreated explants than CH-treated explants. MSCs harvested from the stimulated groups were more proliferative than in the unstimulated explants, but this effect was absent from CH treated explants. In contrast to the marrow, neither LMMS nor CH treatment affected bone formation as measured by mineralising surface. Computational models indicated that LMMS does not induce bone strain, and the reported effects were thus attributed to shear stress in the marrow. From a clinical perspective, genetic or pharmaceutical alterations of cilia expression may affect marrow health and function.

  18. DNAH11 Localization in the Proximal Region of Respiratory Cilia Defines Distinct Outer Dynein Arm Complexes

    PubMed Central

    Dougherty, Gerard W.; Loges, Niki T.; Klinkenbusch, Judith A.; Olbrich, Heike; Pennekamp, Petra; Menchen, Tabea; Raidt, Johanna; Wallmeier, Julia; Werner, Claudius; Westermann, Cordula; Ruckert, Christian; Mirra, Virginia; Hjeij, Rim; Memari, Yasin; Durbin, Richard; Kolb-Kokocinski, Anja; Praveen, Kavita; Kashef, Mohammad A.; Kashef, Sara; Eghtedari, Fardin; Häffner, Karsten; Valmari, Pekka; Baktai, György; Aviram, Micha; Bentur, Lea; Amirav, Israel; Davis, Erica E.; Katsanis, Nicholas; Brueckner, Martina; Shaposhnykov, Artem; Pigino, Gaia; Dworniczak, Bernd

    2016-01-01

    Primary ciliary dyskinesia (PCD) is a recessively inherited disease that leads to chronic respiratory disorders owing to impaired mucociliary clearance. Conventional transmission electron microscopy (TEM) is a diagnostic standard to identify ultrastructural defects in respiratory cilia but is not useful in approximately 30% of PCD cases, which have normal ciliary ultrastructure. DNAH11 mutations are a common cause of PCD with normal ciliary ultrastructure and hyperkinetic ciliary beating, but its pathophysiology remains poorly understood. We therefore characterized DNAH11 in human respiratory cilia by immunofluorescence microscopy (IFM) in the context of PCD. We used whole-exome and targeted next-generation sequence analysis as well as Sanger sequencing to identify and confirm eight novel loss-of-function DNAH11 mutations. We designed and validated a monoclonal antibody specific to DNAH11 and performed high-resolution IFM of both control and PCD-affected human respiratory cells, as well as samples from green fluorescent protein (GFP)–left–right dynein mice, to determine the ciliary localization of DNAH11. IFM analysis demonstrated native DNAH11 localization in only the proximal region of wild-type human respiratory cilia and loss of DNAH11 in individuals with PCD with certain loss-of-function DNAH11 mutations. GFP–left–right dynein mice confirmed proximal DNAH11 localization in tracheal cilia. DNAH11 retained proximal localization in respiratory cilia of individuals with PCD with distinct ultrastructural defects, such as the absence of outer dynein arms (ODAs). TEM tomography detected a partial reduction of ODAs in DNAH11-deficient cilia. DNAH11 mutations result in a subtle ODA defect in only the proximal region of respiratory cilia, which is detectable by IFM and TEM tomography. PMID:26909801

  19. Ectopic cilia associated with an orbital dermoid cyst and sinus tract: case report.

    PubMed

    Krahulík, David; Karhanová, Marta; Vaverka, Miroslav; Brychtová, Světlana; Pospíšilová, Dagmar

    2015-08-01

    Ectopic cilia are extremely rare congenital anomalies in which eyelash follicles appear in an abnormal place on the eyelid, most typically on the lateral quadrant of the anterior surface of the upper eyelid. In the majority of cases, simple surgical excision of ectopic cilia is indicated because of its cosmetic aspect. There is usually no associated medical co-morbidity with this anomaly. The authors report an unusual case of ectopic cilia associated with an orbital dermoid cyst and sinus tract. A 3-year-old boy was initially diagnosed with ectopic cilia on the left upper eyelid. There was no history of inflammation or swelling of the eyelid. An ophthalmological examination revealed only 1 mm of ptosis; no proptosis, inferior displacement, or palpable orbital mass was present. During surgical excision of the ectopic cilia, a thin sinus tract was identified, leading posteriorly to the orbit. Magnetic resonance imaging performed after the excision showed a supraorbital extraconal mass just below the roof of the left orbit. A supraorbital 2-piece craniotomy was performed with total extirpation of the dermoid cyst. The cyst was removed en bloc without damage to the extraocular muscles, but the sinus tract could no longer be identified. Follow-up MRI was performed 6 months after surgery and showed no evidence of recurrence. A follow-up ophthalmological examination showed no signs of inferior displacement or proptosis. To the best of the authors' knowledge, this case is the first reported instance of ectopic cilia associated with a dermoid cyst and sinus tract in which no typical clinical signs and symptoms of possible orbital pathology were present. This case highlights the value of radiological examination in all cases of ectopic cilia prior to surgical excision.

  20. Primary cilia regulate the osmotic stress response of renal epithelial cells through TRPM3.

    PubMed

    Siroky, Brian J; Kleene, Nancy K; Kleene, Steven J; Varnell, Charles D; Comer, Raven G; Liu, Jialiu; Lu, Lu; Pachciarz, Nolan W; Bissler, John J; Dixon, Bradley P

    2017-04-01

    Primary cilia sense environmental conditions, including osmolality, but whether cilia participate in the osmotic response in renal epithelial cells is not known. The transient receptor potential (TRP) channels TRPV4 and TRPM3 are osmoresponsive. TRPV4 localizes to cilia in certain cell types, while renal subcellular localization of TRPM3 is not known. We hypothesized that primary cilia are required for maximal activation of the osmotic response of renal epithelial cells and that ciliary TRPM3 and TRPV4 mediate that response. Ciliated [murine epithelial cells from the renal inner medullary collecting duct (mIMCD-3) and 176-5] and nonciliated (176-5Δ) renal cells expressed Trpv4 and Trpm3 Ciliary expression of TRPM3 was observed in mIMCD-3 and 176-5 cells and in wild-type mouse kidney tissue. TRPV4 was identified in cilia and apical membrane of mIMCD-3 cells by electrophysiology and in the cell body by immunofluorescence. Hyperosmolal stress at 500 mOsm/kg (via NaCl addition) induced the osmotic response genes betaine/GABA transporter (Bgt1) and aldose reductase (Akr1b3) in all ciliated cell lines. This induction was attenuated in nonciliated cells. A TRPV4 agonist abrogated Bgt1 and Akr1b3 induction in ciliated and nonciliated cells. A TRPM3 agonist attenuated Bgt1 and Akr1b3 induction in ciliated cells only. TRPM3 knockout attenuated Akr1b3 induction. Viability under osmotic stress was greater in ciliated than nonciliated cells. Akr1b3 induction was also less in nonciliated than ciliated cells when mannitol was used to induce hyperosmolal stress. These findings suggest that primary cilia are required for the maximal osmotic response in renal epithelial cells and that TRPM3 is involved in this mechanism. TRPV4 appears to modulate the osmotic response independent of cilia.

  1. Esophageal motility in eosinophilic esophagitis.

    PubMed

    Weiss, A H; Iorio, N; Schey, R

    2015-01-01

    Eosinophilic esophagitis (EoE) is characterized by eosinophilic infiltration of the esophagus and is a potential cause of dysphagia and food impaction, most commonly affecting young men. Esophageal manometry findings vary from normal motility to aperistalsis, simultaneous contractions, diffuse esophageal spasm, nutcracker esophagus or hypotonic lower esophageal sphincter (LES). It remains unclear whether esophageal dysmotility plays a significant role in the clinical symptoms of EoE. Our aim is to review the pathogenesis, diagnosis, and effect of treatment on esophageal dysmotility in EoE. A literature search utilizing the PubMed database was performed using keywords: eosinophilic esophagitis, esophageal dysmotility, motility, manometry, impedance planimetry, barium esophagogram, endoscopic ultrasound, and dysphagia. Fifteen studies, totaling 387 patients with eosinophilic esophagitis were identified as keeping in accordance with the aim of this study and included in this review. The occurrence of abnormal esophageal manometry was reported to be between 4 and 87% among patients with EoE. Esophageal motility studies have shown reduced distensibility, abnormal peristalsis, and hypotonicity of the LES in patients with EoE, which may also mimic other esophageal motility disorders such as achalasia or nutcracker esophagus. Studies have shown conflicting results regarding the presence of esophageal dysmotility and symptoms with some reports suggesting a higher rate of food impaction, while others report no correlation between motor function and dysphagia. Motility dysfunction of the esophagus in EoE has not been well reported in the literature and studies have reported conflicting evidence regarding the clinical significance of dysmotility seen in EoE. The correlation between esophageal dysmotility and symptoms of EoE remains unclear. Larger studies are needed to investigate the incidence of esophageal dysmotility, clinical implications, and effect of treatment on

  2. Ion Channels that Control Fertility in Mammalian Spermatozoa

    PubMed Central

    Navarro, Betsy; Kirichok, Yuriy; Chung, Jean-Ju; Clapham, David E.

    2015-01-01

    Whole-cell voltage clamp of mammalian spermatozoa was first achieved in 2006. This technical advance, combined with genetic deletion strategies, makes unambiguous identification of sperm ion channel currents possible. This review summarizes the ion channel currents that have been directly measured in mammalian sperm, and their physiological roles in fertilization. The predominant currents are 1) a Ca2+-selective current requiring expression of the 4 mCatSper genes, and 2) a delayed rectifier K+ current with properties most similar to mSlo3. Intracellular alkalinization activates both channels and induces hyperactivated motility. PMID:18649274

  3. Purification and Characterization of a Sperm Motility Inhibiting Factor from Caprine Epididymal Plasma

    PubMed Central

    Das, Sujoy; Saha, Sudipta; Majumder, Gopal Chandra; Dungdung, Sandhya Rekha

    2010-01-01

    Several studies have been reported on the occurrence of sperm motility inhibiting factors in the male reproductive fluids of different mammalian species, but these proteins have not been adequately purified and characterized. A novel sperm motility inhibiting factor (MIF-II) has been purified from caprine epididymal plasma (EP) by Hydroxylapatite gel adsorption chromatography, DEAE-Cellulose ion-exchange chromatography and chromatofocusing. The MIF-II has been purified to apparent homogeneity and the molecular weight estimated by Sephacryl S-300 gel filtration is 160 kDa. MIF-II is a dimeric protein, made up of two subunits each having a molecular mass of 80 kDa as shown by SDS-PAGE. The isoelectric point of MIF-II is 5.1 as determined by chromatofocusing and isoelectric focusing. It is a heat labile protein and maximal active at the pH 6.9 to 7.5. The sperm motility inhibiting protein factor at 2 µg/ml (12.5 nM) level showed maximal motility-inhibiting activity. The observation that the epididymal plasma factor lowered the intracellular cAMP level of spermatozoa in a concentration-dependent manner suggests that it may block the motility of caprine cauda spermatozoa by interfering the cAMP dependent motility function. The results revealed that the purified protein factor has the potential of sperm motility inhibition and may serve as a vaginal contraceptive. The antibody raised against the MIF-II has the potential for enhancement of forward motility of cauda-spermatozoa. This antibody may thus be useful for solving some of the problems of male infertility due to low sperm motility. PMID:20706623

  4. The sense of smell, its signalling pathways, and the dichotomy of cilia and microvilli in olfactory sensory cells

    PubMed Central

    2007-01-01

    Smell is often regarded as an ancillary perception in primates, who seem so dominated by their sense of vision. In this paper, we will portray some aspects of the significance of olfaction to human life and speculate on what evolutionary factors contribute to keeping it alive. We then outline the functional architecture of olfactory sensory neurons and their signal transduction pathways, which are the primary detectors that render olfactory perception possible. Throughout the phylogenetic tree, olfactory neurons, at their apical tip, are either decorated with cilia or with microvilli. The significance of this dichotomy is unknown. It is generally assumed that mammalian olfactory neurons are of the ciliary type only. The existance of so-called olfactory microvillar cells in mammals, however, is well documented, but their nature remains unclear and their function orphaned. This paper discusses the possibility, that in the main olfactory epithelium of mammals ciliated and microvillar sensory cells exist concurrently. We review evidence related to this hypothesis and ask, what function olfactory microvillar cells might have and what signalling mechanisms they use. PMID:17903277

  5. Secreted frizzled-related protein disrupts PCP in eye lens fiber cells that have polarised primary cilia

    PubMed Central

    Sugiyama, Yuki; Stump, Richard J. W.; Nguyen, Anke; Wen, Li; Chen, Yongjuan; Wang, Yanshu; Murdoch, Jennifer N.; Lovicu, Frank J.; McAvoy, John W.

    2009-01-01

    Planar cell polarity (PCP) signaling polarises cells along tissue axes. Although pathways involved are becoming better understood, outstanding issues include; (i) existence/identity of cues that orchestrate global polarisation in tissues, and (ii) the generality of the link between polarisation of primary cilia and asymmetric localisation of PCP proteins. Mammalian lenses are mainly comprised of epithelial-derived fiber cells. Concentrically arranged fibers are precisely aligned as they elongate along the anterior-posterior axis and orientate towards lens poles where they meet fibers from other segments to form characteristic sutures. We show that lens exhibits PCP, with each fiber cell having a apically situated cilium and in most cases this is polarised towards the anterior pole. Frizzled and other PCP proteins are also asymmetrically localised along the equatorial-anterior axis. Mutations in core PCP genes Van Gogh-like 2 and Celsr1 perturb oriented fiber alignment and suture formation. Suppression of the PCP pathway by overexpressing Sfrp2, shows that whilst local groups of fibers are often similarly oriented, they lack global orientation; consequently when local groups of fibers with different orientations meet they form multiple, small, ectopic suture-like configurations. This indicates that this extracellular inhibitor disrupts a global polarising signal that utilises a PCP-mediated mechanism to coordinate the global alignment and orientation of fibers to lens poles. PMID:19968984

  6. “Mating Behavior, Male Sensory Cilia, and Polycystins in C. elegans” Chapter

    PubMed Central

    Barr, Maureen M.

    2015-01-01

    The investigation of C. elegans males and the male-specific sensory neurons required for mating behaviors has provided insight into the molecular function of polycystins and mechanisms that are needed for polycystin ciliary localization. In humans, polycystin 1 and polycystin 2 are needed for kidney function; loss of polycystin function leads to autosomal dominant polycystic kidney disease (ADPKD). Polycystins localize to cilia in C. elegans and mammals, a finding that has guided research into ADPKD. The discovery that the polycystins form ciliary receptors in male-specific neurons needed for mating behaviors has also helped to unlock insights into two additional exciting new areas: the secretion of extracellular vesicles; and mechanisms of ciliary specialization. First, we will summarize the studies done in C. elegans regarding the expression, localization, and function of the polycystin 1 and 2 homologs, LOV-1 and PKD-2, and discuss insights gained from this basic research. Molecules that are co-expressed with the polycystins in the male-specific neurons may identify evolutionarily conserved molecular mechanisms for polycystin function and localization. We will discuss the finding that polycystins are secreted in extracellular vesicles that evoke behavioral change in males, suggesting that such vesicles provide a novel form of communication to conspecifics in the environment. In humans, polycystin-containing extracellular vesicles are secreted in urine and can be taken up by cilia, and quickly internalized. Therefore, communication by polycystin-containing extracellular vesicles may also use mechanisms that are evolutionarily conserved from nematode to human. Lastly, different cilia display structural and functional differences that specialize them for particular tasks, despite the fact that virtually all cilia are built by a conserved Intraflagellar Transport (IFT) mechanism and share some basic structural features. Comparative analysis of the male

  7. Topography of calcium phosphate ceramics regulates primary cilia length and TGF receptor recruitment associated with osteogenesis.

    PubMed

    Zhang, Jingwei; Dalbay, Melis T; Luo, Xiaoman; Vrij, Erik; Barbieri, Davide; Moroni, Lorenzo; de Bruijn, Joost D; van Blitterswijk, Clemens A; Chapple, J Paul; Knight, Martin M; Yuan, Huipin

    2017-07-15

    The surface topography of synthetic biomaterials is known to play a role in material-driven osteogenesis. Recent studies show that TGFβ signalling also initiates osteogenic differentiation. TGFβ signalling requires the recruitment of TGFβ receptors (TGFβR) to the primary cilia. In this study, we hypothesize that the surface topography of calcium phosphate ceramics regulates stem cell morphology, primary cilia structure and TGFβR recruitment to the cilium associated with osteogenic differentiation. We developed a 2D system using two types of tricalcium phosphate (TCP) ceramic discs with identical chemistry. One sample had a surface topography at micron-scale (TCP-B, with a bigger surface structure dimension) whilst the other had a surface topography at submicron scale (TCP-S, with a smaller surface structure dimension). In the absence of osteogenic differentiation factors, human bone marrow stromal cells (hBMSCs) were more spread on TCP-S than on TCP-B with alterations in actin organization and increased primary cilia prevalence and length. The cilia elongation on TCP-S was similar to that observed on glass in the presence of osteogenic media and was followed by recruitment of transforming growth factor-β RII (p-TGFβ RII) to the cilia axoneme. This was associated with enhanced osteogenic differentiation of hBMSCs on TCP-S, as shown by alkaline phosphatase activity and gene expression for key osteogenic markers in the absence of additional osteogenic growth factors. Similarly, in vivo after a 12-week intramuscular implantation in dogs, TCP-S induced bone formation while TCP-B did not. It is most likely that the surface topography of calcium phosphate ceramics regulates primary cilia length and ciliary recruitment of p-TGFβ RII associated with osteogenesis and bone formation. This bioengineering control of osteogenesis via primary cilia modulation may represent a new type of biomaterial-based ciliotherapy for orthopedic, dental and maxillofacial surgery

  8. Association of Lis1 with outer arm dynein is modulated in response to alterations in flagellar motility

    PubMed Central

    Rompolas, Panteleimon; Patel-King, Ramila S.; King, Stephen M.

    2012-01-01

    The cytoplasmic dynein regulatory factor Lis1, which induces a persistent tight binding to microtubules and allows for transport of cargoes under high-load conditions, is also present in motile cilia/flagella. We observed that Lis1 levels in flagella of Chlamydomonas strains that exhibit defective motility due to mutation of various axonemal substructures were greatly enhanced compared with wild type; this increase was absolutely dependent on the presence within the flagellum of the outer arm dynein α heavy chain/light chain 5 thioredoxin unit. To assess whether cells might interpret defective motility as a “high-load environment,” we reduced the flagellar beat frequency of wild-type cells through enhanced viscous load and by reductive stress; both treatments resulted in increased levels of flagellar Lis1, which altered the intrinsic beat frequency of the trans flagellum. Differential extraction of Lis1 from wild-type and mutant axonemes suggests that the affinity of outer arm dynein for Lis1 is directly modulated. In cytoplasm, Lis1 localized to two punctate structures, one of which was located near the base of the flagella. These data reveal that the cell actively monitors motility and dynamically modulates flagellar levels of the dynein regulatory factor Lis1 in response to imposed alterations in beat parameters. PMID:22855525

  9. Uromodulin is expressed in renal primary cilia and UMOD mutations result in decreased ciliary uromodulin expression

    PubMed Central

    Zaucke, Frank; Boehnlein, Joana M.; Steffens, Sarah; Polishchuk, Roman S.; Rampoldi, Luca; Fischer, Andreas; Pasch, Andreas; Boehm, Christoph W. A.; Baasner, Anne; Attanasio, Massimo; Hoppe, Bernd; Hopfer, Helmut; Beck, Bodo B.; Sayer, John A.; Hildebrandt, Friedhelm; Wolf, Matthias T. F.

    2010-01-01

    Uromodulin (UMOD) mutations are responsible for three autosomal dominant tubulo-interstitial nephropathies including medullary cystic kidney disease type 2 (MCKD2), familial juvenile hyperuricemic nephropathy and glomerulocystic kidney disease. Symptoms include renal salt wasting, hyperuricemia, gout, hypertension and end-stage renal disease. MCKD is part of the ‘nephronophthisis–MCKD complex’, a group of cystic kidney diseases. Both disorders have an indistinguishable histology and renal cysts are observed in either. For most genes mutated in cystic kidney disease, their proteins are expressed in the primary cilia/basal body complex. We identified seven novel UMOD mutations and were interested if UMOD protein was expressed in the primary renal cilia of human renal biopsies and if mutant UMOD would show a different expression pattern compared with that seen in control individuals. We demonstrate that UMOD is expressed in the primary cilia of renal tubules, using immunofluorescent studies in human kidney biopsy samples. The number of UMOD-positive primary cilia in UMOD patients is significantly decreased when compared with control samples. Additional immunofluorescence studies confirm ciliary expression of UMOD in cell culture. Ciliary expression of UMOD is also confirmed by electron microscopy. UMOD localization at the mitotic spindle poles and colocalization with other ciliary proteins such as nephrocystin-1 and kinesin family member 3A is demonstrated. Our data add UMOD to the group of proteins expressed in primary cilia, where mutations of the gene lead to cystic kidney disease. PMID:20172860

  10. Dynamics of self-oscillating cilia designed from active polymer gels

    NASA Astrophysics Data System (ADS)

    Dayal, Pratyush; Bhattacharya, Amitabh; Kuksenok, Olga; Balazs, Anna C.

    2012-02-01

    Using theory and simulations, we design active synthetic surfaces which are capable of replicating functionalities of biological cilia. In order to design such exquisite biomimetic systems we harness unique properties of polymer gels that undergo photosensitive Belousov-Zhabotinsky (BZ) reaction. Powered by internalized BZ reaction these polymer gels swell and de-swell autonomously by chemo-mechanical transduction and therefore are ideal materials for designing our system. In order to simulate the dynamics of the BZ cilia in surrounding fluid we have developed a nonlinear hybrid 3D model which captures elasto-dynamics of polymer gel and diffusive exchange of BZ reagents between the gel and the fluid. Here we show that the geometrical arrangement of cilia and the distribution of BZ activator in the fluid determine the dynamic response of the cilia. We further show that using light as an external stimulus we can sequentially modulate height of individual cilium and thereby create the ``piano effect''. Finally, we demonstrate that synchronized oscillations in the cilia result from the distribution of BZ-activator in the surrounding fluid. Our findings can be used to design active surfaces which can be remotely tuned depending upon the magnitude of external stimuli.

  11. Simulation by using the lattice Boltzmann method of microscopic particle motion induced by artificial cilia

    NASA Astrophysics Data System (ADS)

    Alapati, Suresh; Che, Woo Seong; Mannoor, Madhusoodanan; Suh, Yong Kweon

    2016-06-01

    In this paper, we present the results obtained from the simulation of particle motion induced by the fluid flow driven by an array of beating artificial cilia inside a micro-channel. A worm-like-chain model is used to simulate the elastic cilia, and the lattice Boltzmann equation is used to compute the fluid flow. We employ a harmonic force at the extreme tip of each cilium to actuate it. Our simulation methods are first validated by applying them to the motion of a single cilium and a freely falling sphere. After validation, we simulate the fluid flow generated by an array of beating cilia and find that a maximum flow rate is achieved at an optimum sperm number. Next, we simulate the motion of a neutrally buoyant spherical particle at this optimum sperm number by tracking the particle motion with a smoothed profile method. We address the effect of the following parameters on the particle velocity: the gap between cilia and particle, the particle size, the cilia density, and the presence of an array of intermediate particles.

  12. Specific α- and β-Tubulin Isotypes Optimize the Functions of Sensory Cilia in Caenorhabditis elegans

    PubMed Central

    Hurd, Daryl D.; Miller, Renee M.; Núñez, Lizbeth; Portman, Douglas S.

    2010-01-01

    Primary cilia have essential roles in transducing signals in eukaryotes. At their core is the ciliary axoneme, a microtubule-based structure that defines cilium morphology and provides a substrate for intraflagellar transport. However, the extent to which axonemal microtubules are specialized for sensory cilium function is unknown. In the nematode Caenorhabditis elegans, primary cilia are present at the dendritic ends of most sensory neurons, where they provide a specialized environment for the transduction of particular stimuli. Here, we find that three tubulin isotypes—the α-tubulins TBA-6 and TBA-9 and the β-tubulin TBB-4—are specifically expressed in overlapping sets of C. elegans sensory neurons and localize to the sensory cilia of these cells. Although cilia still form in mutants lacking tba-6, tba-9, and tbb-4, ciliary function is often compromised: these mutants exhibit a variety of sensory deficits as well as the mislocalization of signaling components. In at least one case, that of the CEM cephalic sensory neurons, cilium architecture is disrupted in mutants lacking specific ciliary tubulins. While there is likely to be some functional redundancy among C. elegans tubulin genes, our results indicate that specific tubulins optimize the functional properties of C. elegans sensory cilia. PMID:20421600

  13. Lithium treatment elongates primary cilia in the mouse brain and in cultured cells

    SciTech Connect

    Miyoshi, Ko; Kasahara, Kyosuke; Miyazaki, Ikuko; Asanuma, Masato

    2009-10-30

    The molecular mechanisms underlying the therapeutic effects of lithium, a first-line antimanic mood stabilizer, have not yet been fully elucidated. Treatment of the algae Chlamydomonas reinhardtii with lithium has been shown to induce elongation of their flagella, which are analogous structures to vertebrate cilia. In the mouse brain, adenylyl cyclase 3 (AC3) and certain neuropeptide receptors colocalize to the primary cilium of neuronal cells, suggesting a chemosensory function for the primary cilium in the nervous system. Here we show that lithium treatment elongates primary cilia in the mouse brain and in cultured cells. Brain sections from mice chronically fed with Li{sub 2}CO{sub 3} were subjected to immunofluorescence study. Primary cilia carrying both AC3 and the receptor for melanin-concentrating hormone (MCH) were elongated in the dorsal striatum and nucleus accumbens of lithium-fed mice, as compared to those of control animals. Moreover, lithium-treated NIH3T3 cells and cultured striatal neurons exhibited elongation of the primary cilia. The present results provide initial evidence that a psychotropic agent can affect ciliary length in the central nervous system, and furthermore suggest that lithium exerts its therapeutic effects via the upregulation of cilia-mediated MCH sensing. These findings thus contribute novel insights into the pathophysiology of bipolar mood disorder and other psychiatric diseases.

  14. Flow induced by ependymal cilia dominates near-wall cerebrospinal fluid dynamics in the lateral ventricles.

    PubMed

    Siyahhan, Bercan; Knobloch, Verena; de Zélicourt, Diane; Asgari, Mahdi; Schmid Daners, Marianne; Poulikakos, Dimos; Kurtcuoglu, Vartan

    2014-05-06

    While there is growing experimental evidence that cerebrospinal fluid (CSF) flow induced by the beating of ependymal cilia is an important factor for neuronal guidance, the respective contribution of vascular pulsation-driven macroscale oscillatory CSF flow remains unclear. This work uses computational fluid dynamics to elucidate the interplay between macroscale and cilia-induced CSF flows and their relative impact on near-wall dynamics. Physiological macroscale CSF dynamics are simulated in the ventricular space using subject-specific anatomy, wall motion and choroid plexus pulsations derived from magnetic resonance imaging. Near-wall flow is quantified in two subdomains selected from the right lateral ventricle, for which dynamic boundary conditions are extracted from the macroscale simulations. When cilia are neglected, CSF pulsation leads to periodic flow reversals along the ventricular surface, resulting in close to zero time-averaged force on the ventricle wall. The cilia promote more aligned wall shear stresses that are on average two orders of magnitude larger compared with those produced by macroscopic pulsatile flow. These findings indicate that CSF flow-mediated neuronal guidance is likely to be dominated by the action of the ependymal cilia in the lateral ventricles, whereas CSF dynamics in the centre regions of the ventricles is driven predominantly by wall motion and choroid plexus pulsation.

  15. Functional aspects of primary cilia in signaling, cell cycle and tumorigenesis

    PubMed Central

    2013-01-01

    Dysfunctional cilia underlie a broad range of cellular and tissue phenotypes and can eventually result in the development of ciliopathies: pathologically diverse diseases that range from clinically mild to highly complex and severe multi-organ failure syndromes incompatible with neonatal life. Given that virtually all cells of the human body have the capacity to generate cilia, it is likely that clinical manifestations attributed to ciliary dysfunction will increase in the years to come. Disputed but nevertheless enigmatic is the notion that at least a subset of tumor phenotypes fit within the ciliopathy disease spectrum and that cilia loss may be required for tumor progression. Contending for the centrosome renders ciliation and cell division mutually exclusive; a regulated tipping of balance promotes either process. The mechanisms involved, however, are complex. If the hypothesis that tumorigenesis results from dysfunctional cilia is true, then why do the classic ciliopathies only show limited hyperplasia at best? Although disassembly of the cilium is a prerequisite for cell proliferation, it does not intrinsically drive tumorigenesis per se. Alternatively, we will explore the emerging evidence suggesting that some tumors depend on ciliary signaling. After reviewing the structure, genesis and signaling of cilia, the various ciliopathy syndromes and their genetics, we discuss the current debate of tumorigenesis as a ciliopathy spectrum defect, and describe recent advances in this fascinating field. PMID:23628112

  16. The IFT-A complex regulates Shh signaling through cilia structure and membrane protein trafficking

    PubMed Central

    Liem, Karel F.; Ashe, Alyson; He, Mu; Satir, Peter; Moran, Jennifer; Beier, David; Wicking, Carol

    2012-01-01

    Two intraflagellar transport (IFT) complexes, IFT-A and IFT-B, build and maintain primary cilia and are required for activity of the Sonic hedgehog (Shh) pathway. A weak allele of the IFT-A gene, Ift144, caused subtle defects in cilia structure and ectopic activation of the Shh pathway. In contrast, strong loss of IFT-A, caused by either absence of Ift144 or mutations in two IFT-A genes, blocked normal ciliogenesis and decreased Shh signaling. In strong IFT-A mutants, the Shh pathway proteins Gli2, Sufu, and Kif7 localized correctly to cilia tips, suggesting that these pathway components were trafficked by IFT-B. In contrast, the membrane proteins Arl13b, ACIII, and Smo failed to localize to primary cilia in the absence of IFT-A. We propose that the increased Shh activity seen in partial loss-of-function IFT-A mutants may be a result of decreased ciliary ACIII and that the loss of Shh activity in the absence of IFT-A is a result of severe disruptions of cilia structure and membrane protein trafficking. PMID:22689656

  17. Flow induced by ependymal cilia dominates near-wall cerebrospinal fluid dynamics in the lateral ventricles

    PubMed Central

    Siyahhan, Bercan; Knobloch, Verena; de Zélicourt, Diane; Asgari, Mahdi; Schmid Daners, Marianne; Poulikakos, Dimos; Kurtcuoglu, Vartan

    2014-01-01

    While there is growing experimental evidence that cerebrospinal fluid (CSF) flow induced by the beating of ependymal cilia is an important factor for neuronal guidance, the respective contribution of vascular pulsation-driven macroscale oscillatory CSF flow remains unclear. This work uses computational fluid dynamics to elucidate the interplay between macroscale and cilia-induced CSF flows and their relative impact on near-wall dynamics. Physiological macroscale CSF dynamics are simulated in the ventricular space using subject-specific anatomy, wall motion and choroid plexus pulsations derived from magnetic resonance imaging. Near-wall flow is quantified in two subdomains selected from the right lateral ventricle, for which dynamic boundary conditions are extracted from the macroscale simulations. When cilia are neglected, CSF pulsation leads to periodic flow reversals along the ventricular surface, resulting in close to zero time-averaged force on the ventricle wall. The cilia promote more aligned wall shear stresses that are on average two orders of magnitude larger compared with those produced by macroscopic pulsatile flow. These findings indicate that CSF flow-mediated neuronal guidance is likely to be dominated by the action of the ependymal cilia in the lateral ventricles, whereas CSF dynamics in the centre regions of the ventricles is driven predominantly by wall motion and choroid plexus pulsation. PMID:24621815

  18. The IFT-A complex regulates Shh signaling through cilia structure and membrane protein trafficking.

    PubMed

    Liem, Karel F; Ashe, Alyson; He, Mu; Satir, Peter; Moran, Jennifer; Beier, David; Wicking, Carol; Anderson, Kathryn V

    2012-06-11

    Two intraflagellar transport (IFT) complexes, IFT-A and IFT-B, build and maintain primary cilia and are required for activity of the Sonic hedgehog (Shh) pathway. A weak allele of the IFT-A gene, Ift144, caused subtle defects in cilia structure and ectopic activation of the Shh pathway. In contrast, strong loss of IFT-A, caused by either absence of Ift144 or mutations in two IFT-A genes, blocked normal ciliogenesis and decreased Shh signaling. In strong IFT-A mutants, the Shh pathway proteins Gli2, Sufu, and Kif7 localized correctly to cilia tips, suggesting that these pathway components were trafficked by IFT-B. In contrast, the membrane proteins Arl13b, ACIII, and Smo failed to localize to primary cilia in the absence of IFT-A. We propose that the increased Shh activity seen in partial loss-of-function IFT-A mutants may be a result of decreased ciliary ACIII and that the loss of Shh activity in the absence of IFT-A is a result of severe disruptions of cilia structure and membrane protein trafficking.

  19. Lack of cilia and differentiation defects in the liver of human foetuses with the Meckel syndrome.

    PubMed

    Clotman, Frédéric; Libbrecht, Louis; Killingsworth, Murray C; Loo, Christine C K; Roskams, Tania; Lemaigre, Frédéric P

    2008-03-01

    Meckel syndrome is an autosomal-recessive disease characterized by a combination of renal cysts, anomalies of the central nervous system, polydactyly and ductal plate malformations (DPM), which are hepatic anomalies consisting of excessive and abnormal foetal biliary structures. Among the genomic loci associated with Meckel syndrome, mutations in four genes were recently identified. These genes code for proteins associated with primary cilia and are possibly involved in cell differentiation. The aim of the present work was to investigate the formation of the primary cilia and the differentiation of the hepatic cells in foetuses with Meckel syndrome. Sections of livers from human foetuses with Meckel syndrome were analysed by immunofluorescence, immunohistochemistry and electron microscopy. The primary cilia of the biliary cells were absent in some Meckel foetuses, but were present in others. In addition, defects in hepatic differentiation were observed in Meckel livers, as evidenced by the presence of hybrid cells co-expressing hepatocytic and biliary markers. Defects in cilia formation occur in some Meckel livers, and most cases show DPM associated with abnormal hepatic cell differentiation. Because differentiation precedes the formation of the cilia during liver development, we propose that defective differentiation may constitute the initial defect in the liver of Meckel syndrome foetuses.

  20. Self-organized cell motility

    NASA Astrophysics Data System (ADS)

    Du, Xinxin; Doubrovinski, Konstantin

    2011-03-01

    Cell migration plays a key role in a wide range of biological phenomena, such as morphogenesis, chemotaxis, and wound healing. Cell locomotion relies on the cytoskeleton, a meshwork of filamentous proteins, intrinsically out of thermodynamic equilibrium and cross-linked by molecular motors, proteins that turn chemical energy into mechanical work. In the course of locomotion, cells remain polarized, i.e. they retain a single direction of motion in the absence of external cues. Traditionally, polarization has been attributed to intracellular signaling. However, recent experiments show that polarization may be a consequence of self-organized cytoskeletal dynamics. Our aim is to elucidate the mechanisms by which persistent unidirectional locomotion may arise through simple mechanical interactions of the cytoskeletal proteins. To this end, we develop a simple physical description of cytoskeletal dynamics. We find that the proposed description accounts for a range of phenomena associated with cell motility, including spontaneous polarization, persistent unidirectional motion, and the co-existence of motile and non-motile states.

  1. Bacterial motility: From propulsion to collective behavior

    NASA Astrophysics Data System (ADS)

    Dombrowski, Christopher C.

    This work explores bacterial motility from the mechanisms of propulsion of an individual cell to the complex behavior of collective motility. The shear modulus of bacterial flagella was measured by stretching isolated flagella with an optical trap and by measuring force extension curves of the stretched flagella shedding light onto the mechanics involved in the motility of single micro-organisms. Experiments in concentrated suspensions of bacteria show collective behavior with large scale mixing on a time and length scale greater than can be understood from the standard model of "run and tumble" motility of a single organism are reported. To further understand the transition from individual to collective motility a novel form of motility where an individual bacterium can reverse direction without changing cell orientation is reported here. These experiments further the understanding of bacterial motility.

  2. New insights into an old organelle: meeting report on biology of cilia and flagella.

    PubMed

    Sengupta, Piali; Barr, Maureen M

    2014-06-01

    The rising interest of the scientific community in cilia biology was evident from the fact that registration for the third FASEB conference on 'The Biology of Cilia and Flagella' closed out before the early bird deadline. Cilia and flagella are organelles of profound medical importance; defects in their structure or function result in a plethora of human diseases called ciliopathies. 240 clinicians and basic scientists from around the world gathered from 23 June 2013 to 28 June 2013 at Sheraton at the Falls, Niagara Falls, NY to present and discuss their research on this intensely studied subcellular structure. The meeting was organized by Gregory Pazour (University of Massachusetts Medical School), Bradley Yoder (University of Alabama-Birmingham), and Maureen Barr (Rutgers University) and was sponsored by the Federation of American Societies for Experimental Biology (FASEB). Here, we report highlights, points of discussion, and emerging themes from this exciting meeting.

  3. Microtubule-depolymerizing kinesins in the regulation of assembly, disassembly, and length of cilia and flagella.

    PubMed

    Hu, Zhangfeng; Liang, Yinwen; Meng, Dan; Wang, Liang; Pan, Junmin

    2015-01-01

    Defects in ciliary assembly, maintenance, and signaling are associated with various human diseases and developmental disorders, termed ciliopathies. Eukaryotic flagella and cilia (interchangeable terms) are microtubule-based organelles. Thus, microtubule dynamics and microtubule-dependent transport are predicted to affect the structural integrity and functionality of cilia profoundly. Kinesin-2 is well known for its role in intraflagellar transport to transport ciliary precursors and signaling molecules. Recently, microtubule-depolymerizing kinesins found in kinesin-8, -13, and -14A families have emerged as regulators of cilia. We first discuss ciliary kinesins identified in the flagellar or ciliary proteome, and then focus on the function and regulation of microtubule-depolymerizing kinesins. Lastly, we review the recent advances of microtubule-depolymerizing kinesins in controlling ciliary assembly, disassembly, and length.

  4. Primary Cilia Modulate IHH Signal Transduction in Response to Hydrostatic Loading of Growth Plate Chondrocytes

    PubMed Central

    Shao, Y, Yvonne Y.; Wang, Lai; Welter, J, Jean F.; Ballock, R. Tracy

    2011-01-01

    Indian Hedgehog (Ihh) is a key component of the regulatory apparatus governing chondrocyte proliferation and differentiation in the growth plate. Recent studies have demonstrated that the primary cilium is the site of Ihh signaling within the cell, and that primary cilia are essential for bone and cartilage formation. Primary cilia are also postulated to act as mechanosensory organelles that transduce mechanical forces acting on the cell into biological signals. In this study, we used a hydrostatic compression system to examine Ihh signal transduction under the influence of mechanical load. Our results demonstrate that hydrostatic compression increased both Ihh gene expression and Ihh-responsive Gli-luciferase activity. These increases were aborted by disrupting the primary cilia structure with chloral hydrate. These results suggest that growth plate chondrocytes respond to hydrostatic loading by increasing Ihh signaling, and that the primary cilium is required for this mechano-biological signal transduction to occur. PMID:21930256

  5. Primary Cilia as a Possible Link between Left-Right Asymmetry and Neurodevelopmental Diseases

    PubMed Central

    Trulioff, Andrey; Ermakov, Alexander; Malashichev, Yegor

    2017-01-01

    Cilia have multiple functions in the development of the entire organism, and participate in the development and functioning of the central nervous system. In the last decade, studies have shown that they are implicated in the development of the visceral left-right asymmetry in different vertebrates. At the same time, some neuropsychiatric disorders, such as schizophrenia, autism, bipolar disorder, and dyslexia, are known to be associated with lateralization failure. In this review, we consider possible links in the mechanisms of determination of visceral asymmetry and brain lateralization, through cilia. We review the functions of seven genes associated with both cilia, and with neurodevelopmental diseases, keeping in mind their possible role in the establishment of the left-right brain asymmetry. PMID:28125008

  6. Inositol polyphosphate 5-phosphatases; new players in the regulation of cilia and ciliopathies.

    PubMed

    Conduit, Sarah E; Dyson, Jennifer M; Mitchell, Christina A

    2012-08-31

    Phosphoinositides regulate numerous cellular events via the recruitment and activation of multiple lipid-binding effector proteins. The precise temporal and spatial regulation of phosphoinositide signals by the co-ordinated activities of phosphoinositide kinases and phosphatases is essential for homeostasis and development. Mutations in two inositol polyphosphate 5-phosphatases, INPP5E and OCRL, cause the cerebrorenal syndromes of Joubert and Lowe's, respectively. INPP5E and OCRL exhibit overlapping phosphoinositide substrate specificity and subcellular localisation, including an association with the primary cilia. Here, we review recent studies that identify a new role for these enzymes in the regulation of primary cilia function. Joubert syndrome has been extensively linked to primary cilia defects, and Lowe's may represent a new class of 'ciliopathy associated' syndromes.

  7. The essential roles of transition fibers in the context of cilia.

    PubMed

    Wei, Qing; Ling, Kun; Hu, Jinghua

    2015-08-01

    Once thought of as a vestigial organelle, the primary cilium is now recognized as a signaling hub for key cellular pathways in vertebrate development. The recent renaissance in cilia studies significantly improved our understanding of how cilia form and function, but little is known about how ciliogenesis is initiated and how ciliary proteins enter cilia. These important ciliary events require transition fibers (TFs) that are positioned at the ciliary base as symmetric nine-bladed propeller fibrous structures. Up until recently, TFs have been the most underappreciated ciliary structures due to limited knowledge about their molecular composition and function. Here, we highlight recent advances in our understanding of TF composition and the indispensable roles of TFs in regulating the initiation of ciliogenesis and the selective import of ciliary proteins.

  8. Role for primary cilia as flow detectors in the cardiovascular system.

    PubMed

    Van der Heiden, Kim; Egorova, Anastasia D; Poelmann, Robert E; Wentzel, Jolanda J; Hierck, Beerend P

    2011-01-01

    The cardiovascular system is exposed to biochemical and biomechanical signals. Various sensors for these signals have been described and they contribute to cardiovascular development, maintenance of vessel integrity during adult life, and to pathogenesis. In the past 10years, primary cilia, membrane-covered, rod-like cellular protrusions, were discovered on multiple cell types of the cardiovascular system. Primary cilia are sensory organelles involved in several key (developmental) signaling pathways and in chemo- and mechanosensing on a myriad of cell types. In the embryonic and adult cardiovascular system, they have been demonstrated to function as shear stress sensors on endothelial cells and could act as strain sensors on smooth muscle cells and cardiomyocytes and as chemosensors on fibroblasts. This review will cover their occurrence and elaborate on established and possible functions of primary cilia in the cardiovascular system.

  9. Morphological Aspects of Ciliary Motility

    PubMed Central

    Satir, Peter

    1967-01-01

    In Elliptio complanatus lateral cilia, two distinct patterns of filament termination can be discerned. In one case, all nine filaments are present and all are single; in the second, at least one filament is missing but doublets are still present. These probably represent different configurations within one cilium in different stroke positions; to get from one to the other, some peripheral filaments must move with respect to others. The data are consistent with the hypothesis that the filaments themselves do not change length, but rather slide past one another to accommodate increasing curvature. The bent regions of the cilium are in the form of circular arcs. In a few cases, apparent displacement of filaments at the tip (Δl) can be shown to be accounted for if we assume that all differences are generated within these arcs. The displacement per degree of bend is 35 A. Regions of bent arc are initially confined to the base of the cilium but move up the shaft as straight regions appear below them. From the relationship between arc length and radius of curvature, a shaft length that is the unit that initially bends and slides may be defined. Quantal displacements of the length of one 14S dynein may perhaps occur at sites between filaments at opposite sides of such a unit as sliding occurs. PMID:6050597

  10. Fundoplication improves disordered esophageal motility.

    PubMed

    Heider, T Ryan; Behrns, Kevin E; Koruda, Mark J; Shaheen, Nicholas J; Lucktong, Tananchai A; Bradshaw, Barbara; Farrell, Timothy M

    2003-02-01

    Patients with gastroesophageal reflux disease (GERD) and disordered esophageal motility are at risk for postoperative dysphagia, and are often treated with partial (270-degree) fundoplication as a strategy to minimize postoperative swallowing difficulties. Complete (360-degree) fundoplication, however, may provide more effective and durable reflux protection over time. Recently we reported that postfundoplication dysphagia is uncommon, regardless of preoperative manometric status and type of fundoplication. To determine whether esophageal function improves after fundoplication, we measured postoperative motility in patients in whom disordered esophageal motility had been documented before fundoplication. Forty-eight of 262 patients who underwent laparoscopic fundoplication between 1995 and 2000 satisfied preoperative manometric criteria for disordered esophageal motility (distal esophageal peristaltic amplitude < or =30 mm Hg and/or peristaltic frequency < or =80%). Of these, 19 had preoperative manometric assessment at our facility and consented to repeat study. Fifteen (79%) of these patients had a complete fundoplication and four (21%) had a partial fundoplication. Each patient underwent repeat four-channel esophageal manometry 29.5 +/- 18.4 months (mean +/- SD) after fundoplication. Distal esophageal peristaltic amplitude and peristaltic frequency were compared to preoperative data by paired t test. After fundoplication, mean peristaltic amplitude in the distal esophagus increased by 47% (56.8 +/- 30.9 mm Hg to 83.5 +/- 36.5 mm Hg; P < 0.001) and peristaltic frequency improved by 33% (66.4 +/- 28.7% to 87.6 +/- 16.3%; P < 0.01). Normal esophageal motor function was present in 14 patients (74%) after fundoplication, whereas in five patients the esophageal motor function remained abnormal (2 improved, 1 worsened, and 2 remained unchanged). Three patients with preoperative peristaltic frequencies of 0%, 10%, and 20% improved to 84%, 88%, and 50%, respectively

  11. The role of Zn-alpha2 glycoprotein in sperm motility is mediated by changes in cyclic AMP.

    PubMed

    Qu, Fei; Ying, Xiaoqian; Guo, Wei; Guo, Qiangsu; Chen, Guowu; Liu, Yue; Ding, Zhide

    2007-10-01

    Sperm motility is essential for male reproduction or natural fertilization. The cyclic AMP (cAMP)/cAMP-dependent protein kinase A (PKA) signaling pathway is generally recognized as one of the significant signaling pathways in the regulation of mammalian spermatozoan motility. Since Zn-alpha2-glycoprotein (ZAG) activity in mammalian adipose tissue is mediated via the beta(3)-adrenoreceptor, with upregulation of the cAMP pathway, we hypothesize that ZAG may play the same role in sperm motility regulation, a new factor of regulation of sperm motility. Therefore, the gene encoding human ZAG was cloned and polyclonal antibodies were generated, and then laser scanning confocal microscopy and flow cytometry were employed to identify this protein in human spermatozoa. The results showed that ZAG protein was mostly localized on the pre-equatorial region covering the acrosome, neck, and middle piece of the flagellum of spermatozoa. Furthermore, using computer-assisted sperm analysis, we found that anti-human ZAG antibodies could significantly reduce the motility of human swim-up spermatozoa after 90- or 120-min incubation (P<0.05 and P<0.01 respectively), together with the decreasing of intracellular cAMP and PKA levels. In conclusion, these data suggest that ZAG is present in human spermatozoa and may be involved in the regulation of sperm motility via the cAMP/PKA signaling pathway.

  12. Longitudinal analysis of Plasmodium sporozoite motility in the dermis reveals component of blood vessel recognition

    PubMed Central

    Hopp, Christine S; Chiou, Kevin; Ragheb, Daniel RT; Salman, Ahmed M; Khan, Shahid M; Liu, Andrea J; Sinnis, Photini

    2015-01-01

    Malaria infection starts with injection of Plasmodium sporozoites by an Anopheles mosquito into the skin of the mammalian host. How sporozoites locate and enter a blood vessel is a critical, but poorly understood process. In this study, we examine sporozoite motility and their interaction with dermal blood vessels, using intravital microscopy in mice. Our data suggest that sporozoites exhibit two types of motility: in regions far from blood vessels, they exhibit ‘avascular motility’, defined by high speed and less confinement, while in the vicinity of blood vessels their motility is more constrained. We find that curvature of sporozoite tracks engaging with vasculature optimizes contact with dermal capillaries. Imaging of sporozoites with mutations in key adhesive proteins highlight the importance of the sporozoite's gliding speed and its ability to modulate adhesive properties for successful exit from the inoculation site. DOI: http://dx.doi.org/10.7554/eLife.07789.001 PMID:26271010

  13. Implementing an open-access CASA software for the assessment of stallion sperm motility: Relationship with other sperm quality parameters.

    PubMed

    Giaretta, Elisa; Munerato, Mauro; Yeste, Marc; Galeati, Giovanna; Spinaci, Marcella; Tamanini, Carlo; Mari, Gaetano; Bucci, Diego

    2017-01-01

    Setting an open-access computer assisted sperm analysis (CASA) may benefit the evaluation of motility in mammalian sperm, especially when economic constraints do not allow the use of a commercial system. There have been successful attempts to develop such a device in Zebra fish sperm and the system has been used in very few studies on mammalian spermatozoa. Against this background, the present study aimed at developing an open-access CASA system for mammalian sperm using the horse as a model and based upon the Image J software previously established for Zebra fish sperm. Along with determining the sperm progressive motility and other kinetic parameters (such as amplitude of lateral head displacement), the "results" window was adjusted to simplify subsequent statistical analyses. The path window was enriched with colored sperm trajectories on the basis of the subpopulation they belong to and a number that allowed the sperm track to be associated to the sperm motility data shown in the "results" window. Data obtained from the novel plugin (named as CASA_bgm) were compared with those of the commercial CASA Hamilton-Thorn IVOS Vers.12, through Bland Altman's plots. While the percentage of total and progressive motile sperm, VCL, VAP, VSL, LIN and STR and ALH were in agreement with those obtained with the commercial system, BCF significantly differed between the two systems probably due to their settings. Interestingly, a positive and significant correlation between the percentages of total motile sperm evaluated through CASA_bgm and those showing high mitochondrial membrane potential evaluated by JC-1 staining was found. In conclusion, CASA_bgm ImageJ plugin could be useful and reliable for stallion sperm motility analysis and it is our aim to apply this system to other mammalian species. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. An increase or a decrease in myosin II phosphorylation inhibits macrophage motility

    PubMed Central

    1991-01-01

    Myosin II purified from mammalian non-muscle cells is phosphorylated on the 20-kD light chain subunit (MLC20) by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase (MLCK). The importance of MLC20 phosphorylation in regulating cell motility was investigated by introducing either antibodies to MLCK (MK-Ab) or a Ca2+/calmodulin- independent, constitutively active form of MLCK (MK-) into macrophages. The effects of these proteins on cell motility were then determined using a quantitative chemotaxis assay. Chemotaxis is significantly diminished in macrophages containing MK-Ab compared to macrophages containing control antibodies. Moreover, there is an inverse relationship between the number of cells that migrate and the amount of MK-Ab introduced into cells. Interestingly, there is also an inverse relationship between the number of cells that migrate and the amount of MK- introduced into cells. Other experiments demonstrated that MK-Ab decreased intracellular MLC20 phosphorylation while MK- increased MLC20 phosphorylation. MK- also increased the amount of myosin associated with the cytoskeleton. These data demonstrate that the regulation of MLCK is an important aspect of cell motility and suggest that MLC20 phosphorylation must be maintained within narrow limits during translational motility by mammalian cells. PMID:2071674

  15. The N-DRC forms a conserved biochemical complex that maintains outer doublet alignment and limits microtubule sliding in motile axonemes

    PubMed Central

    Bower, Raqual; Tritschler, Douglas; VanderWaal, Kristyn; Perrone, Catherine A.; Mueller, Joshua; Fox, Laura; Sale, Winfield S.; Porter, M. E.

    2013-01-01

    The nexin–dynein regulatory complex (N-DRC) is proposed to coordinate dynein arm activity and interconnect doublet microtubules. Here we identify a conserved region in DRC4 critical for assembly of the N-DRC into the axoneme. At least 10 subunits associate with DRC4 to form a discrete complex distinct from other axonemal substructures. Transformation of drc4 mutants with epitope-tagged DRC4 rescues the motility defects and restores assembly of missing DRC subunits and associated inner-arm dyneins. Four new DRC subunits contain calcium-signaling motifs and/or AAA domains and are nearly ubiquitous in species with motile cilia. However, drc mutants are motile and maintain the 9 + 2 organization of the axoneme. To evaluate the function of the N-DRC, we analyzed ATP-induced reactivation of isolated axonemes. Rather than the reactivated bending observed with wild-type axonemes, ATP addition to drc-mutant axonemes resulted in splaying of doublets in the distal region, followed by oscillatory bending between pairs of doublets. Thus the N-DRC provides some but not all of the resistance to microtubule sliding and helps to maintain optimal alignment of doublets for productive flagellar motility. These findings provide new insights into the mechanisms that regulate motility and further highlight the importance of the proximal region of the axoneme in generating flagellar bending. PMID:23427265

  16. Emerging role of primary cilia as mechanosensors in osteocytes.

    PubMed

    Nguyen, An M; Jacobs, Christopher R

    2013-06-01

    The primary cilium is a solitary, immotile microtubule-based extension present on nearly every mammalian cell. This organelle has established mechanosensory roles in several contexts including kidney, liver, and the embryonic node. Mechanical load deflects the cilium, triggering biochemical responses. Defects in cilium function have been associated with numerous human diseases. Recent research has implicated the primary cilium as a mechanosensor in bone. In this review, we discuss the cilium, the growing evidence for its mechanosensory role in bone, and areas of future study.

  17. DYX1C1 is required for axonemal dynein assembly and ciliary motility

    PubMed Central

    Tarkar, Aarti; Loges, Niki T.; Slagle, Christopher E.; Francis, Richard; Dougherty, Gerard W.; Tamayo, Joel V.; Shook, Brett; Cantino, Marie; Schwartz, Daniel; Jahnke, Charlotte; Olbrich, Heike; Werner, Claudius; Raidt, Johanna; Pennekamp, Petra; Abouhamed, Marouan; Hjeij, Rim; Köhler, Gabriele; Griese, Matthias; Li, You; Lemke, Kristi; Klena, Nikolas; Liu, Xiaoqin; Gabriel, George; Tobita, Kimimasa; Jaspers, Martine; Morgan, Lucy C.; Shapiro, Adam J.; Letteboer, Stef J.F.; Mans, Dorus A.; Carson, Johnny L.; Leigh, Margaret W.; Wolf, Whitney E.; Chen, Serafine; Lucas, Jane S.; Onoufriadis, Alexandros; Plagnol, Vincent; Schmidts, Miriam; Boldt, Karsten; Roepman, Ronald; Zariwala, Maimoona; Lo, Cecilia W.; Mitchison, Hannah M.; Knowles, Michael R.; Burdine, Rebecca D.; LoTurco, Joseph J.; Omran, Heymut

    2014-01-01

    SUMMARY Dyx1c1 has been associated with dyslexia and neuronal migration in the developing neocortex. Unexpectedly, we found that deletion of Dyx1c1 exons 2–4 in mice caused a phenotype resembling primary ciliary dyskinesia (PCD), a genetically heterogeneous disorder characterized by chronic airway disease, laterality defects, and male infertility. This phenotype was confirmed independently in mice with a Dyx1c1c.T2A start codon mutation recovered from an ENU mutagenesis screen. Morpholinos targeting dyx1c1 in zebrafish also created laterality and ciliary motility defects. In humans, recessive loss-of-function DYX1C1 mutations were identified in twelve PCD individuals. Ultrastructural and immunofluorescence analyses of DYX1C1-mutant motile cilia in mice and humans revealed disruptions of outer and inner dynein arms (ODA/IDA). DYX1C1 localizes to the cytoplasm of respiratory epithelial cells, its interactome is enriched for molecular chaperones, and it interacts with the cytoplasmic ODA/IDA assembly factor DNAAF2/KTU. Thus, we propose that DYX1C1 is a newly identified dynein axonemal assembly factor (DNAAF4). PMID:23872636

  18. Distinctive features of cilia in metazoans and their significance for systematics.

    PubMed

    Tyler, S

    1979-01-01

    A comparative study of epidermal cilia in the Turbellaria and Nemertea has revealed features in these organelles that are specific to certain taxonomic groups. Turbellarians of the order Acoela, in particular, have a characteristic pattern of axonemal filament termination in the distal tips of their cilia and a characteristic ciliary rootlet system that is not seen in other turbellarian orders nor in other metazoans. Each epidermal cilium in acoels has a typical 9 + 2 axonemal pattern through the main part of its length, but near its distal tip there is an abrupt shelf-life narrowing at which filaments 4-7 terminate; filaments 1, 2, 8 and 9 continue into the thinner distal-most part of the shaft along with singlet microtubules from the axonemal center. The rootlet system in acoel cilia involves an interconnecting pattern with lateral connectives. The unique structure of these cilia has systematic and phylogenetic significance for the Acoela, and it is argued that ultrastructural characters in general, including characters of organelles, can be validly applied to the phylogeny and systematics of the Metazoa.

  19. Loss of Primary Cilia Upregulates Renal Hypertrophic Signaling and Promotes Cystogenesis

    PubMed Central

    Fitzgibbon, Wayne; Sas, Kelli; Stenbit, Antine E.; Amria, May; Houston, Amber; Reichert, Ryan; Gilley, Sandra; Siegal, Gene P.; Bissler, John; Bilgen, Mehmet; Chou, Peter Cheng-te; Guay-Woodford, Lisa; Yoder, Brad; Haycraft, Courtney J.; Siroky, Brian

    2011-01-01

    Primary cilia dysfunction alters renal tubular cell proliferation and differentiation and associates with accelerated cyst formation in polycystic kidney disease. However, the mechanism leading from primary ciliary dysfunction to renal cyst formation is unknown. We hypothesize that primary cilia prevent renal cyst formation by suppressing pathologic tubular cell hypertrophy and proliferation. Unilateral nephrectomy initiates tubular cell hypertrophy and proliferation in the contralateral kidney and provides a tool to examine primary cilia regulation of renal hypertrophy. Conditional knockout of the primary cilia ift88 gene leads to delayed, adult-onset renal cystic disease, which provides a window of opportunity to conduct unilateral nephrectomy and examine downstream kinetics of renal hypertrophy and cyst formation. In wild-type animals, unilateral nephrectomy activated the mTOR pathway and produced appropriate structural and functional hypertrophy without renal cyst formation. However, in ift88 conditional knockout animals, unilateral nephrectomy triggered increased renal hypertrophy and accelerated renal cyst formation, leading to renal dysfunction. mTOR signaling also increased compared with wild-type animals, suggesting a mechanistic cascade starting with primary ciliary dysfunction, leading to excessive mTOR signaling and renal hypertrophic signaling and culminating in cyst formation. These data suggest that events initiating hypertrophic signaling, such as structural or functional loss of renal mass, may accelerate progression of adult polycystic kidney disease toward end-stage renal disease. PMID:21493775

  20. The primary cilia, a 'Rab-id' transit system for hedgehog signaling.

    PubMed

    Oro, Anthony E

    2007-12-01

    Intense focus has been centered around how the primary cilia transduces the hedgehog (Hh) signal from smoothened (Smo) to the Gli transcription factors. New data indicate that ligand and signaling lipids help regulate small GTPase-dependent accumulation and activity of signaling components.

  1. Symplectic and antiplectic waves in an array of beating cilia attached to a closed body

    NASA Astrophysics Data System (ADS)

    Ghorbani, Aref; Najafi, Ali

    2017-05-01

    By taking into account the hydrodynamic interactions in a one dimensional array of model cilia attached to a no-slip cylinderical surface, we investigate their synchronized motion. We show how the emergence of metachronal waves depends on the initial state of the system and investigate the conditions under which the formation of symplectic and antiplectic waves are possible.

  2. Dynamics of Individual cilia to external loading- A simple one dimensional picture

    NASA Astrophysics Data System (ADS)

    Swaminathan, Vinay; Hill, David; Superfine, R.

    2008-10-01

    From being called the cellular janitors to swinging debauchers, cilia have captured the fascinations of researchers for over 200 years. In cystic fibrosis and chronic obstructive pulmonary disease where the cilia loses it's function, the protective mucus layer in the lung thickens and mucociliary clearance breaks down, leading to inflammation along the airways and an increased rate of infection. The mechanistic understanding of mucus clearance depends on a quantitative assessment of the axoneme dynamics and the maximum force the cilia are capable of generating and imparting to the mucus layer. Similar to the situation in molecular motors, detailed quantitative measurements of dynamics under applied load conditions are expected to be essential in developing predictive models. Based on our measurements of the dynamics of individual ciliary motion in the human bronchial epithelial cell under the application of an applied load, we present a simple one dimensional model for the axoneme dynamics and quantify the axoneme stiffness, the internal force generated by the axoneme, the stall force and show how the dynamics sheds insight on the time dependence of the internal force generation. The internal force generated by the axoneme is related to the ability of cilia to propel fluids and to their potential role in force sensing.

  3. Real-time remote control of artificial cilia actuation using fingertip drawing for efficient micromixing.

    PubMed

    Chen, Chia-Yuan; Yao, Chih-Yuan; Lin, Cheng-Yi; Hung, Shih-Hsuan

    2014-10-01

    Low-efficiency diffusion mechanism poses a significant barrier to the enhancement of micromixing efficiency in microfluidics. Actuating artificial cilia to increase the contact area of two flow streams during micromixing provides a promising alternative to enhance the mixing performance. Real-time adjustment of beating behavior in artificial cilia is necessary to accommodate various biological/chemical reagents with different hydrodynamic properties that are processed in a single microfluidic platform during micromixing. Equipping the microfluidic device with a self-troubleshooting feature for the end user, such as a bubble removal function during the process of multiple chemical solution injections, is also essential for robust micromixing. To meet these requirements, we initiated a new beating control concept by controlling the beating behavior of the artificial cilia through remote and simultaneous actuation of human fingertip drawing. A series of micromixing test cases under extreme flow conditions (Re < 10(-3)) was conducted in the designed micromixer with high mixing performance. Satisfactory micromixing efficiency was achieved even with a rapid beating trajectory of the artificial cilia actuated through the fingertip motion of end users. The analytical paradigm and results allow end users to troubleshoot technical difficulties encountered during micromixing operations.

  4. Levonorgestrel decreases cilia beat frequency of human fallopian tubes and rat oviducts without changing morphological structure.

    PubMed

    Zhao, Weihong; Zhu, Qian; Yan, Mingxing; Li, Cheng; Yuan, Jiangjing; Qin, Guojuan; Zhang, Jian

    2015-02-01

    Levonorgestrel, a derivative of progesterone, effectively protects women against unwanted pregnancy as an emergency contraceptive. Previous studies have not been successful in determining the mechanism by which levonorgestrel acts. In the present study we analysed cilia beat action and cilia morphology following levonorgestrel exposure in vitro and in vivo using both light and electron microscopy. There was a significant decrease in the ciliary beat frequency (CBF) of human fallopian tubes between mucosal explants bathed in 5 μmol/L levonorgestrel and those bathed in medium alone (P < 0.05). There was a tendency for CBF to decrease more in the ampulla than in isthmus, but there were no differences between the proliferative and secretory phases. In rat oviducts, levonorgestrel produced a similar reduction in CBF (~ 10%) compared with the saline control group (P < 0.05). Histological and ultrastructural analysis demonstrated no changes in the percentage of ciliated cells or in the classic '9 + 2' structure of cilia following levonorgestrel treatment in either system. Thus, levonorgestrel reduces CBF without damaging cilia morphology. Decreases in CBF may indicate a pathological role for levonorgestrel in the transportation of the ovum and zygote in the fallopian tube. © 2014 Wiley Publishing Asia Pty Ltd.

  5. A Cilia Independent Role of Ift88/Polaris during Cell Migration

    PubMed Central

    Hamann, Christoph; Powelske, Christian; Mergen, Miriam; Herbst, Henriette; Kotsis, Fruzsina; Nitschke, Roland; Kuehn, E. Wolfgang

    2015-01-01

    Ift88 is a central component of the intraflagellar transport (Ift) complex B, essential for the building of cilia and flagella from single cell organisms to mammals. Loss of Ift88 results in the absence of cilia and causes left-right asymmetry defects, disordered Hedgehog signaling, and polycystic kidney disease, all of which are explained by aberrant ciliary function. In addition, a number of extraciliary functions of Ift88 have been described that affect the cell-cycle, mitosis, and targeting of the T-cell receptor to the immunological synapse. Similarly, another essential ciliary molecule, the kinesin-2 subunit Kif3a, which transports Ift-B in the cilium, affects microtubule (MT) dynamics at the leading edge of migrating cells independently of cilia. We now show that loss of Ift88 impairs cell migration irrespective of cilia. Ift88 is required for the polarization of migrating MDCK cells, and Ift88 depleted cells have fewer MTs at the leading edge. Neither MT dynamics nor MT nucleation are dependent on Ift88. Our findings dissociate the function of Ift88 from Kif3a outside the cilium and suggest a novel extraciliary function for Ift88. Future studies need to address what unifying mechanism underlies the different extraciliary functions of Ift88. PMID:26465598

  6. Olfactory response termination involves Ca2+-ATPase in vertebrate olfactory receptor neuron cilia

    PubMed Central

    Antolin, Salome; Reisert, Johannes

    2010-01-01

    In vertebrate olfactory receptor neurons (ORNs), odorant-induced activation of the transduction cascade culminates in production of cyclic AMP, which opens cyclic nucleotide–gated channels in the ciliary membrane enabling Ca2+ influx. The ensuing elevation of the intraciliary Ca2+ concentration opens Ca2+-activated Cl− channels, which mediate an excitatory Cl− efflux from the cilia. In order for the response to terminate, the Cl− channel must close, which requires that the intraciliary Ca2+ concentration return to basal levels. Hitherto, the extrusion of Ca2+ from the cilia has been thought to depend principally on a Na+–Ca2+ exchanger. In this study, we show using simultaneous suction pipette recording and Ca2+-sensitive dye fluorescence measurements that in fire salamander ORNs, withdrawal of external Na+ from the solution bathing the cilia, which incapacitates Na+–Ca2+exchange, has only a modest effect on the recovery of the electrical response and the accompanying decay of intraciliary Ca2+ concentration. In contrast, exposure of the cilia to vanadate or carboxyeosin, a manipulation designed to block Ca2+-ATPase, has a substantial effect on response recovery kinetics. Therefore, we conclude that Ca2+-ATPase contributes to Ca2+ extrusion in ORNs, and that Na+–Ca2+exchange makes only a modest contribution to Ca2+ homeostasis in this species. PMID:20351061

  7. INTU is essential for oncogenic Hh signaling through regulating primary cilia formation in basal cell carcinoma.

    PubMed

    Yang, N; Leung, E L-H; Liu, C; Li, L; Eguether, T; Jun Yao, X-J; Jones, E C; Norris, D A; Liu, A; Clark, R A; Roop, D R; Pazour, G J; Shroyer, K R; Chen, J

    2017-08-31

    Inturned (INTU), a cilia and planar polarity effector, performs prominent ciliogenic functions during morphogenesis, such as in the skin. INTU is expressed in adult tissues but its role in tissue maintenance is unknown. Here, we report that the expression of the INTU gene is aberrantly elevated in human basal cell carcinoma (BCC), coinciding with increased primary cilia formation and activated hedgehog (Hh) signaling. Disrupting Intu in an oncogenic mutant Smo (SmoM2)-driven BCC mouse model prevented the formation of BCC through suppressing primary cilia formation and Hh signaling, suggesting that Intu performs a permissive role during BCC formation. INTU is essential for intraflagellar transport A complex assembly during ciliogenesis. To further determine whether Intu is directly involved in the activation of Hh signaling downstream of ciliogenesis, we examined the Hh signaling pathway in mouse embryonic fibroblasts, which readily responds to the Hh pathway activation. Depleting Intu blocked Smo agonist-induced Hh pathway activation, whereas the expression of Gli2ΔN, a constitutively active Gli2, restored Hh pathway activation in Intu-deficient cells, suggesting that INTU functions upstream of Gli2 activation. In contrast, overexpressing Intu did not promote ciliogenesis or Hh signaling. Taken together, data obtained from this study suggest that INTU is indispensable during BCC tumorigenesis and that its aberrant upregulation is likely a prerequisite for primary cilia formation during Hh-dependent tumorigenesis.

  8. The retrograde IFT machinery of C. elegans cilia: two IFT dynein complexes?

    PubMed

    Hao, Limin; Efimenko, Evgeni; Swoboda, Peter; Scholey, Jonathan M

    2011-01-01

    We analyzed the relatively poorly understood IFT-dynein (class DYNC2)-driven retrograde IFT pathway in C. elegans cilia, which yielded results that are surprising in the context of current models of IFT. Assays of C. elegans dynein gene expression and intraflagellar transport (IFT) suggest that conventional IFT-dynein contains essential heavy (CHE-3), light-intermediate (XBX-1), plus three light polypeptide chains that participate in IFT, but no "essential" intermediate chain. IFT assays of XBX-1::YFP suggest that IFT-dynein is transported as cargo to the distal tip of the cilium by kinesin-2 motors, but independent of the IFT-particle/BBSome complexes. Finally, we were surprised to find that the subset of cilia present on the OLQ (outer labial quadrant) neurons assemble independently of conventional "CHE-3" IFT-dynein, implying that there is a second IFT-dynein acting in these cilia. We have found a novel gene encoding a dynein heavy chain, DHC-3, and two light chains, in OLQ neurons, which could constitute an IFT-dynein complex in OLQ neuronal cilia. Our results underscore several surprising features of retrograde IFT that require clarification.

  9. Spatial Distribution of Calcium-Gated Chloride Channels in Olfactory Cilia

    PubMed Central

    French, Donald A.; Badamdorj, Dorjsuren; Kleene, Steven J.

    2010-01-01

    Background In vertebrate olfactory receptor neurons, sensory cilia transduce odor stimuli into changes in neuronal membrane potential. The voltage changes are primarily caused by the sequential openings of two types of channel: a cyclic-nucleotide-gated (CNG) cationic channel and a calcium-gated chloride channel. In frog, the cilia are 25 to 200 µm in length, so the spatial distributions of the channels may be an important determinant of odor sensitivity. Principal Findings To determine the spatial distribution of the chloride channels, we recorded from single cilia as calcium was allowed to diffuse down the length of the cilium and activate the channels. A computational model of this experiment allowed an estimate of the spatial distribution of the chloride channels. On average, the channels were concentrated in a narrow band centered at a distance of 29% of the ciliary length, measured from the base of the cilium. This matches the location of the CNG channels determined previously. This non-uniform distribution of transduction proteins is consistent with similar findings in other cilia. Conclusions On average, the two types of olfactory transduction channel are concentrated in the same region of the cilium. This may contribute to the efficient detection of weak stimuli. PMID:21209888

  10. Mechanism of Actin-Based Motility

    NASA Astrophysics Data System (ADS)

    Pantaloni, Dominique; Le Clainche, Christophe; Carlier, Marie-France

    2001-05-01

    Spatially controlled polymerization of actin is at the origin of cell motility and is responsible for the formation of cellular protrusions like lamellipodia. The pathogens Listeria monocytogenes and Shigella flexneri, which undergo actin-based propulsion, are acknowledged models of the leading edge of lamellipodia. Actin-based motility of the bacteria or of functionalized microspheres can be reconstituted in vitro from only five pure proteins. Movement results from the regulated site-directed treadmilling of actin filaments, consistent with observations of actin dynamics in living motile cells and with the biochemical properties of the components of the synthetic motility medium.

  11. Genetic Analysis Reveals a Hierarchy of Interactions between Polycystin-Encoding Genes and Genes Controlling Cilia Function during Left-Right Determination.

    PubMed

    Grimes, Daniel T; Keynton, Jennifer L; Buenavista, Maria T; Jin, Xingjian; Patel, Saloni H; Kyosuke, Shinohara; Vibert, Jennifer; Williams, Debbie J; Hamada, Hiroshi; Hussain, Rohanah; Nauli, Surya M; Norris, Dominic P

    2016-06-01

    During mammalian development, left-right (L-R) asymmetry is established by a cilia-driven leftward fluid flow within a midline embryonic cavity called the node. This 'nodal flow' is detected by peripherally-located crown cells that each assemble a primary cilium which contain the putative Ca2+ channel PKD2. The interaction of flow and crown cell cilia promotes left side-specific expression of Nodal in the lateral plate mesoderm (LPM). Whilst the PKD2-interacting protein PKD1L1 has also been implicated in L-R patterning, the underlying mechanism by which flow is detected and the genetic relationship between Polycystin function and asymmetric gene expression remains unknown. Here, we characterize a Pkd1l1 mutant line in which Nodal is activated bilaterally, suggesting that PKD1L1 is not required for LPM Nodal pathway activation per se, but rather to restrict Nodal to the left side downstream of nodal flow. Epistasis analysis shows that Pkd1l1 acts as an upstream genetic repressor of Pkd2. This study therefore provides a genetic pathway for the early stages of L-R determination. Moreover, using a system in which cultured cells are supplied artificial flow, we demonstrate that PKD1L1 is sufficient to mediate a Ca2+ signaling response after flow stimulation. Finally, we show that an extracellular PKD domain within PKD1L1 is crucial for PKD1L1 function; as such, destabilizing the domain causes L-R defects in the mouse. Our demonstration that PKD1L1 protein can mediate a response to flow coheres with a mechanosensation model of flow sensation in which the force of fluid flow drives asymmetric gene expression in the embryo.

  12. Genetic Analysis Reveals a Hierarchy of Interactions between Polycystin-Encoding Genes and Genes Controlling Cilia Function during Left-Right Determination

    PubMed Central

    Grimes, Daniel T.; Keynton, Jennifer L.; Buenavista, Maria T.; Jin, Xingjian; Patel, Saloni H.; Kyosuke, Shinohara; Williams, Debbie J.; Hamada, Hiroshi; Hussain, Rohanah; Nauli, Surya M.; Norris, Dominic P.

    2016-01-01

    During mammalian development, left-right (L-R) asymmetry is established by a cilia-driven leftward fluid flow within a midline embryonic cavity called the node. This ‘nodal flow’ is detected by peripherally-located crown cells that each assemble a primary cilium which contain the putative Ca2+ channel PKD2. The interaction of flow and crown cell cilia promotes left side-specific expression of Nodal in the lateral plate mesoderm (LPM). Whilst the PKD2-interacting protein PKD1L1 has also been implicated in L-R patterning, the underlying mechanism by which flow is detected and the genetic relationship between Polycystin function and asymmetric gene expression remains unknown. Here, we characterize a Pkd1l1 mutant line in which Nodal is activated bilaterally, suggesting that PKD1L1 is not required for LPM Nodal pathway activation per se, but rather to restrict Nodal to the left side downstream of nodal flow. Epistasis analysis shows that Pkd1l1 acts as an upstream genetic repressor of Pkd2. This study therefore provides a genetic pathway for the early stages of L-R determination. Moreover, using a system in which cultured cells are supplied artificial flow, we demonstrate that PKD1L1 is sufficient to mediate a Ca2+ signaling response after flow stimulation. Finally, we show that an extracellular PKD domain within PKD1L1 is crucial for PKD1L1 function; as such, destabilizing the domain causes L-R defects in the mouse. Our demonstration that PKD1L1 protein can mediate a response to flow coheres with a mechanosensation model of flow sensation in which the force of fluid flow drives asymmetric gene expression in the embryo. PMID:27272319

  13. A WASp-binding type II phosphatidylinositol 4-kinase required for actin polymerization-driven endosome motility

    PubMed Central

    Chang, Fanny S.; Han, Gil-Soo; Carman, George M.; Blumer, Kendall J.

    2005-01-01

    Endosomes in yeast have been hypothesized to move through the cytoplasm by the momentum gained after actin polymerization has driven endosome abscision from the plasma membrane. Alternatively, after abscission, ongoing actin polymerization on endosomes could power transport. Here, we tested these hypotheses by showing that the Arp2/3 complex activation domain (WCA) of Las17 (Wiskott-Aldrich syndrome protein [WASp] homologue) fused to an endocytic cargo protein (Ste2) rescued endosome motility in las17ΔWCA mutants, and that capping actin filament barbed ends inhibited endosome motility but not endocytic internalization. Motility therefore requires continual actin polymerization on endosomes. We also explored how Las17 is regulated. Endosome motility required the Las17-binding protein Lsb6, a type II phosphatidylinositol 4-kinase. Catalytically inactive Lsb6 interacted with Las17 and promoted endosome motility. Lsb6 therefore is a novel regulator of Las17 that mediates endosome motility independent of phosphatidylinositol 4-phosphate synthesis. Mammalian type II phosphatidylinositol 4-kinases may regulate WASp proteins and endosome motility. PMID:16216926

  14. A WASp-binding type II phosphatidylinositol 4-kinase required for actin polymerization-driven endosome motility.

    PubMed

    Chang, Fanny S; Han, Gil-Soo; Carman, George M; Blumer, Kendall J

    2005-10-10

    Endosomes in yeast have been hypothesized to move through the cytoplasm by the momentum gained after actin polymerization has driven endosome abscision from the plasma membrane. Alternatively, after abscission, ongoing actin polymerization on endosomes could power transport. Here, we tested these hypotheses by showing that the Arp2/3 complex activation domain (WCA) of Las17 (Wiskott-Aldrich syndrome protein [WASp] homologue) fused to an endocytic cargo protein (Ste2) rescued endosome motility in las17DeltaWCA mutants, and that capping actin filament barbed ends inhibited endosome motility but not endocytic internalization. Motility therefore requires continual actin polymerization on endosomes. We also explored how Las17 is regulated. Endosome motility required the Las17-binding protein Lsb6, a type II phosphatidylinositol 4-kinase. Catalytically inactive Lsb6 interacted with Las17 and promoted endosome motility. Lsb6 therefore is a novel regulator of Las17 that mediates endosome motility independent of phosphatidylinositol 4-phosphate synthesis. Mammalian type II phosphatidylinositol 4-kinases may regulate WASp proteins and endosome motility.

  15. The effect of halothane and pentobarbital sodium on brain ependymal cilia

    PubMed Central

    2012-01-01

    Background The effect of anesthetic agents on ependymal ciliary function is unknown. The aim of this study was to determine the effect of halothane and pentobarbital sodium on brain ependymal ciliary function. Methods We used an ex vivo rat brain slice model to measure ependymal ciliary beat frequency by high speed video photography at 37°C. Results Exposure to halothane caused a significant reduction in ciliary beat frequency of 2 % (P = 0.006), 15.5 % (P < 0.001), and 21.5 % (P < 0.001) for halothane concentrations of 1.8 %, 3.4 % and 4.4 %, respectively, compared to controls. Following a one-hour wash-out period, there was no significant difference between control samples and cilia that had been exposed to 1.8 % (P = 0.5) and 3.4 % (P = 0.3) halothane. The beat frequency of cilia exposed to 4.4 % halothane had increased following the wash-out period but cilia were still beating significantly more slowly than cilia from the control group (P = <0.001). Pentobarbitone at concentrations of 25 and 50 μg/ml had no effect on ciliary beat frequency compared to controls (P = 0.6 and 0.4 respectively). A significant (P = 0.002) decrease in ciliary beat frequency was seen following incubation with a pentobarbitone concentration of 250 μg/ml (mean (SD) frequency, 24(8) Hz compared to controls, 38(9) Hz). Conclusions Halothane reversibly inhibits the rate at which ependymal cilia beat. Pentobarbitone has no effect on ciliary activity at levels used for anesthesia. It is unclear whether the slowing of ependymal ciliary by halothane is responsible for some of the secondary central nervous system effects of volatile anesthetic agents. PMID:23351190

  16. Current topics of functional links between primary cilia and cell cycle.

    PubMed

    Izawa, Ichiro; Goto, Hidemasa; Kasahara, Kousuke; Inagaki, Masaki

    2015-01-01

    Primary cilia, microtubule-based sensory structures, orchestrate various critical signals during development and tissue homeostasis. In view of the rising interest into the reciprocal link between ciliogenesis and cell cycle, we discuss here several recent advances to understand the molecular link between the individual step of ciliogenesis and cell cycle control. At the onset of ciliogenesis (the transition from centrosome to basal body), distal appendage proteins have been established as components indispensable for the docking of vesicles at the mother centriole. In the initial step of axonemal extension, CP110, Ofd1, and trichoplein, key negative regulators of ciliogenesis, are found to be removed by a kinase-dependent mechanism, autophagy, and ubiquitin-proteasome system, respectively. Of note, their disposal functions as a restriction point to decide that the axonemal nucleation and extension begin. In the elongation step, Nde1, a negative regulator of ciliary length, is revealed to be ubiquitylated and degraded by CDK5-SCF(Fbw7) in a cell cycle-dependent manner. With regard to ciliary length control, it has been uncovered in flagellar shortening of Chlamydomonas that cilia itself transmit a ciliary length signal to cytoplasm. At the ciliary resorption step upon cell cycle re-entry, cilia are found to be disassembled not only by Aurora A-HDAC6 pathway but also by Nek2-Kif24 and Plk1-Kif2A pathways through their microtubule-depolymerizing activity. On the other hand, it is becoming evident that the presence of primary cilia itself functions as a structural checkpoint for cell cycle re-entry. These data suggest that ciliogenesis and cell cycle intimately link each other, and further elucidation of these mechanisms will contribute to understanding the pathology of cilia-related disease including cancer and discovering targets of therapeutic interventions.

  17. Putative odour receptors localize in cilia of olfactory receptor cells in rat and mouse: a freeze-substitution ultrastructural study.

    PubMed

    Menco, B P; Cunningham, A M; Qasba, P; Levy, N; Reed, R R

    1997-05-01

    Two different polyclonal antibodies were raised to synthetic peptides corresponding to distinct putative odour receptors of rat and mouse. Both antibodies selectively labelled olfactory cilia as seen with cryofixation and immunogold ultrastructural procedures. Regions of the olfactory organ where label was detected were consistent with those found at LM levels. Immunopositive cells were rare; only up to about 0.4% of these receptor cells were labelled. Despite chemical, species, and topographic differences both antibodies behaved identically in their ultrastructural labelling patterns. For both antibodies, labelling was very specific for olfactory cilia; both bound amply to the thick proximal and the thinner and long distal parts of the cilia. Dendritic knobs showed little labelling if any. Dendritic receptor cell structures below the knobs, supporting cell structures, and respiratory cilia did not immunolabel. There were no obvious differences in morphology between labelled and unlabelled receptor cells and their cilia. Labelling could be followed up to a distance of about 15 microns from the knobs along the distal parts of the cilia. When labelled cells were observed, this signal was detectable in two, sometimes three, sections taken through these cells while being consistently absent in neighbouring cells. This pattern argues strongly for the specificity of the labelling. In conclusion, very few receptor cells labelled with the antibodies to putative odour receptors. Additionally the olfactory cilia, the cellular regions that first encounter odour molecules and that are thought to transduce the odorous signal, displayed the most intense labelling with both antibodies. Consequently, the results showed these cilia as having many copies of the putative receptors. Finally, similar patterns of subcellular labelling were displayed in two different species, despite the use of different antibodies. Thus, this study provides compelling evidence that the heptahelical

  18. Putative odour receptors localize in cilia of olfactory receptor cells in rat and mouse: a freeze-substitution ultrastructural study.

    PubMed

    Menco, B P; Cunningham, A M; Qasba, P; Levy, N; Reed, R R

    1997-10-01

    Two different polyclonal antibodies were raised to synthetic peptides corresponding to distinct putative odour receptors of rat and mouse. Both antibodies selectively labelled olfactory cilia as seen with cryofixation and immunogold ultrastructural procedures. Regions of the olfactory organ where label was detected were consistent with those found at LM levels. Immunopositive cells were rare; only up to about 0.4% of these receptor cells were labelled. Despite chemical, species, and topographic differences both antibodies behaved identically in their ultrastructural labelling patterns. For both antibodies, labelling was very specific for olfactory cilia; both bound amply to the thick proximal and the thinner and long distal parts of the cilia. Dendritic knobs showed little labelling if any. Dendritic receptor cell structures below the knobs, supporting cell structures, and respiratory cilia did not immunolabel. There were no obvious differences in morphology between labelled and unlabelled receptor cells and their cilia. Labelling could be followed up to a distance of about 15 microns from the knobs along the distal parts of the cilia. When labelled cells were observed, this signal was detectable in two, sometimes three, sections taken through these cells while being consistently absent in neighbouring cells. This pattern argues strongly for the specificity of the labelling. In conclusion, very few receptor cells labelled with the antibodies to putative odour receptors. Additionally the olfactory cilia, the cellular regions that first encounter odour molecules and that are thought to transduce the odorous signal, displayed the most intense labelling with both antibodies. Consequently, the results showed these cilia as having many copies of the putative receptors. Finally, similar patterns of subcellular labelling were displayed in two different species, despite the use of different antibodies. Thus, this study provides compelling evidence that the heptahelical

  19. Langevin Dynamics Deciphers the Motility Pattern of Swimming Parasites

    NASA Astrophysics Data System (ADS)

    Zaburdaev, Vasily; Uppaluri, Sravanti; Pfohl, Thomas; Engstler, Markus; Friedrich, Rudolf; Stark, Holger

    2011-05-01

    The parasite African trypanosome swims in the bloodstream of mammals and causes the highly dangerous human sleeping sickness. Cell motility is essential for the parasite’s survival within the mammalian host. We present an analysis of the random-walk pattern of a swimming trypanosome. From experimental time-autocorrelation functions for the direction of motion we identify two relaxation times that differ by an order of magnitude. They originate from the rapid deformations of the cell body and a slower rotational diffusion of the average swimming direction. Velocity fluctuations are athermal and increase for faster cells whose trajectories are also straighter. We demonstrate that such a complex dynamics is captured by two decoupled Langevin equations that decipher the complex trajectory pattern by referring it to the microscopic details of cell behavior.

  20. Coupling Active Hair Bundle Mechanics, Fast Adaptation, and Somatic Motility in a Cochlear Model

    PubMed Central

    Meaud, Julien; Grosh, Karl

    2011-01-01

    One of the central questions in the biophysics of the mammalian cochlea is determining the contributions of the two active processes, prestin-based somatic motility and hair bundle (HB) motility, to cochlear amplification. HB force generation is linked to fast adaptation of the transduction current via a calcium-dependent process and somatic force generation is driven by the depolarization caused by the transduction current. In this article, we construct a global mechanical-electrical-acoustical mathematical model of the cochlea based on a three-dimensional fluid representation. The global cochlear model is coupled to linearizations of nonlinear somatic motility and HB activity as well as to the micromechanics of the passive structural and electrical elements of the cochlea. We find that the active HB force alone is not sufficient to power high frequency cochlear amplification. However, somatic motility can overcome resistor-capacitor filtering by the basolateral membrane and deliver sufficient mechanical energy for amplification at basal locations. The results suggest a new theory for high frequency active cochlear mechanics, in which fast adaptation controls the transduction channel sensitivity and thereby the magnitude of the energy delivered by somatic motility. PMID:21641302

  1. The flagellar motility of Chlamydomonas pf25 mutant lacking an AKAP-binding protein is overtly sensitive to medium conditions.

    PubMed

    Yang, Chun; Yang, Pinfen

    2006-01-01

    Radial spokes are a conserved axonemal structural complex postulated to regulate the motility of 9 + 2 cilia and flagella via a network of phosphoenzymes and regulatory proteins. Consistently, a Chlamydomonas radial spoke protein, RSP3, has been identified by RII overlays as an A-kinase anchoring protein (AKAP) that localizes the cAMP-dependent protein kinase (PKA) holoenzyme by binding to the RIIa domain of PKA RII subunit. However, the highly conserved docking domain of PKA is also found in the N termini of several AKAP-binding proteins unrelated to PKA as well as a 24-kDa novel spoke protein, RSP11. Here, we report that RSP11 binds to RSP3 directly in vitro and colocalizes with RSP3 toward the spoke base near outer doublets and dynein motors in axonemes. Importantly, RSP11 mutant pf25 displays a spectrum of motility, from paralysis with flaccid or twitching flagella as other spoke mutants to wildtype-like swimming. The wide range of motility changes reversibly depending on the condition of liquid media without replacing defective proteins. We postulate that radial spokes use the RIIa/AKAP module to regulate ciliary and flagellar beating; absence of the spoke RIIa protein exposes a medium-sensitive regulatory mechanism that is not obvious in wild-type Chlamydomonas.

  2. 21 CFR 876.1725 - Gastrointestinal motility monitoring system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Gastrointestinal motility monitoring system. 876... Gastrointestinal motility monitoring system. (a) Identification. A gastrointestinal motility monitoring system is a... esophageal motility monitor and tube, the gastrointestinal motility (electrical) system, and...

  3. 21 CFR 876.1725 - Gastrointestinal motility monitoring system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Gastrointestinal motility monitoring system. 876... Gastrointestinal motility monitoring system. (a) Identification. A gastrointestinal motility monitoring system is a... esophageal motility monitor and tube, the gastrointestinal motility (electrical) system, and...

  4. 21 CFR 876.1725 - Gastrointestinal motility monitoring system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Gastrointestinal motility monitoring system. 876... Gastrointestinal motility monitoring system. (a) Identification. A gastrointestinal motility monitoring system is a... esophageal motility monitor and tube, the gastrointestinal motility (electrical) system, and...

  5. Automated real-time measurement of chemotactic cell motility.

    PubMed

    Hadjout, N; Laevsky, G; Knecht, D A; Lynes, M A

    2001-11-01

    We have developed a novel method, (ECIS/taxis), for monitoring cell movement in response to chemotactic and chemokinetic factors. In this system, cells migrate in an under-agarose environment, and their positions are monitored using the electric cell-substrate impedance sensor technology to measure the impedance change at a target electrode, that is lithographed onto the substrate, as the cells arrive at the target. In the studies reported here, Dictyostelium discoideum was used as a prototypical, motile eukaryotic cell. Using the ECIS/taxis system, the arrival of cells at the target electrode was proportional to the dose offolate used to stimulate the cells and could be assessed by changes in resistance at the electrode. ECIS/taxis was readily able to distinguish between wild-type cells and a mutant that is deficient in its chemotactic response. Finally, we have shown that an agent that interferes with chemotactic motility leads to the delayed arrival of cells at the target electrode. The multi-well assay configuration allows for simultaneous automated screening of many samples for chemotactic or anti-chemotactic activity. This assay system is compatible with measurements of mammalian cell movement and should be valuable in the assessment of both agonists and antagonists of cell movement.

  6. miR-34/449 miRNAs are required for motile ciliogenesis by repressing cp110

    PubMed Central

    Song, Rui; Walentek, Peter; Sponer, Nicole; Klimke, Alexander; Lee, Joon Sub; Dixon, Gary; Harland, Richard; Wan, Ying; Lishko, Polina; Lize, Muriel; Kessel, Michael; He, Lin

    2014-01-01

    Summary The miR-34/449 family consists of six homologous miRNAs at three genomic loci. Redundancy of miR-34/449 miRNAs and their dominant expression in multiciliated epithelia suggest a functional significance in ciliogenesis. Here, we report that mice deficient for all miR-34/449 miRNAs exhibited postnatal mortality, infertility, and strong respiratory dysfunction caused by defective mucociliary clearance. In both mouse and Xenopus, miR-34/449-deficient multiciliated cells (MCCs) exhibited a significant decrease in cilia length and number, due to defective basal body maturation and apical docking. The effect of miR-34/449 on ciliogenesis was mediated, at least in part, by post-transcriptional repression of Cp110, a centriolar protein suppressing cilia assembly. cp110 knockdown in miR-34/449-deficient MCCs restored ciliogenesis by rescuing basal body maturation and docking. Altogether, our findings elucidate conserved cellular and molecular mechanisms through which miR-34/449 regulate motile ciliogenesis. PMID:24899310

  7. Gaslike model of social motility.

    PubMed

    Parravano, A; Reyes, L M

    2008-08-01

    We propose a model to represent the motility of social elements. The model is completely deterministic, possesses a small number of parameters, and exhibits a series of properties that are reminiscent of the behavior of communities in social-ecological competition; these are (i) similar individuals attract each other; (ii) individuals can form stable groups; (iii) a group of similar individuals breaks into subgroups if it reaches a critical size; (iv) interaction between groups can modify the distribution of the elements as a result of fusion, fission, or pursuit; (v) individuals can change their internal state by interaction with their neighbors. The simplicity of the model and its richness of emergent behaviors, such as, for example, pursuit between groups, make it a useful toy model to explore a diversity of situations by changing the rule by which the internal state of individuals is modified by the interactions with the environment.

  8. The actin gene ACT1 is required for phagocytosis, motility, and cell separation of Tetrahymena thermophila.

    PubMed

    Williams, Norman E; Tsao, Che-Chia; Bowen, Josephine; Hehman, Gery L; Williams, Ruth J; Frankel, Joseph

    2006-03-01

    A previously identified Tetrahymena thermophila actin gene (C. G. Cupples and R. E. Pearlman, Proc. Natl. Acad. Sci. USA 83:5160-5164, 1986), here called ACT1, was disrupted by insertion of a neo3 cassette. Cells in which all expressed copies of this gene were disrupted exhibited intermittent and extremely slow motility and severely curtailed phagocytic uptake. Transformation of these cells with inducible genetic constructs that contained a normal ACT1 gene restored motility. Use of an epitope-tagged construct permitted visualization of Act1p in the isolated axonemes of these rescued cells. In ACT1Delta mutant cells, ultrastructural abnormalities of outer doublet microtubules were present in some of the axonemes. Nonetheless, these cells were still able to assemble cilia after deciliation. The nearly paralyzed ACT1Delta cells completed cleavage furrowing normally, but the presumptive daughter cells often failed to separate from one another and later became reintegrated. Clonal analysis revealed that the cell cycle length of the ACT1Delta cells was approximately double that of wild-type controls. Clones could nonetheless be maintained for up to 15 successive fissions, suggesting that the ACT1 gene is not essential for cell viability or growth. Examination of the cell cortex with monoclonal antibodies revealed that whereas elongation of ciliary rows and formation of oral structures were normal, the ciliary rows of reintegrated daughter cells became laterally displaced and sometimes rejoined indiscriminately across the former division furrow. We conclude that Act1p is required in Tetrahymena thermophila primarily for normal ciliary motility and for phagocytosis and secondarily for the final separation of daughter cells.

  9. Active gel model of amoeboid cell motility

    NASA Astrophysics Data System (ADS)

    Callan-Jones, A. C.; Voituriez, R.

    2013-02-01

    We develop a model of amoeboid cell motility based on active gel theory. Modeling the motile apparatus of a eukaryotic cell as a confined layer of finite length of poroelastic active gel permeated by a solvent, we first show that, due to active stress and gel turnover, an initially static and homogeneous layer can undergo a contractile-type instability to a polarized moving state in which the rear is enriched in gel polymer. This agrees qualitatively with motile cells containing an actomyosin-rich uropod at their rear. We find that the gel layer settles into a steadily moving, inhomogeneous state at long times, sustained by a balance between contractility and filament turnover. In addition, our model predicts an optimal value of the gel-substrate adhesion leading to maximum layer speed, in agreement with cell motility assays. The model may be relevant to motility of cells translocating in complex, confining environments that can be mimicked experimentally by cell migration through microchannels.

  10. Sperm motility under conditions of weightlessness.

    PubMed

    Engelmann, U; Krassnigg, F; Schill, W B

    1992-01-01

    The aim of this study was to determine the differences in motility of frozen and thawed bull spermatozoa under conditions of weightlessness compared with ground conditions. The tests were performed within a series of scientific and technologic experiments under microgravity using sounding rockets in the Technologische Experimente unter Schwerelosigkeit (TEXUS) program launched in Kiruna, North Sweden. Using a computerized sperm motility analyzer, significant differences were found in sperm motility under microgravity compared with sperm under gravitational conditions on earth. Computer analysis showed alterations in straight line and curvilinear velocity, as well as in linearity values. The amount of progressively motile spermatozoa, including all spermatozoa with a velocity > 20 microns/second, increased significantly from 24% +/- 9.5% in the reference test to 49% +/- 7.6% in the microgravity test. In conclusion, there is strong evidence that gravity influences sperm motility.

  11. Odorant-stimulated phosphoinositide signaling in mammalian olfactory receptor neurons

    PubMed Central

    Klasen, K.; Corey, E.A.; Kuck, F.; Wetzel, C.H.; Hatt, H.; Ache, B.W.

    2009-01-01

    Recent evidence has revived interest in the idea that phosphoinositides (PIs) may play a role in signal transduction in mammalian olfactory receptor neurons (ORNs). To provide direct evidence that odorants indeed activate PI signaling in ORNs, we used adenoviral vectors carrying two different fluorescently tagged probes, the pleckstrin homology (PH) domains of phospholipase Cδ1 (PLCδ1) and the general receptor of phosphoinositides (GRP1), to monitor PI activity in the dendritic knobs of ORNs in vivo. Odorants mobilized PI(4,5)P2/IP3 and PI(3,4,5)P3, the substrates and products of PLC and PI3K. We then measured odorant activation of PLC and PI3K in olfactory ciliary-enriched membranes in vitro using a phospholipid overlay assay and ELISAs. Odorants activated both PLC and PI3K in the olfactory cilia within 2 sec of odorant stimulation. Odorant-dependent activation of PLC and PI3K in the olfactory epithelium could be blocked by enzyme-specific inhibitors. Odorants activated PLC and PI3K with partially overlapping specificity. These results provide direct evidence that odorants indeed activate PI signaling in mammalian ORNs in a manner that is consistent with the idea that PI signaling plays a role in olfactory transduction. PMID:19781634

  12. Primary cilia are critical for Sonic hedgehog-mediated dopaminergic neurogenesis in the embryonic midbrain.

    PubMed

    Gazea, Mary; Tasouri, Evangelia; Tolve, Marianna; Bosch, Viktoria; Kabanova, Anna; Gojak, Christian; Kurtulmus, Bahtiyar; Novikov, Orna; Spatz, Joachim; Pereira, Gislene; Hübner, Wolfgang; Brodski, Claude; Tucker, Kerry L; Blaess, Sandra

    2016-01-01

    Midbrain dopaminergic (mDA) neurons modulate various motor and cognitive functions, and their dysfunction or degeneration has been implicated in several psychiatric diseases. Both Sonic Hedgehog (Shh) and Wnt signaling pathways have been shown to be essential for normal development of mDA neurons. Primary cilia are critical for the development of a number of structures in the brain by serving as a hub for essential developmental signaling cascades, but their role in the generation of mDA neurons has not been examined. We analyzed mutant mouse lines deficient in the intraflagellar transport protein IFT88, which is critical for primary cilia function. Conditional inactivation of Ift88 in the midbrain after E9.0 results in progressive loss of primary cilia, a decreased size of the mDA progenitor domain, and a reduction in mDA neurons. We identified Shh signaling as the primary cause of these defects, since conditional inactivation of the Shh signaling pathway after E9.0, through genetic ablation of Gli2 and Gli3 in the midbrain, results in a phenotype basically identical to the one seen in Ift88 conditional mutants. Moreover, the expansion of the mDA progenitor domain observed when Shh signaling is constitutively activated does not occur in absence of Ift88. In contrast, clusters of Shh-responding progenitors are maintained in the ventral midbrain of the hypomorphic Ift88 mouse mutant, cobblestone. Despite the residual Shh signaling, the integrity of the mDA progenitor domain is severely disturbed, and consequently very few mDA neurons are generated in cobblestone mutants. Our results identify for the first time a crucial role of primary cilia in the induction of mDA progenitors, define a narrow time window in which Shh-mediated signaling is dependent upon normal primary cilia function for this purpose, and suggest that later Wnt signaling-dependent events act independently of primary cilia. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Redox regulation of mammalian sperm capacitation

    PubMed Central

    O’Flaherty, Cristian

    2015-01-01

    Capacitation is a series of morphological and metabolic changes necessary for the spermatozoon to achieve fertilizing ability. One of the earlier happenings during mammalian sperm capacitation is the production of reactive oxygen species (ROS) that will trigger and regulate a series of events including protein phosphorylation, in a time-dependent fashion. The identity of the sperm oxidase responsible for the production of ROS involved in capacitation is still elusive, and several candidates are discussed in this review. Interestingly, ROS-induced ROS formation has been described during human sperm capacitation. Redox signaling during capacitation is associated with changes in thiol groups of proteins located on the plasma membrane and subcellular compartments of the spermatozoon. Both, oxidation of thiols forming disulfide bridges and the increase on thiol content are necessary to regulate different sperm proteins associated with capacitation. Reducing equivalents such as NADH and NADPH are necessary to support capacitation in many species including humans. Lactate dehydrogenase, glucose-6-phospohate dehydrogenase, and isocitrate dehydrogenase are responsible in supplying NAD (P) H for sperm capacitation. Peroxiredoxins (PRDXs) are newly described enzymes with antioxidant properties that can protect mammalian spermatozoa; however, they are also candidates for assuring the regulation of redox signaling required for sperm capacitation. The dysregulation of PRDXs and of enzymes needed for their reactivation such as thioredoxin/thioredoxin reductase system and glutathione-S-transferases impairs sperm motility, capacitation, and promotes DNA damage in spermatozoa leading to male infertility. PMID:25926608

  14. Primary cilia on porcine testicular somatic cells and their role in hedgehog signaling and tubular morphogenesis in vitro.

    PubMed

    Dores, Camila; Alpaugh, Whitney; Su, Lin; Biernaskie, Jeff; Dobrinski, Ina

    2017-04-01

    The primary cilium is a microtubule-based sensory organelle found on nearly all eukaryotic cells but little is understood about its function in the testis. We investigate the role of primary cilia on testis cells in vitro by inhibiting formation of the primary cilium with Ciliobrevin D, a cell-permeable, reversible chemical inhibitor of ATPase motor cytoplasmic dynein. We analyzed cultured cells for the presence of primary cilia and their involvement in hedgehog signaling. Primary cilia were present on 89.3 ± 2.3 % of untreated testicular somatic cells compared to 3.1 ± 2.5 % cells with primary cilia for Ciliobrevin D-treated cells. Protein levels of Gli-2 and Smoothened were lower on Western blots after suppression of cilia with Ciliobrevin D. The inhibitor did not affect centrosome localization or cell proliferation, indicating that changes were due to ablation of the primary cilium. Testicular somatic cells have the ability to form three-dimensional tubules in vitro. In vitro-formed tubules were significantly longer and wider in the control group than in the Ciliobrevin D-treated group (9.91 ± 0.35 vs. 5.540 ± 1.08 mm and 339.8 ± 55.78 vs. 127.2 ± 11.9 μm, respectively) indicating that primary cilia play a role in tubule formation. Our results establish that the inhibition of ATPase motor cytoplasmic dynein perturbs formation of primary cilia in testicular somatic cells, affects the hedgehog signaling pathway and impairs tubule formation in vitro. These findings provide evidence for a role of cilia in the testis in cell signaling and tubular morphogenesis in vitro.

  15. Mammalian development in space

    NASA Technical Reports Server (NTRS)

    Ronca, April E.

    2003-01-01

    Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young

  16. Mammalian development in space

    NASA Technical Reports Server (NTRS)

    Ronca, April E.

    2003-01-01

    Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young

  17. Mammalian development in space.

    PubMed

    Ronca, April E

    2003-01-01

    Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young

  18. Effects of pharmacological agents on gastrointestinal motility.

    PubMed

    Gerring, E L

    1989-08-01

    The control mechanisms of gastrointestinal motility are complex. Extrinsic neurohormonal effects modulate an intrinsic system, often called the "gut brain," composed of nervous and neuropeptide components. To exert pharmacologic influence on GI motility, use is made of agents that mimic the external control system. Agents that stimulate opioid receptors, block adrenoceptors, block or facilitate acetylcholine action, or antagonize the action of prostaglandins are used to effect changes in GI motility. The major indications for pharmacologic intervention are to increase motility in constipation, to reduce it in most cases of diarrhea, and to restore propulsive coordination in postoperative ileus. In cases of clinical colic the primary requirement is control of pain. Agents used for this purpose may adversely affect motility, and choice requires knowledge of their actions in this respect. In addition, drugs used for other purposes, anthelmintics for instance, may also influence gut motility. A synopsis of the actions of the agents commonly employed in GI motility control and some associated drugs are displayed in Table 3. Recent advances in the understanding of drug action on the gut should help in the selection of drugs for clinical use.

  19. Regulation of flagellar motility during biofilm formation

    PubMed Central

    Guttenplan, Sarah B.; Kearns, Daniel B.

    2013-01-01

    Many bacteria swim in liquid or swarm over solid surfaces by synthesizing rotary flagella. The same bacteria that are motile also commonly form non-motile multicellular aggregates held together by an extracellular matrix called biofilms. Biofilms are an important part of the lifestyle of pathogenic bacteria and it is assumed that there is a motility-to-biofilm transition wherein the inhibition of motility promotes biofilm formation. The transition is largely inferred from regulatory mutants that reveal the opposite regulation of the two phenotypes. Here we review the regulation of motility during biofilm formation in Bacillus, Pseudomonas, Vibrio, and Escherichia, and we conclude that the motility-to-biofilm transition, if necessary, likely involves two steps. In the short term, flagella are functionally regulated to either inhibit rotation or modulate the basal flagellar reversal frequency. Over the long term, flagellar gene transcription is inhibited and in the absence of de novo synthesis, flagella are likely diluted to extinction through growth. Both short term and long term control is likely important to the motility-to-biofilm transition to stabilize aggregates and optimize resource investment. We emphasize the newly discovered classes of flagellar functional regulators and speculate that others await discovery in the context of biofilm formation. PMID:23480406

  20. An ovine hepatorenal fibrocystic model of a Meckel-like syndrome associated with dysmorphic primary cilia and TMEM67 mutations.

    PubMed

    Stayner, C; Poole, C A; McGlashan, S R; Pilanthananond, M; Brauning, R; Markie, D; Lett, B; Slobbe, L; Chae, A; Johnstone, A C; Jensen, C G; McEwan, J C; Dittmer, K; Parker, K; Wiles, A; Blackburne, W; Leichter, A; Leask, M; Pinnapureddy, A; Jennings, M; Horsfield, J A; Walker, R J; Eccles, M R

    2017-05-09

    Meckel syndrome (MKS) is an inherited autosomal recessive hepatorenal fibrocystic syndrome, caused by mutations in TMEM67, characterized by occipital encephalocoele, renal cysts, hepatic fibrosis, and polydactyly. Here we describe an ovine model of MKS, with kidney and liver abnormalities, without polydactyly or occipital encephalocoele. Homozygous missense p.(Ile681Asn; Ile687Ser) mutations identified in ovine TMEM67 were pathogenic in zebrafish phenotype rescue assays. Meckelin protein was expressed in affected and unaffected kidney epithelial cells by immunoblotting, and in primary cilia of lamb kidney cyst epithelial cells by immunofluorescence. In contrast to primary cilia of relatively consistent length and morphology in unaffected kidney cells, those of affected cyst-lining cells displayed a range of short and extremely long cilia, as well as abnormal morphologies, such as bulbous regions along the axoneme. Putative cilia fragments were also consistently located within the cyst luminal contents. The abnormal ciliary phenotype was further confirmed in cultured interstitial fibroblasts from affected kidneys. These primary cilia dysmorphologies and length control defects were significantly greater in affected cells compared to unaffected controls. In conclusion, we describe abnormalities involving primary cilia length and morphology in the first reported example of a large animal model of MKS, in which we have identified TMEM67 mutations.

  1. Arl13b in primary cilia regulates the migration and placement of interneurons in the developing cerebral cortex.

    PubMed

    Higginbotham, Holden; Eom, Tae-Yeon; Mariani, Laura E; Bachleda, Amelia; Hirt, Joshua; Gukassyan, Vladimir; Cusack, Corey L; Lai, Cary; Caspary, Tamara; Anton, E S

    2012-11-13

    Coordinated migration and placement of interneurons and projection neurons lead to functional connectivity in the cerebral cortex; defective neuronal migration and the resultant connectivity changes underlie the cognitive defects in a spectrum of neurological disorders. Here we show that primary cilia play a guiding role in the migration and placement of postmitotic interneurons in the developing cerebral cortex and that this process requires the ciliary protein, Arl13b. Through live imaging of interneuronal cilia, we show that migrating interneurons display highly dynamic primary cilia and we correlate cilia dynamics with the interneuron's migratory state. We demonstrate that the guidance cue receptors essential for interneuronal migration localize to interneuronal primary cilia, but their concentration and dynamics are altered in the absence of Arl13b. Expression of Arl13b variants known to cause Joubert syndrome induce defective interneuronal migration, suggesting that defects in cilia-dependent interneuron migration may in part underlie the neurological defects in Joubert syndrome patients. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. The mammalian Cos2 homolog Kif7 plays an essential role in modulating Hh signal transduction during development.

    PubMed

    Endoh-Yamagami, Setsu; Evangelista, Marie; Wilson, Deanna; Wen, Xiaohui; Theunissen, Jan-Willem; Phamluong, Khanhky; Davis, Matti; Scales, Suzie J; Solloway, Mark J; de Sauvage, Frederic J; Peterson, Andrew S

    2009-08-11

    The Hedgehog (Hh) signaling pathway regulates development in animals ranging from flies to humans. Although its framework is conserved, differences in pathway components have been reported. A kinesin-like protein, Costal2 (Cos2), plays a central role in the Hh pathway in flies. Knockdown of a zebrafish homolog of Cos2, Kif7, results in ectopic Hh signaling, suggesting that Kif7 acts primarily as a negative regulator of Hh signal transduction. However, in vitro analysis of the function of mammalian Kif7 and the closely related Kif27 has led to the conclusion that neither protein has a role in Hh signaling. Using Kif7 knockout mice, we demonstrate that mouse Kif7, like its zebrafish and Drosophila homologs, plays a role in transducing the Hh signal. We show that Kif7 accumulates at the distal tip of the primary cilia in a Hh-dependent manner. We also demonstrate a requirement for Kif7 in the efficient localization of Gli3 to cilia in response to Hh and for the processing of Gli3 to its repressor form. These results suggest a role for Kif7 in coordinating Hh signal transduction at the tip of cilia and preventing Gli3 cleavage into a repressor form in the presence of Hh.

  3. Katanin p80 Regulates Human Cortical Development by Limiting Centriole and Cilia Number

    PubMed Central

    Hu, Wen F.; Pomp, Oz; Ben-Omran, Tawfeg; Kodani, Andrew; Henke, Katrin; Mochida, Ganeshwaran H.; Yu, Timothy W.; Woodworth, Mollie B.; Bonnard, Carine; Raj, Grace Selva; Tan, Thong Teck; Hamamy, Hanan; Masri, Amira; Shboul, Mohammad; Al Saffar, Muna; Partlow, Jennifer N.; Al-Dosari, Mohammed; Alazami, Anas; Alowain, Mohammed; Alkuraya, Fowzan S.; Reiter, Jeremy F.; Harris, Matthew P.; Reversade, Bruno; Walsh, Christopher A.

    2015-01-01

    SUMMARY Katanin is a microtubule-severing complex whose catalytic activities are well characterized, but whose in vivo functions are incompletely understood. Human mutations in KATNB1, which encodes the noncatalytic regulatory p80 subunit of katanin, cause severe microlissencephaly. Loss of Katnb1 in mice confirms essential roles in neurogenesis and cell survival, while loss of zebrafish katnb1 reveals specific roles for katnin p80 in early and late developmental stages. Surprisingly, Katnb1 null mutant mouse embryos display hallmarks of aberrant Sonic hedgehog signaling, including holoprosencephaly. KATNB1-deficient human cells show defective proliferation and spindle structure, while Katnb1 null fibroblasts also demonstrate a remarkable excess of centrioles, with supernumerary cilia but deficient Hedgehog signaling. Our results reveal unexpected functions for KATNB1 in regulating overall centriole, mother centriole, and cilia number, and as an essential gene for normal Hedgehog signaling during neocortical development. PMID:25521379

  4. A molecular ruler determines the repeat length in eukaryotic cilia and flagella.

    PubMed

    Oda, Toshiyuki; Yanagisawa, Haruaki; Kamiya, Ritsu; Kikkawa, Masahide

    2014-11-14

    Existence of cellular structures with specific size raises a fundamental question in biology: How do cells measure length? One conceptual answer to this question is by a molecular ruler, but examples of such rulers in eukaryotes are lacking. In this work, we identified a molecular ruler in eukaryotic cilia and flagella. Using cryo-electron tomography, we found that FAP59 and FAP172 form a 96-nanometer (nm)-long complex in Chlamydomonas flagella and that the absence of the complex disrupted 96-nm repeats of axonemes. Furthermore, lengthening of the FAP59/172 complex by domain duplication resulted in extension of the repeats up to 128 nm, as well as duplication of specific axonemal components. Thus, the FAP59/172 complex is the molecular ruler that determines the 96-nm repeat length and arrangements of components in cilia and flagella.

  5. A Smoothened-Evc2 complex transduces the Hedgehog signal at primary cilia.

    PubMed

    Dorn, Karolin V; Hughes, Casey E; Rohatgi, Rajat

    2012-10-16

    Vertebrate Hedgehog (Hh) signaling is initiated at primary cilia by the ligand-triggered accumulation of Smoothened (Smo) in the ciliary membrane. The underlying biochemical mechanisms remain unknown. We find that Hh agonists promote the association between Smo and Evc2, a ciliary protein that is defective in two human ciliopathies. The formation of the Smo-Evc2 complex is under strict spatial control, being restricted to a distinct ciliary compartment, the EvC zone. Mutant Evc2 proteins that localize in cilia but are displaced from the EvC zone are dominant inhibitors of Hh signaling. Disabling Evc2 function blocks Hh signaling at a specific step between Smo and the downstream regulators protein kinase A and Suppressor of Fused, preventing activation of the Gli transcription factors. Our data suggest that the Smo-Evc2 signaling complex at the EvC zone is required for Hh signal transmission and elucidate the molecular basis of two human ciliopathies.

  6. Imaging fluid flow and cilia beating pattern in Xenopus brain ventricles.

    PubMed

    Miskevich, Frank

    2010-05-30

    Brain development and health depends upon the efficient movement of the cerebrospinal fluid inside of brain ventricles. When disrupted either through mutation, disease, or physiological damage, brain function becomes significantly impaired. Here I present a simple method of following cerebrospinal fluid circulation in Xenopus tadpoles using fluorescent microspheres which can be applied to imaging fluid circulation in any transparent embryo. In particular, cilia may be labeled with these microspheres to study their dynamics and movement patterns in vivo while simultaneously measuring bulk fluid flow. This technique will facilitate the analysis of fluid dynamics in developing embryos and aid in understanding the regulation of cilia dependent fluid flow in vivo. (c) 2010 Elsevier B.V. All rights reserved.

  7. Identification of Elongated Primary Cilia with Impaired Mechanotransduction in Idiopathic Scoliosis Patients

    PubMed Central

    Oliazadeh, Niaz; Gorman, Kristen F.; Eveleigh, Robert; Bourque, Guillaume; Moreau, Alain

    2017-01-01

    The primary cilium is an outward projecting antenna-like organelle with an important role in bone mechanotransduction. The capacity to sense mechanical stimuli can affect important cellular and molecular aspects of bone tissue. Idiopathic scoliosis (IS) is a complex pediatric disease of unknown cause, defined by abnormal spinal curvatures. We demonstrate significant elongation of primary cilia in IS patient bone cells. In response to mechanical stimulation, these IS cells differentially express osteogenic factors, mechanosensitive genes, and signaling genes. Considering that numerous ciliary genes are associated with a scoliosis phenotype, among ciliopathies and knockout animal models, we expected IS patients to have an accumulation of rare variants in ciliary genes. Instead, our SKAT-O analysis of whole exomes showed an enrichment among IS patients for rare variants in genes with a role in cellular mechanotransduction. Our data indicates defective cilia in IS bone cells, which may be linked to heterogeneous gene variants pertaining to cellular mechanotransduction. PMID:28290481

  8. Effects of freezing/thawing on motile sperm subpopulations of boar and donkey ejaculates.

    PubMed

    Flores, E; Taberner, E; Rivera, M M; Peña, A; Rigau, T; Miró, J; Rodríguez-Gil, J E

    2008-10-01

    The main aim of this study is to assess the influence of freeze/thawing on motile sperm subpopulations in ejaculates from two phylogenetically different mammalian species, boar and donkey. Our results indicate that, whereas boar and donkey sperm respond very differently in their mean motion characteristics to freezing/thawing, this process did not change the existence of a 4-subpopulations structure in the ejaculates in either species when these subpopulations were defined by taking values of curvilinear velocity (VCL) as reference. Moreover, the freezing/thawing-linked changes in mean sperm-motion characteristics in both boar and donkey semen were especially due to changes in the proportion among each concrete subpopulation. In this way, the freezing/thawing-induced mean increase in motion characteristics observed in boar sperm was a result of the decrease in the percentage of sperm in Subpopulation 1 (from 53.9%+/-4.7% to 31.2%+/-3.9% after thawing) and a concomitant increase of sperm from Subpopulations 3 (from 13.3%+/-2.5% to 32.6%+/-3.9% after thawing) and 4 (from 3.4%+/-0.9% to 8.0%+/-1.1% after thawing). On the contrary, changes in mean moti