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Sample records for motile mammalian cilia

  1. Cyclic GMP and Cilia Motility

    PubMed Central

    Wyatt, Todd A.

    2015-01-01

    Motile cilia of the lungs respond to environmental challenges by increasing their ciliary beat frequency in order to enhance mucociliary clearance as a fundamental tenant of innate defense. One important second messenger in transducing the regulable nature of motile cilia is cyclic guanosine 3′,5′-monophosphate (cGMP). In this review, the history of cGMP action is presented and a survey of the existing data addressing cGMP action in ciliary motility is presented. Nitric oxide (NO)-mediated regulation of cGMP in ciliated cells is presented in the context of alcohol-induced cilia function and dysfunction. PMID:26264028

  2. 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

  3. 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

  4. Motile Cilia of Human Airway Epithelia Are Chemosensory

    PubMed Central

    Shah, Alok S; Ben-Shahar, Yehuda; Moninger, Thomas O; Kline, Joel N; Welsh, Michael J

    2010-01-01

    Cilia are microscopic projections that extend from eukaryotic cells. There are two general types of cilia; primary cilia serve as sensory organelles, whereas motile cilia exert mechanical force. The motile cilia emerging from human airway epithelial cells propel harmful inhaled material out of the lung. We found that these cells express sensory bitter taste receptors, which localized on motile cilia. Bitter compounds increased the intracellular Ca2+ concentration and stimulated ciliary beat frequency. Thus, airway epithelia contain a cell-autonomous system in which motile cilia both sense noxious substances entering airways and initiate a defensive mechanical mechanism to eliminate the offending compound. Hence, like primary cilia, classical motile cilia also contain sensors to detect the external environment. PMID:19628819

  5. 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.

  6. Tubulin glycylases and glutamylases have distinct functions in stabilization and motility of ependymal cilia

    PubMed Central

    Bosch Grau, Montserrat; Gonzalez Curto, Gloria; Rocha, Cecilia; Magiera, Maria M.; Marques Sousa, Patricia; Giordano, Tiziana

    2013-01-01

    Microtubules are subject to a variety of posttranslational modifications that potentially regulate cytoskeletal functions. Two modifications, glutamylation and glycylation, are highly enriched in the axonemes of most eukaryotes, and might therefore play particularly important roles in cilia and flagella. Here we systematically analyze the dynamics of glutamylation and glycylation in developing mouse ependymal cilia and the expression of the corresponding enzymes in the brain. By systematically screening enzymes of the TTLL family for specific functions in ependymal cilia, we demonstrate that the glycylating enzymes TTLL3 and TTLL8 were required for stability and maintenance of ependymal cilia, whereas the polyglutamylase TTLL6 was necessary for coordinated beating behavior. Our work provides evidence for a functional separation of glutamylating and glycylating enzymes in mammalian ependymal cilia. It further advances the elucidation of the functions of tubulin posttranslational modifications in motile cilia of the mammalian brain and their potential importance in brain development and disease. PMID:23897886

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

    PubMed

    Lewis, Wesley R; Malarkey, Erik B; Tritschler, Douglas; Bower, Raqual; Pasek, Raymond C; Porath, Jonathan D; Birket, Susan E; Saunier, Sophie; Antignac, Corinne; Knowles, Michael R; Leigh, Margaret W; Zariwala, Maimoona A; Challa, Anil K; Kesterson, Robert A; Rowe, Steven M; Drummond, Iain A; Parant, John M; Hildebrandt, Friedhelm; Porter, Mary E; Yoder, Bradley K; Berbari, Nicolas F

    2016-07-01

    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 in

  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. Sensory functions of motile cilia and implication for bronchiectasis

    PubMed Central

    Jain, Raksha; Javidan-Nejad, Cylen; Alexander-Brett, Jennifer; Horani, Amjad; Cabellon, Michelle C.; Walter, Michael J.; Brody, Steven L.

    2013-01-01

    Cilia are specialized organelles that extend from the surface of cells into the local environment. Airway epithelial cell cilia are motile to provide mucociliary clearance for host defense. On other cells, solitary cilia are specialized to detect chemical or mechanosensory signals. Sensory proteins in motile cilia have recently been identified that detect shear stress, osmotic force, fluid flow, bitter taste and sex hormones. The relationship of sensory function in human motile cilia to disease is now being revealed. One example is polycystin-1 and polycystin-2. As a complex, these proteins function as a flow sensor in cilia and are mutated in autosomal dominant polycystic kidney disease (ADPKD). The polycystins are also expressed in motile cilia of the airways, potentially operating as sensors in the lung. Computed tomography studies from patients with ADPKD revealed radiographic evidence for bronchiectasis, suggesting that polycystin-1 and -2 are important in lung function. The expression of this complex and sensory channel TRPV4, and bitter taste and sex hormones receptors in motile cilia indicate that the cell is wired to interpret environmental cues to regulate cilia beat frequency and other functions. Defective signaling of sensory proteins may result in a ciliopathy that includes lung disease. PMID:22202111

  10. 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.

  11. ADENYLATE CYCLASE REGULATES ELONGATION OF MAMMALIAN PRIMARY CILIA

    PubMed Central

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

    2011-01-01

    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-3β 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. PMID:19576885

  12. Specialized Cilia in Mammalian Sensory Systems

    PubMed Central

    Falk, Nathalie; Lösl, Marlene; Schröder, Nadja; Gießl, Andreas

    2015-01-01

    Cilia and flagella are highly conserved and important microtubule-based organelles that project from the surface of eukaryotic cells and act as antennae to sense extracellular signals. Moreover, cilia have emerged as key players in numerous physiological, developmental, and sensory processes such as hearing, olfaction, and photoreception. Genetic defects in ciliary proteins responsible for cilia formation, maintenance, or function underlie a wide array of human diseases like deafness, anosmia, and retinal degeneration in sensory systems. Impairment of more than one sensory organ results in numerous syndromic ciliary disorders like the autosomal recessive genetic diseases Bardet-Biedl and Usher syndrome. Here we describe the structure and distinct functional roles of cilia in sensory organs like the inner ear, the olfactory epithelium, and the retina of the mouse. The spectrum of ciliary function in fundamental cellular processes highlights the importance of elucidating ciliopathy-related proteins in order to find novel potential therapies. PMID:26378583

  13. 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. PMID:24067530

  14. c21orf59/kurly Controls Both Cilia Motility and Polarization.

    PubMed

    Jaffe, Kimberly M; Grimes, Daniel T; Schottenfeld-Roames, Jodi; Werner, Michael E; Ku, Tse-Shuen J; Kim, Sun K; Pelliccia, Jose L; Morante, Nicholas F C; Mitchell, Brian J; Burdine, Rebecca D

    2016-03-01

    Cilia are microtubule-based projections that function in the movement of extracellular fluid. This requires cilia to be: (1) motile and driven by dynein complexes and (2) correctly polarized on the surface of cells, which requires planar cell polarity (PCP). Few factors that regulate both processes have been discovered. We reveal that C21orf59/Kurly (Kur), a cytoplasmic protein with some enrichment at the base of cilia, is needed for motility; zebrafish mutants exhibit characteristic developmental abnormalities and dynein arm defects. kur was also required for proper cilia polarization in the zebrafish kidney and the larval skin of Xenopus laevis. CRISPR/Cas9 coupled with homologous recombination to disrupt the endogenous kur locus in Xenopus resulted in the asymmetric localization of the PCP protein Prickle2 being lost in mutant multiciliated cells. Kur also makes interactions with other PCP components, including Disheveled. This supports a model wherein Kur plays a dual role in cilia motility and polarization. PMID:26904945

  15. 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.

  16. 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

  17. 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

  18. 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

    2014-08-01

    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. PMID:25043421

  19. Robust estimation of the motile cilia beating frequency.

    PubMed

    Meste, O; Brau, F; Guyon, A

    2015-10-01

    The estimation of the cilia beating frequency (CBF) is of great interest in understanding how the CBF modulates liquid fluxes and how it is controlled by the ciliated cell intra- and/or extracellular medium composition in physiological processes. Motion artifacts and camera defaults may hinder the computation of the frequency variations during long-lasting experiments. We have developed a new analysis approach consisting of a preliminary corrective step (removal of a grid pattern on the image sequence and shift compensation), followed by a harmonic model of the observed cilia using a maximum likelihood estimator framework. It is shown that a more accurate estimation of the frequency can be obtained by averaging the squared Fourier transform of individual pixels followed by a particular summation over the different frequencies, namely the compressed spectrum. Robustness of the proposed method over traditional approaches is shown by several examples and simulations. The method is then applied to images of samples containing ciliated ependymal cells located in the third cerebral ventricle of mouse brains, showing that even small variations in CBF in response to changes in the amount of oxygenation, pH or glucose were clearly visible in the computed frequencies. As a conclusion, this method reveals a fine metabolic tuning of the cilia beating in ependimocytes lining the third cerebral ventricle. Such regulations are likely to participate in homeostatic mechanisms regulating CSF movements and brain energy supply. PMID:26215519

  20. Ex vivo method for high resolution imaging of cilia motility in rodent airway epithelia.

    PubMed

    Francis, Richard; Lo, Cecilia

    2013-01-01

    An ex vivo technique for imaging mouse airway epithelia for quantitative analysis of motile cilia function important for insight into mucociliary clearance function has been established. Freshly harvested mouse trachea is cut longitudinally through the trachealis muscle and mounted in a shallow walled chamber on a glass-bottomed dish. The trachea sample is positioned along its long axis to take advantage of the trachealis muscle to curl longitudinally. This allows imaging of ciliary motion in the profile view along the entire tracheal length. Videos at 200 frames/sec are obtained using differential interference contrast microscopy and a high speed digital camera to allow quantitative analysis of cilia beat frequency and ciliary waveform. With the addition of fluorescent beads during imaging, cilia generated fluid flow also can be determined. The protocol time spans approximately 30 min, with 5 min for chamber preparation, 5-10 min for sample mounting, and 10-15 min for videomicroscopy. PMID:23963287

  1. Loss of Bardet–Biedl syndrome proteins alters the morphology and function of motile cilia in airway epithelia

    PubMed Central

    Shah, Alok S.; Farmen, Sara L.; Moninger, Thomas O.; Businga, Thomas R.; Andrews, Michael P.; Bugge, Kevin; Searby, Charles C.; Nishimura, Darryl; Brogden, Kim A.; Kline, Joel N.; Sheffield, Val C.; Welsh, Michael J.

    2008-01-01

    Mutations in a group of genes that contribute to ciliary function cause Bardet–Biedl syndrome (BBS). Most studies of BBS have focused on primary, sensory cilia. Here, we asked whether loss of BBS proteins would also affect motile cilia lining the respiratory tract. We found that BBS genes were expressed in human airway epithelia, and BBS2 and BBS4 localized to cellular structures associated with motile cilia. Although BBS proteins were not required for ciliogenesis, their loss caused structural defects in a fraction of cilia covering mouse airway epithelia. The most common abnormality was bulges filled with vesicles near the tips of cilia. We discovered this same misshapen appearance in airway cilia from Bbs1, Bbs2, Bbs4, and Bbs6 mutant mice. The structural abnormalities were accompanied by functional defects; ciliary beat frequency was reduced in Bbs mutant mice. Previous reports suggested BBS might increase the incidence of asthma. However, compared with wild-type controls, neither airway hyperresponsiveness nor inflammation increased in Bbs2−/− or Bbs4−/− mice immunized with ovalbumin. Instead, these animals were partially protected from airway hyperresponsiveness. These results emphasize the role of BBS proteins in both the structure and function of motile cilia. They also invite additional scrutiny of motile cilia dysfunction in patients with this disease. PMID:18299575

  2. The role of hair cells, cilia and ciliary motility in otolith formation in the zebrafish otic vesicle.

    PubMed

    Stooke-Vaughan, Georgina A; Huang, Peng; Hammond, Katherine L; Schier, Alexander F; Whitfield, Tanya T

    2012-05-01

    Otoliths are biomineralised structures required for the sensation of gravity, linear acceleration and sound in the zebrafish ear. Otolith precursor particles, initially distributed throughout the otic vesicle lumen, become tethered to the tips of hair cell kinocilia (tether cilia) at the otic vesicle poles, forming two otoliths. We have used high-speed video microscopy to investigate the role of cilia and ciliary motility in otolith formation. In wild-type ears, groups of motile cilia are present at the otic vesicle poles, surrounding the immotile tether cilia. A few motile cilia are also found on the medial wall, but most cilia (92-98%) in the otic vesicle are immotile. In mutants with defective cilia (iguana) or ciliary motility (lrrc50), otoliths are frequently ectopic, untethered or fused. Nevertheless, neither cilia nor ciliary motility are absolutely required for otolith tethering: a mutant that lacks cilia completely (MZovl) is still capable of tethering otoliths at the otic vesicle poles. In embryos with attenuated Notch signalling [mindbomb mutant or Su(H) morphant], supernumerary hair cells develop and otolith precursor particles bind to the tips of all kinocilia, or bind directly to the hair cells' apical surface if cilia are absent [MZovl injected with a Su(H)1+2 morpholino]. However, if the first hair cells are missing (atoh1b morphant), otolith formation is severely disrupted and delayed. Our data support a model in which hair cells produce an otolith precursor-binding factor, normally localised to tether cell kinocilia. We also show that embryonic movement plays a minor role in the formation of normal otoliths. PMID:22461562

  3. 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

  4. Tubulin tyrosine ligase-like genes ttll3 and ttll6 maintain zebrafish cilia structure and motility.

    PubMed

    Pathak, Narendra; Austin, Christina A; Drummond, Iain A

    2011-04-01

    Tubulin post-translational modifications generate microtubule heterogeneity and modulate microtubule function, and are catalyzed by tubulin tyrosine ligase-like (TTLL) proteins. Using antibodies specific to monoglycylated, polyglycylated, and glutamylated tubulin in whole mount immunostaining of zebrafish embryos, we observed distinct, tissue-specific patterns of tubulin modifications. Tubulin modification patterns in cilia correlated with the expression of ttll3 and ttll6 in ciliated cells. Expression screening of all zebrafish tubulin tyrosine ligase-like genes revealed additional tissue-specific expression of ttll1 in brain neurons, ttll4 in muscle, and ttll7 in otic placodes. Knockdown of ttll3 eliminated cilia tubulin glycylation but had surprisingly mild effects on cilia structure and motility. Similarly, knockdown of ttll6 strongly reduced cilia tubulin glutamylation but only partially affected cilia structure and motility. Combined loss of function of ttll3 and ttll6 caused near complete loss of cilia motility and induced a variety of axonemal ultrastructural defects similar to defects previously observed in zebrafish fleer mutants, which were shown to lack tubulin glutamylation. Consistently, we find that fleer mutants also lack tubulin glycylation. These results indicate that tubulin glycylation and glutamylation have overlapping functions in maintaining cilia structure and motility and that the fleer/dyf-1 TPR protein is required for both types of tubulin post-translational modification. PMID:21262966

  5. Caenorhabditis elegans DYF-11, an orthologue of mammalian Traf3ip1/MIP-T3, is required for sensory cilia formation.

    PubMed

    Kunitomo, Hirofumi; Iino, Yuichi

    2008-01-01

    Cilia and flagella play critical roles in cell motility, development and sensory perception in animals. Formation and maintenance of cilia require a conserved protein transport system called intraflagellar transport (IFT). Here, we show that Caenorhabditis elegans dyf-11 encodes an evolutionarily conserved protein required for cilium biogenesis. dyf-11 is expressed in most of the ciliated neurons and is regulated by DAF-19, a crucial transcription factor for ciliary genes in C. elegans. dyf-11 mutants exhibit stunted cilia, fluorescent dye-filling defects (Dyf) of sensory neurons, and abnormal chemotaxis (Che). Cell- and stage-specific rescue experiments indicated that DYF-11 is required for formation and maintenance of sensory cilia in cell-autonomous manner. Fluorescent protein-tagged DYF-11 localizes to cilia and moves antero- and retrogradely via IFT. Analysis of DYF-11 movement in bbs mutants further suggested that DYF-11 is likely associated with IFT complex B. Domain analysis using DYF-11 deletion constructs revealed that the coiled-coil region is required for proper localization and ciliogenesis. We further show that Traf3ip1/MIP-T3, the mammalian orthologue of DYF-11, localizes to cilia in the MDCK renal epithelial cells. PMID:18173744

  6. RC/BTB2 is Essential for Formation of Primary Cilia in Mammalian Cells

    PubMed Central

    Zhang, Ling; Li, Wei; Ni, Jin; Wu, Jinghua; Liu, Junping; Zhang, Zhengang; Zhang, Yong; Li, Hongfei; Shi, Yuqin; Teves, Maria E; Song, Shizheng; Strauss, Jerome F.; Zhang, Zhibing

    2016-01-01

    RC/BTB2 is a binding partner of sperm associated antigen 16S (SPAG16S), which is regulator of spermiogenesis in mice, a process during which sperm flagella are formed. The expression of Rc/btb2 is also regulated by multicilin, a protein that controls ciliogenesis. Given that mouse Rc/btb2 mRNA is not only expressed in tissues bearing motile cilia, but also in tissues without motile cilia, we investigated whether RC/BTB2 plays a role in the general process of ciliogenesis by studying two somatic cells lines that have primary cilia, NIH3T3 and IMCD3. We discovered that the subcellular localization of RT/BTB2 in the NIH3T3 and IMCD3 cells encompasses the pathway for ciliogenesis. RC/BTB2 was found in the Golgi bodies and centrosomes, two key structures essential for normal ciliogenesis. Knockdown of Rc/btb2 gene expression in these cell lines disrupted ciliogenesis. The percentage of cells with primary cilia was significantly reduced in stable cell lines transduced with specific Rc/btb2 shRNA viruses compared to the control cells. When cilia were formed in the knockdown cells, they were significantly shorter than those in the control cells. Knockdown of Rc/btb2 expression did not affect cell proliferation and the cell cycle. Exogenous expression of RC/BTB2 in these stable knockdown cells restored ciliogenesis. These findings suggest that RC/BTB2 is a necessary component of the process of formation of primary cilia in somatic cells, perhaps through the transportation of cargos from Golgi bodies to centrosomes for cilia assembling. PMID:25762510

  7. 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.

  8. A complex of BBS1 and NPHP7 is required for cilia motility in zebrafish.

    PubMed

    Kim, Yun Hee; Epting, Daniel; Slanchev, Krasimir; Engel, Christina; Walz, Gerd; Kramer-Zucker, Albrecht

    2013-01-01

    Bardet-Biedl syndrome (BBS) and nephronophthisis (NPH) are hereditary autosomal recessive disorders, encoded by two families of diverse genes. BBS and NPH display several overlapping phenotypes including cystic kidney disease, retinitis pigmentosa, liver fibrosis, situs inversus and cerebellar defects. Since most of the BBS and NPH proteins localize to cilia and/or their appendages, BBS and NPH are considered ciliopathies. In this study, we characterized the function of the transcription factor Nphp7 in zebrafish, and addressed the molecular connection between BBS and NPH. The knockdown of zebrafish bbs1 and nphp7.2 caused similar phenotypic changes including convergent extension defects, curvature of the body axis, hydrocephalus, abnormal heart looping and cystic pronephros, all consistent with an altered ciliary function. Immunoprecipitation assays revealed a physical interaction between BBS1 and NPHP7, and the simultaneous knockdown of zbbs1 and znphp7.2 enhanced the cystic pronephros phenotype synergistically, suggesting a genetic interaction between zbbs1 and znphp7.2 in vivo. Deletion of zBbs1 or zNphp7.2 did not compromise cilia formation, but disrupted cilia motility. Although NPHP7 has been shown to act as transcriptional repressor, our studies suggest a crosstalk between BBS1 and NPHP7 in regulating normal function of the cilium. PMID:24069149

  9. 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

  10. Direct evidence for BBSome-associated intraflagellar transport reveals distinct properties of native mammalian cilia

    PubMed Central

    Williams, Corey L.; McIntyre, Jeremy C.; Norris, Stephen R.; Jenkins, Paul M.; Zhang, Lian; Pei, Qinglin; Verhey, Kristen; Martens, Jeffrey R.

    2014-01-01

    Cilia dysfunction underlies a class of human diseases with variable penetrance in different organ systems. Across eukaryotes, intraflagellar transport (IFT) facilitates cilia biogenesis and cargo trafficking, but our understanding of mammalian IFT is insufficient. Here we perform live analysis of cilia ultrastructure, composition and cargo transport in native mammalian tissue using olfactory sensory neurons. Proximal and distal axonemes of these neurons show no bias towards IFT kinesin-2 choice, and Kif17 homodimer is dispensable for distal segment IFT. We identify Bardet–Biedl syndrome proteins (BBSome) as bona fide constituents of IFT in olfactory sensory neurons, and show that they exist in 1:1 stoichiometry with IFT particles. Conversely, subpopulations of peripheral membrane proteins, as well as transmembrane olfactory signalling pathway components, are capable of IFT but with significantly less frequency and/or duration. Our results yield a model for IFT and cargo trafficking in native mammalian cilia and may explain the penetrance of specific ciliopathy phenotypes in olfactory neurons. PMID:25504142

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

    PubMed Central

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

    2015-01-01

    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. PMID:26446842

  12. RFX3 governs growth and beating efficiency of motile cilia in mouse and controls the expression of genes involved in human ciliopathies.

    PubMed

    El Zein, Loubna; Ait-Lounis, Aouatef; Morlé, Laurette; Thomas, Joëlle; Chhin, Brigitte; Spassky, Nathalie; Reith, Walter; Durand, Bénédicte

    2009-09-01

    Cilia are cellular organelles that play essential physiological and developmental functions in various organisms. They can be classified into two categories, primary cilia and motile cilia, on the basis of their axonemal architecture. Regulatory factor X (RFX) transcription factors have been shown to be involved in the assembly of primary cilia in Caenorhabditis elegans, Drosophila and mice. Here, we have taken advantage of a novel primary-cell culture system derived from mouse brain to show that RFX3 is also necessary for biogenesis of motile cilia. We found that the growth and beating efficiencies of motile cilia are impaired in multiciliated Rfx3(-/-) cells. RFX3 was required for optimal expression of the FOXJ1 transcription factor, a key player in the differentiation program of motile cilia. Furthermore, we demonstrate for the first time that RFX3 regulates the expression of axonemal dyneins involved in ciliary motility by binding directly to the promoters of their genes. In conclusion, RFX proteins not only regulate genes involved in ciliary assembly, but also genes that are involved in ciliary motility and that are associated with ciliopathies such as primary ciliary dyskinesia in humans. PMID:19671664

  13. Primary cilia utilize glycoprotein-dependent adhesion mechanisms to stabilize long-lasting cilia-cilia contacts

    PubMed Central

    2012-01-01

    Background The central tenet of cilia function is sensing and transmitting information. The capacity to directly contact extracellular surfaces would empower primary cilia to probe the environment for information about the nature and location of nearby surfaces. It has been well established that flagella and other motile cilia perform diverse cellular functions through adhesion. We hypothesized that mammalian primary cilia also interact with the extracellular environment through direct physical contact. Methods We identified cilia in rod photoreceptors and cholangiocytes in fixed mouse tissues and examined the structures that these cilia contact in vivo. We then utilized an MDCK cell culture model to characterize the nature of the contacts we observed. Results In retina and liver tissue, we observed that cilia from nearby cells touch one another. Using MDCK cells, we found compelling evidence that these contacts are stable adhesions that form bridges between two cells, or networks between many cells. We examined the nature and duration of the cilia-cilia contacts and discovered primary cilia movements that facilitate cilia-cilia encounters. Stable adhesions form as the area of contact expands from a single point to a stretch of tightly bound, adjacent cilia membranes. The cilia-cilia contacts persisted for hours and were resistant to several harsh treatments such as proteases and DTT. Unlike many other cell adhesion mechanisms, calcium was not required for the formation or maintenance of cilia adhesion. However, swainsonine, which blocks maturation of N-linked glycoproteins, reduced contact formation. We propose that cellular control of adhesion maintenance is active because cilia adhesion did not prevent cell division; rather, contacts dissolved during mitosis as cilia were resorbed. Conclusions The demonstration that mammalian primary cilia formed prolonged, direct, physical contacts supports a novel paradigm: that mammalian primary cilia detect features of the

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

    PubMed

    Patel-King, Ramila S; King, Stephen M

    2016-04-15

    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 planarianSchmidtea mediterraneaand 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

  15. 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

  16. 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

  17. Primary cilia and kidney injury: current research status and future perspectives

    PubMed Central

    Wang, Shixuan

    2013-01-01

    Cilia, membrane-enclosed organelles protruding from the apical side of cells, can be divided into two classes: motile and primary cilia. During the past decades, motile cilia have been intensively studied. However, it was not until the 1990s that people began to realize the importance of primary cilia as cellular-specific sensors, particularly in kidney tubular epithelial cells. Furthermore, accumulating evidence indicates that primary cilia may be involved in the regulation of cell proliferation, differentiation, apoptosis, and planar cell polarity. Many signaling pathways, such as Wnt, Notch, Hedgehog, and mammalian target of rapamycin, have been located to the primary cilia. Thus primary cilia have been regarded as a hub that integrates signals from the extracellular environment. More importantly, dysfunction of this organelle may contribute to the pathogenesis of a large spectrum of human genetic diseases, named ciliopathies. The significance of primary cilia in acquired human diseases such as hypertension and diabetes has gradually drawn attention. Interestingly, recent reports disclosed that cilia length varies during kidney injury, and shortening of cilia enhances the sensitivity of epithelial cells to injury cues. This review briefly summarizes the current status of cilia research and explores the potential mechanisms of cilia-length changes during kidney injury as well as provides some thoughts to allure more insightful ideas and promotes the further study of primary cilia in the context of kidney injury. PMID:23904226

  18. 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. PMID:5678451

  19. 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

  20. Cilia dysfunction in lung disease.

    PubMed

    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 composed 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, to repeated chest infections, and to the 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

  1. 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.

  2. Evaluating efficiency and robustness in cilia design.

    PubMed

    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. PMID:27078459

  3. 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

  4. 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. PMID:27193298

  5. 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

  6. 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

  7. Left-right asymmetry: cilia and calcium revisited.

    PubMed

    Blum, Martin; Vick, Philipp

    2015-03-01

    Leftward flow generated by motile cilia is known to underlie left-right asymmetry in vertebrate embryos. A new study now links intraciliary calcium oscillations to cilia motility and the downstream nodal signaling cascade that drives left-sided development. PMID:25734272

  8. Olfactory Cilia: Linking Sensory Cilia Function and Human Disease

    PubMed Central

    Jenkins, Paul M.; McEwen, Dyke P.

    2009-01-01

    The olfactory system gives us an awareness of our immediate environment by allowing us to detect airborne stimuli. The components necessary for detection of these odorants are compartmentalized in the cilia of olfactory sensory neurons. Cilia are microtubule-based organelles, which can be found projecting from the surface of almost any mammalian cell, and are critical for proper olfactory function. Mislocalization of ciliary proteins and/or the loss of cilia cause impaired olfactory function, which is now recognized as a clinical manifestation of a broad class of human diseases, termed ciliopathies. Future work investigating the mechanisms of olfactory cilia function will provide us important new information regarding the pathogenesis of human sensory perception diseases. PMID:19406873

  9. Altering the motility of Trypanosoma cruzi with rabbit polyclonal anti-peptide antibodies reduces infection to susceptible mammalian cells.

    PubMed

    Finkelsztein, Eli J; Diaz-Soto, Juan C; Vargas-Zambrano, Juan C; Suesca, Elizabeth; Guzmán, Fanny; López, Manuel C; Thomas, M Carmen; Forero-Shelton, Manu; Cuellar, Adriana; Puerta, Concepción J; González, John M

    2015-03-01

    Trypanosoma cruzi's trypomastigotes are highly active and their incessant motility seems to be important for mammalian host cell infection. The kinetoplastid membrane protein-11 (KMP-11) is a protein expressed in all parasite stages, which induces a cellular and humoral immune response in the infected host, and is hypothesized to participate in the parasite's motility. An N-terminal peptide from KMP-11, termed K1 or TcTLE, induced polyclonal antibodies that inhibit parasitic invasion of Vero cells. The goal of this study was to evaluate the motility and infectivity of T. cruzi when exposed to polyclonal anti-TcTLE antibodies. Rabbits were immunized with TcTLE peptide along with FIS peptide as an immunomodulator. ELISA assay results showed that post-immunization sera contained high titers of polyclonal anti-TcTLE antibodies, which were also reactive against the native KMP-11 protein and live parasites as detected by immunofluorescence and flow cytometry assays. Trypomastigotes of T. cruzi were incubated with pre- or post-immunization sera, and infectivity to human astrocytes was assessed by Giemsa staining/light microscope and flow cytometry using carboxyfluorescein diacetate succinimidyl ester (CFSE) labeled parasites. T. cruzi infection in astrocytes decreased approximately by 30% upon incubation with post-immunization sera compared with pre-immunization sera. Furthermore, trypomastigotes were recorded by video microscopy and the parasite's flagellar speed was calculated by tracking the flagella. Trypomastigotes exposed to post-immunization sera had qualitative alterations in motility and significantly slower flagella (45.5 µm/s), compared with those exposed to pre-immunization sera (69.2 µm/s). In summary, polyclonal anti-TcTLE serum significantly reduced the parasite's flagellar speed and cell infectivity. These findings support that KMP-11 could be important for parasite motility, and that by targeting its N-terminal peptide infectivity can be reduced

  10. 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.

  11. JAM-A is present in Mammalian Spermatozoa where it is Essential for Normal Motility

    PubMed Central

    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 ∼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 hyperactived 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 protein 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. PMID:18022613

  12. Sensory roles of neuronal cilia: cilia development, morphogenesis, and function in C. elegans.

    PubMed

    Bae, Young-Kyung; Barr, Maureen M

    2008-01-01

    In the free-living nematode Caenorhabditis elegans, cilia are found on the dendritic endings of sensory neurons. C. elegans cilia are classified as 'primary' or 'sensory' according to the '9+0' axonemal ultrastructure (nine doublet outer microtubules with no central microtubule pair) and lack of motility, characteristics of '9+2' cilia. The C. elegans ciliated nervous system allows the animal to perceive environmental stimuli and make appropriate developmental, physiological, and behavioral decisions. In vertebrates, the biological significance of primary cilia had been largely neglected. Recent findings have placed primary/sensory cilia in the center of cellular signaling and developmental processes. Studies using genetic model organisms such as C. elegans identified the link between ciliary dysfunction and human ciliopathies. Future studies in the worm will address important basic questions regarding ciliary development, morphogenesis, specialization, and signaling functions. PMID:18508635

  13. 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.

  14. Calmodulin-dependent protein kinases mediate calcium-induced slow motility of mammalian outer hair cells.

    PubMed

    Puschner, B; Schacht, J

    1997-08-01

    Cochlear outer hair cells in vitro respond to elevation of intracellular calcium with slow shape changes over seconds to minutes ('slow motility'). This process is blocked by general calmodulin antagonists suggesting the participation of calcium/calmodulin-dependent enzymatic reactions. The present study proposes a mechanism for these reactions. Length changes of outer hair cells isolated from the guinea pig cochlea were induced by exposure to the calcium ionophore ionomycin. ATP levels remained unaffected by this treatment ruling out depletion of ATP (by activation of calcium-dependent ATPases) as a cause of the observed shape changes. Involvement of protein kinases was suggested by the inhibition of shape changes by K252a, a broad-spectrum inhibitor of protein kinase activity. Furthermore, the inhibitors ML-7 and ML-9 blocked the shape changes at concentrations compatible with inhibition of myosin light chain kinase (MLCK). KN-62, an inhibitor of Ca2+/calmodulin-dependent protein kinase II (CaMKII), also attenuated the length changes. Inhibitors with selectivity for cyclic nucleotide-dependent protein kinases (H-89, staurosporine) were tested to assess potential additional contributions by such enzymes. The dose dependence of their action supported the notion that the most likely mechanism of slow motility involves phosphorylation reactions catalyzed by MLCK or CaMKII or both. PMID:9282907

  15. 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

  16. Autophagy and regulation of cilia function and assembly

    PubMed Central

    Orhon, I; Dupont, N; Pampliega, O; Cuervo, A M; Codogno, P

    2015-01-01

    Motile and primary cilia (PC) are microtubule-based structures located at the cell surface of many cell types. Cilia govern cellular functions ranging from motility to integration of mechanical and chemical signaling from the environment. Recent studies highlight the interplay between cilia and autophagy, a conserved cellular process responsible for intracellular degradation. Signaling from the PC recruits the autophagic machinery to trigger autophagosome formation. Conversely, autophagy regulates ciliogenesis by controlling the levels of ciliary proteins. The cross talk between autophagy and ciliated structures is a novel aspect of cell biology with major implications in development, physiology and human pathologies related to defects in cilium function. PMID:25361082

  17. Methods for imaging individual cilia in living echinoid embryos.

    PubMed

    Morris, Robert L; Pope, Hans W; Sholi, Adam N; Williams, Leah M; Ettinger, Chelsea R; Beacham, Gwendolyn M; Shintaku, Tatsushi; Abbott, Zachary D; Doherty, Elyse M

    2015-01-01

    The embryos of echinoids (sea urchins and sand dollars) serve as excellent models for studying cilia differentiation and stages of the cilia life cycle including ciliogenic initiation, growth, maintenance, and retraction. Early in echinoid development, uniform motile cilia form on all cells simultaneously but then rapidly differentiate into multiple cilia types that differ in morphology, motility, and signaling sensitivity. Metal ion treatments that shift germ layer boundaries and thereby "animalize" or "vegetalize" embryos can be used to enrich for low-abundance cilia types rendering those specialized cilia and the differentiation processes they exhibit much easier to study. The experimental advantages of having robust cilia growth and differentiation is tempered by the challenge of restraining ciliated embryos well enough to view the process of ciliogenesis live. We have developed four observation chambers as modifications of the Kiehart chamber for long-term light microscopic imaging of ciliated echinoid embryos. One of these systems employs paramagnetic beads to render ciliated larvae magnetic so they can be gently and reversibly trapped directly under the objective lens. With this magnetic trapping system, the larva can be positioned and repositioned until they achieve the orientation with the clearest view of any cilia of interest. These methods of gentle embryo restraint allow normal embryo development and the normal ciliogenic cycle and ciliary differentiation processes to continue in direct view. Sequential image series can then be collected and analyzed to quantitatively study the wide spectrum of cilia behaviors and properties that arise in developing echinoid embryos. PMID:25837394

  18. Primary cilia and forebrain development.

    PubMed

    Willaredt, Marc August; Tasouri, Evangelia; Tucker, Kerry L

    2013-01-01

    With a microtubule-based axoneme supporting its plasma membrane-ensheathed projection from the basal body of almost all cell types in the human body, and present in only one copy per cell, the primary cilium can be considered an organelle sui generis. Although it was first observed and recorded in histological studies from the late 19th century, the tiny structure was essentially forgotten for many decades. In the past ten years, however, scientists have turned their eyes once again upon primary cilia and realized that they are very important for the development of almost all organs in the mammalian body, especially those dependent upon the signaling from members Hedgehog family, such as Indian and Sonic hedgehog. In this review, we outline the roles that primary cilia play in forebrain development, not just in the crucial transduction of Sonic hedgehog signaling, but also new results showing that cilia are important for cell cycle progression in proliferating neural precursors. We will focus upon cerebral cortex development but will also discuss the importance of cilia for the embryonic hippocampus, olfactory bulb, and diencephalon. PMID:23085524

  19. Cilia driven flow networks in the brain

    NASA Astrophysics Data System (ADS)

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

    Neurons exchange soluble substances via the cerebrospinal fluid (CSF) that fills the ventricular system. The walls of the ventricular cavities are covered with motile cilia that constantly beat and thereby induce a directional flow. We recently discovered that cilia in the third ventricle generate a complex flow pattern leading to partitioning of the ventricular volume and site-directed transport paths along the walls. Transient and daily recurrent alterations in the cilia beating direction lead to changes in the flow pattern. This has consequences for delivery of CSF components along the near wall flow. The contribution of this cilia-induced flow to overall CSF flow remains to be investigated. The state-of-art lattice Boltzmann method is adapted for studying the CFS flow. The 3D geometry of the third ventricle at high resolution was reconstructed. Simulation of CSF flow without cilia in this geometry confirmed that the previous idea about unidirectional flow does not explain how different components of CSF can be delivered to their various target sites. We study the contribution of the cilia-induced flow pattern to overall CSF flow and identify target areas for site-specific delivery of CSF-constituents with respect to the temporal changes.

  20. 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. PMID:26597000

  1. 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. PMID:24930722

  2. Cildb: a knowledgebase for centrosomes and cilia

    PubMed Central

    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 PMID:20428338

  3. Using Xenopus Skin to Study Cilia Development and Function

    PubMed Central

    Werner, Michael E.; Mitchell, Brian J.

    2015-01-01

    Cilia are prevalent biological structures that are important for cell signaling and for generating fluid flow (or motility). Cilia are found throughout biology from single-celled organisms to vertebrates, and many model systems have been employed for their analysis. Here, we describe the use of Xenopus larval skin as a system for the study of ciliogenesis and ciliary function. In particular, we describe basic molecular and embryological manipulations and imaging techniques that have proven particularly useful for understanding the polarized beating of cilia and the generation of directed fluid flow (Werner & Mitchell, 2012). However, these same tools have the potential to benefit a large number of cilia-related biological questions. PMID:23522471

  4. Primary cilia are not calcium-responsive mechanosensors.

    PubMed

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

    2016-03-31

    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 cilium. This Ca(2+)-responsive mechanosensor 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 cancers. Here we report the complete lack of mechanically induced calcium increases in primary cilia, in tissues upon which this hypothesis has been based. We developed a transgenic mouse, Arl13b-mCherry-GECO1.2, expressing a ratiometric genetically encoded calcium indicator 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 Ca(2+) 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 signalling. PMID:27007841

  5. 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

  6. 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. PMID:26787743

  7. 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

  8. 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

  9. Assembly and dynamics of synthetic cilia

    NASA Astrophysics Data System (ADS)

    Sanchez, Tim

    2012-02-01

    From motility of simple protists to determining the handedness of complex vertebrates, highly conserved eukaryotic cilia and flagella are essential for the reproduction and survival of many biological organisms. Despite extensive studies, the exact mechanism by which individual components coordinate to produce ciliary beating patterns remains unknown. We describe a novel approach towards studying ciliary beating. Instead of deconstructing a fully functional organelle from the top-down, we describe a process by which synthetic cilia-like structures are assembled from the bottom-up. We find that simple mixtures of microtubules, kinesin clusters, and a bundling agent produce spontaneous oscillations in MT bundles, suggesting that self-organized beating may be a generic feature of internally driven bundles. Furthermore, bundles in close proximity spontaneously coordinate their beating to generate metachronal traveling waves, reminiscent of the waves seen in ciliary fields. These findings and future refinements of the system can potentially provide insights into general design principles required for engineering synthetic cilia as well as understanding the biological analogues.

  10. The zebrafish fleer gene encodes an essential regulator of cilia tubulin polyglutamylation.

    PubMed

    Pathak, Narendra; Obara, Tomoko; Mangos, Steve; Liu, Yan; Drummond, Iain A

    2007-11-01

    Cilia and basal bodies are essential organelles for a broad spectrum of functions, including the development of left-right asymmetry, kidney function, cerebrospinal fluid transport, generation of photoreceptor outer segments, and hedgehog signaling. Zebrafish fleer (flr) mutants exhibit kidney cysts, randomized left-right asymmetry, hydrocephalus, and rod outer segment defects, suggesting a pleiotropic defect in ciliogenesis. Positional cloning flr identified a tetratricopeptide repeat protein homologous to the Caenorhabditis elegans protein DYF1 that was highly expressed in ciliated cells. flr pronephric cilia were shortened and showed a reduced beat amplitude, and olfactory cilia were absent in mutants. flr cilia exhibited ultrastructural defects in microtubule B-tubules, similar to axonemes that lack tubulin posttranslational modifications (polyglutamylation or polyglycylation). flr cilia showed a dramatic reduction in cilia polyglutamylated tubulin, indicating that flr encodes a novel modulator of tubulin polyglutamylation. We also found that the C. elegans flr homologue, dyf-1, is also required for tubulin polyglutamylation in sensory neuron cilia. Knockdown of zebrafish Ttll6, a tubulin polyglutamylase, specifically eliminated tubulin polyglutamylation and cilia formation in olfactory placodes, similar to flr mutants. These results are the first in vivo evidence that tubulin polyglutamylation is required for vertebrate cilia motility and structure, and, when compromised, results in failed ciliogenesis. PMID:17761526

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

    PubMed

    Marra, Amanda N; Li, Yue; Wingert, Rebecca A

    2016-09-01

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

    PubMed

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

    2015-09-01

    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

  13. 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

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

    PubMed

    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

  15. 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. PMID:9453710

  16. Kinesin-13 regulates the quantity and quality of tubulin inside cilia

    PubMed Central

    Vasudevan, Krishna Kumar; Jiang, Yu-Yang; Lechtreck, Karl F.; Kushida, Yasuharu; Alford, Lea M.; Sale, Winfield S.; Hennessey, Todd; Gaertig, Jacek

    2015-01-01

    Kinesin-13, an end depolymerizer of cytoplasmic and spindle microtubules, also affects the length of cilia. However, in different models, depletion of kinesin-13 either lengthens or shortens cilia, and therefore the exact function of kinesin-13 in cilia remains unclear. We generated null mutations of all kinesin-13 paralogues in the ciliate Tetrahymena. One of the paralogues, Kin13Ap, localizes to the nuclei and is essential for nuclear divisions. The remaining two paralogues, Kin13Bp and Kin13Cp, localize to the cell body and inside assembling cilia. Loss of both Kin13Bp and Kin13Cp resulted in slow cell multiplication and motility, overgrowth of cell body microtubules, shortening of cilia, and synthetic lethality with either paclitaxel or a deletion of MEC-17/ATAT1, the α-tubulin acetyltransferase. The mutant cilia assembled slowly and contained abnormal tubulin, characterized by altered posttranslational modifications and hypersensitivity to paclitaxel. The mutant cilia beat slowly and axonemes showed reduced velocity of microtubule sliding. Thus kinesin-13 positively regulates the axoneme length, influences the properties of ciliary tubulin, and likely indirectly, through its effects on the axonemal microtubules, affects the ciliary dynein-dependent motility. PMID:25501369

  17. 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

  18. Analysis of primary cilia in the developing mouse brain.

    PubMed

    Paridaen, Judith T M L; Huttner, Wieland B; Wilsch-Bräuninger, Michaela

    2015-01-01

    Stem and progenitor cells in the developing mammalian brain are highly polarized cells that carry a primary cilium protruding into the brain ventricles. Here, cilia detect signals present in the cerebrospinal fluid that fills the ventricles. Recently, striking observations have been made regarding the dynamics of primary cilia in mitosis and cilium reformation after cell division. In neural progenitors, primary cilia are not completely disassembled during cell division, and some ciliary membrane remnant can be inherited by one daughter cell that tends to maintain a progenitor fate. Furthermore, newborn differentiating cells grow a primary cilium on their basolateral plasma membrane, in spite of them possessing apical membrane and adherens junctions, and thus change the environment to which the primary cilium is exposed. These phenomena are proposed to be involved in cell fate determination and delamination of daughter cells in conjunction with the production of neurons. Here, we describe several methods that can be used to study the structure, localization, and dynamics of primary cilia in the developing mouse brain; these include time-lapse imaging of live mouse embryonic brain tissues, and analysis of primary cilia structure and localization using correlative light- and electron- and serial-block-face scanning electron microscopy. PMID:25837388

  19. 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.

  20. [Cilia and renal cysts].

    PubMed

    Paces-Fessy, Mélanie

    2014-11-01

    Advances in genomics, bioinformatics and the creation of model organisms have identified many genes associated with polycystic kidney diseases. Historically, these genes were not necessarily associated with ciliopathies, but it appeared that many connections can be made between the cystic kidney disease and function of the primary cilium. Indeed, the proteins encoded by these genes are localized to the cilium itself, to the basal body or are known to regulate the expression and localization of ciliary proteins. The goal of this article is to describe the multiple cellular processes that may lead to the development of renal cysts if they are deregulated. These include changes in proliferation rate, cell polarity or signaling pathways involved in embryonic kidney development. To highlight the role of the primary cilium in cystogenesis, I will discuss several studies investigating the function of ciliary genes and cilia in the kidneys of different model organisms. PMID:25388585

  1. Intracellular and extracellular forces drive primary cilia movement.

    PubMed

    Battle, Christopher; Ott, Carolyn M; Burnette, Dylan T; Lippincott-Schwartz, Jennifer; Schmidt, Christoph F

    2015-02-01

    Primary cilia are ubiquitous, microtubule-based organelles that play diverse roles in sensory transduction in many eukaryotic cells. They interrogate the cellular environment through chemosensing, osmosensing, and mechanosensing using receptors and ion channels in the ciliary membrane. Little is known about the mechanical and structural properties of the cilium and how these properties contribute to ciliary perception. We probed the mechanical responses of primary cilia from kidney epithelial cells [Madin-Darby canine kidney-II (MDCK-II)], which sense fluid flow in renal ducts. We found that, on manipulation with an optical trap, cilia deflect by bending along their length and pivoting around an effective hinge located below the basal body. The calculated bending rigidity indicates weak microtubule doublet coupling. Primary cilia of MDCK cells lack interdoublet dynein motors. Nevertheless, we found that the organelles display active motility. 3D tracking showed correlated fluctuations of the cilium and basal body. These angular movements seemed random but were dependent on ATP and cytoplasmic myosin-II in the cell cortex. We conclude that force generation by the actin cytoskeleton surrounding the basal body results in active ciliary movement. We speculate that actin-driven ciliary movement might tune and calibrate ciliary sensory functions. PMID:25605896

  2. Intracellular and extracellular forces drive primary cilia movement

    PubMed Central

    Battle, Christopher; Ott, Carolyn M.; Burnette, Dylan T.; Lippincott-Schwartz, Jennifer; Schmidt, Christoph F.

    2015-01-01

    Primary cilia are ubiquitous, microtubule-based organelles that play diverse roles in sensory transduction in many eukaryotic cells. They interrogate the cellular environment through chemosensing, osmosensing, and mechanosensing using receptors and ion channels in the ciliary membrane. Little is known about the mechanical and structural properties of the cilium and how these properties contribute to ciliary perception. We probed the mechanical responses of primary cilia from kidney epithelial cells [Madin–Darby canine kidney-II (MDCK-II)], which sense fluid flow in renal ducts. We found that, on manipulation with an optical trap, cilia deflect by bending along their length and pivoting around an effective hinge located below the basal body. The calculated bending rigidity indicates weak microtubule doublet coupling. Primary cilia of MDCK cells lack interdoublet dynein motors. Nevertheless, we found that the organelles display active motility. 3D tracking showed correlated fluctuations of the cilium and basal body. These angular movements seemed random but were dependent on ATP and cytoplasmic myosin-II in the cell cortex. We conclude that force generation by the actin cytoskeleton surrounding the basal body results in active ciliary movement. We speculate that actin-driven ciliary movement might tune and calibrate ciliary sensory functions. PMID:25605896

  3. 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.

  4. Sperm-Associated Antigen 6 (SPAG6) Deficiency and Defects in Ciliogenesis and Cilia Function: Polarity, Density, and Beat

    PubMed Central

    Teves, Maria E.; Sears, Patrick R.; Li, Wei; Zhang, Zhengang; Tang, Waixing; van Reesema, Lauren; Costanzo, Richard M.; Davis, C. William; Knowles, Michael R.; Strauss, Jerome F.; Zhang, Zhibing

    2014-01-01

    SPAG6, an axoneme central apparatus protein, is essential for function of ependymal cell cilia and sperm flagella. A significant number of Spag6-deficient mice die with hydrocephalus, and surviving males are sterile because of sperm motility defects. In further exploring the ciliary dysfunction in Spag6-null mice, we discovered that cilia beat frequency was significantly reduced in tracheal epithelial cells, and that the beat was not synchronized. There was also a significant reduction in cilia density in both brain ependymal and trachea epithelial cells, and cilia arrays were disorganized. The orientation of basal feet, which determines the direction of axoneme orientation, was apparently random in Spag6-deficient mice, and there were reduced numbers of basal feet, consistent with reduced cilia density. The polarized epithelial cell morphology and distribution of intracellular mucin, α-tubulin, and the planar cell polarity protein, Vangl2, were lost in Spag6-deficient tracheal epithelial cells. Polarized epithelial cell morphology and polarized distribution of α-tubulin in tracheal epithelial cells was observed in one-week old wild-type mice, but not in the Spag6-deficient mice of the same age. Thus, the cilia and polarity defects appear prior to 7 days post-partum. These findings suggest that SPAG6 not only regulates cilia/flagellar motility, but that in its absence, ciliogenesis, axoneme orientation, and tracheal epithelial cell polarity are altered. PMID:25333478

  5. Comparative study of the primary cilia in thyrocytes of adult mammals.

    PubMed

    Utrilla, J C; Gordillo-Martínez, F; Gómez-Pascual, A; Fernández-Santos, J M; Garnacho, C; Vázquez-Román, V; Morillo-Bernal, J; García-Marín, R; Jiménez-García, A; Martín-Lacave, I

    2015-10-01

    Since their discovery in different human tissues by Zimmermann in 1898, primary cilia have been found in the vast majority of cell types in vertebrates. Primary cilia are considered to be cellular antennae that occupy an ideal cellular location for the interpretation of information both from the environment and from other cells. To date, in mammalian thyroid gland, primary cilia have been found in the thyrocytes of humans and dogs (fetuses and adults) and in rat embryos. The present study investigated whether the existence of this organelle in follicular cells is a general event in the postnatal thyroid gland of different mammals, using both immunolabeling by immunofluorescence and electron microscopy. Furthermore, we aimed to analyse the presence of primary cilia in various thyroid cell lines. According to our results, primary cilia are present in the adult thyroid gland of most mammal species we studied (human, pig, guinea pig and rabbit), usually as a single copy per follicular cell. Strikingly, they were not found in rat or mouse thyroid tissues. Similarly, cilia were also observed in all human thyroid cell lines tested, both normal and neoplastic follicular cells, but not in cultured thyrocytes of rat origin. We hypothesize that primary cilia could be involved in the regulation of normal thyroid function through specific signaling pathways. Nevertheless, further studies are needed to shed light on the permanence of these organelles in the thyroid gland of most species during postnatal life. PMID:26228270

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

    PubMed Central

    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 PMID:24596149

  7. Cilia and cilia-associated proteins in cancer

    PubMed Central

    Seeger-Nukpezah, Tamina; Little, Joy L.; Serzhanova, Victoria; Golemis, Erica A.

    2013-01-01

    The primary cilium is a well-established target in the pathogenesis of numerous developmental and chronic disorders, and more recently is attracting interest as a structure relevant to cancer. Here we discuss mechanisms by which changes in cilia can contribute to the formation and growth of tumors. We emphasize the cancer-relevance of cilia-dependent signaling pathways and proteins including mTOR, VHL, TSC, WNT, Aurora-A, NEDD9, and Hedgehog, and highlight the emerging role of ciliary dysfunction in renal cell carcinoma, medulloblastoma, and breast cancer. PMID:24982684

  8. Cytoplasmic carboxypeptidase 5 regulates tubulin glutamylation and zebrafish cilia formation and function

    PubMed Central

    Pathak, Narendra; Austin-Tse, Christina A.; Liu, Yan; Vasilyev, Aleksandr; Drummond, Iain A.

    2014-01-01

    Glutamylation is a functionally important tubulin posttranslational modification enriched on stable microtubules of neuronal axons, mitotic spindles, centrioles, and cilia. In vertebrates, balanced activities of tubulin glutamyl ligase and cytoplasmic carboxypeptidase deglutamylase enzymes maintain organelle- and cell type–specific tubulin glutamylation patterns. Tubulin glutamylation in cilia is regulated via restricted subcellular localization or expression of tubulin glutamyl ligases (ttlls) and nonenzymatic proteins, including the zebrafish TPR repeat protein Fleer/Ift70. Here we analyze the expression patterns of ccp deglutamylase genes during zebrafish development and the effects of ccp gene knockdown on cilia formation, morphology, and tubulin glutamylation. The deglutamylases ccp2, ccp5, and ccp6 are expressed in ciliated cells, whereas ccp1 expression is restricted to the nervous system. Only ccp5 knockdown increases cilia tubulin glutamylation, induces ciliopathy phenotypes, including axis curvature, hydrocephalus, and pronephric cysts, and disrupts multicilia motility, suggesting that Ccp5 is the principal tubulin deglutamylase that maintains functional levels of cilia tubulin glutamylation. The ability of ccp5 knockdown to restore cilia tubulin glutamylation in fleer/ift70 mutants and rescue pronephric multicilia formation in both fleer- and ift88-deficient zebrafish indicates that tubulin glutamylation is a key driver of ciliogenesis. PMID:24743595

  9. 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. PMID:25837391

  10. Transport of the outer dynein arm complex to cilia requires a cytoplasmic protein Lrrc6.

    PubMed

    Inaba, Yasuko; Shinohara, Kyosuke; Botilde, Yanick; Nabeshima, Ryo; Takaoka, Katsuyoshi; Ajima, Rieko; Lamri, Lynda; Takeda, Hiroyuki; Saga, Yumiko; Nakamura, Tetsuya; Hamada, Hiroshi

    2016-07-01

    Lrrc6 encodes a cytoplasmic protein that is expressed specifically in cells with motile cilia including the node, trachea and testes of the mice. A mutation of Lrrc6 has been identified in human patients with primary ciliary dyskinesia (PCD). Mutant mice lacking Lrrc6 show typical PCD defects such as hydrocephalus and laterality defects. We found that in the absence of Lrrc6, the morphology of motile cilia remained normal, but their motility was completely lost. The 9 + 2 arrangement of microtubules remained normal in Lrrc6(-/-) mice, but the outer dynein arms (ODAs), the structures essential for the ciliary beating, were absent from the cilia. In the absence of Lrrc6, ODA proteins such as DNAH5, DNAH9 and IC2, which are assembled in the cytoplasm and transported to the ciliary axoneme, remained in the cytoplasm and were not transported to the ciliary axoneme. The IC2-IC1 interaction, which is the first step of ODA assembly, was normal in Lrrc6(-/-) mice testes. Our results suggest that ODA proteins may be transported from the cytoplasm to the cilia by an Lrrc6-dependent mechanism. PMID:27353389

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

    PubMed Central

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

    2015-01-01

    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;Ift88fl/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. SIGNIFICANCE STATEMENT 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

  12. The zebrafish foxj1a transcription factor regulates cilia function in response to injury and epithelial stretch.

    PubMed

    Hellman, Nathan E; Liu, Yan; Merkel, Erin; Austin, Christina; Le Corre, Stephanie; Beier, David R; Sun, Zhaoxia; Sharma, Neeraj; Yoder, Bradley K; Drummond, Iain A

    2010-10-26

    Cilia are essential for normal organ function and developmental patterning, but their role in injury and regeneration responses is unknown. To probe the role of cilia in injury, we analyzed the function of foxj1, a transcriptional regulator of cilia genes, in response to tissue damage and renal cyst formation. Zebrafish foxj1a, but not foxj1b, was rapidly induced in response to epithelial distension and stretch, kidney cyst formation, acute kidney injury by gentamicin, and crush injury in spinal cord cells. Obstruction-induced up-regulation of foxj1a was not inhibited by cycloheximide, identifying foxj1a as a primary response gene to epithelial injury. Foxj1 was also dramatically up-regulated in murine cystic kidney disease epithelia [jck/jck (nek8) and Ift88Tg737Rpw(-/-)] as well as in response to kidney ischemia-reperfusion injury. Obstruction of the zebrafish pronephric tubule caused a rapid increase in cilia beat rate that correlated tightly with expanded tubule diameter and epithelial stretch. Zebrafish foxj1a was specifically required for cilia motility. Enhanced foxj1a expression in obstructed tubules induced cilia motility target genes efhc1, tektin-1, and dnahc9. foxj1a-deficient embryos failed to up-regulate efhc1, tektin-1, and dnahc9 and could not maintain enhanced cilia beat rates after obstruction, identifying an essential role for foxj1 in modulating cilia function after injury. These studies reveal that activation of a Foxj1 transcriptional network of ciliogenic genes is an evolutionarily conserved response to multiple forms of tissue damage and highlight enhanced cilia function as a previously uncharacterized component of organ homeostasis. PMID:20937855

  13. Ophiobolin A from Bipolaris oryzae Perturbs Motility and Membrane Integrities of Porcine Sperm and Induces Cell Death on Mammalian Somatic Cell Lines

    PubMed Central

    Bencsik, Ottó; Papp, Tamás; Berta, Máté; Zana, Annamária; Forgó, Péter; Dombi, György; Andersson, Maria A.; Salkinoja-Salonen, Mirja; Vágvölgyi, Csaba; Szekeres, András

    2014-01-01

    Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1–2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration. PMID:25251540

  14. Ophiobolin A from Bipolaris oryzae perturbs motility and membrane integrities of porcine sperm and induces cell death on mammalian somatic cell lines.

    PubMed

    Bencsik, Ottó; Papp, Tamás; Berta, Máté; Zana, Annamária; Forgó, Péter; Dombi, György; Andersson, Maria A; Salkinoja-Salonen, Mirja; Vágvölgyi, Csaba; Szekeres, András

    2014-09-01

    Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1-2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration. PMID:25251540

  15. 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

  16. 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. PMID:25921740

  17. Multiple cilia suppress tumour formation.

    PubMed

    Eberhart, Charles

    2016-04-01

    Primary cilia are cellular structures that have important functions in development and disease. The suppression of multiciliate differentiation of choroid plexus precursors, and maintenance of a single primary cilium by Notch1, is now shown to be involved in choroid plexus tumour formation. PMID:27027488

  18. The more we know, the more we have to discover: an exciting future for understanding cilia and ciliopathies.

    PubMed

    Benmerah, Alexandre; Durand, Bénédicte; Giles, Rachel H; Harris, Tess; Kohl, Linda; Laclef, Christine; Meilhac, Sigolène M; Mitchison, Hannah M; Pedersen, Lotte B; Roepman, Ronald; Swoboda, Peter; Ueffing, Marius; Bastin, Philippe

    2015-01-01

    The Cilia 2014 conference was organised by four European networks: the Ciliopathy Alliance, the Groupement de Recherche CIL, the Nordic Cilia and Centrosome Network and the EU FP7 programme SYSCILIA. More than 400 delegates from 27 countries gathered at the Institut Pasteur conference centre in Paris, including 30 patients and patient representatives. The meeting offered a unique opportunity for exchange between different scientific and medical communities. Major highlights included new discoveries about the roles of motile and immotile cilia during development and homeostasis, the mechanism of cilium construction, as well as progress in diagnosis and possible treatment of ciliopathies. The contributions to the cilia field of flagellated infectious eukaryotes and of systems biology were also presented. PMID:25974046

  19. 3D structure of eukaryotic flagella/cilia by cryo-electron tomography

    PubMed Central

    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. PMID:27493552

  20. General and cell-type specific mechanisms target TRPP2/PKD-2 to cilia.

    PubMed

    Bae, Young-Kyung; Qin, Hongmin; Knobel, Karla M; Hu, Jinghua; Rosenbaum, Joel L; Barr, Maureen M

    2006-10-01

    Ciliary localization of the transient receptor potential polycystin 2 channel (TRPP2/PKD-2) is evolutionarily conserved, but how TRPP2 is targeted to cilia is not known. In this study, we characterize the motility and localization of PKD-2, a TRPP2 homolog, in C. elegans sensory neurons. We demonstrate that GFP-tagged PKD-2 moves bidirectionally in the dendritic compartment. Furthermore, we show a requirement for different molecules in regulating the ciliary localization of PKD-2. PKD-2 is directed to moving dendritic particles by the UNC-101/adaptor protein 1 (AP-1) complex. When expressed in non-native neurons, PKD-2 remains in cell bodies and is not observed in dendrites or cilia, indicating that cell-type specific factors are required for directing PKD-2 to the dendrite. PKD-2 stabilization in cilia and cell bodies requires LOV-1, a functional partner and a TRPP1 homolog. In lov-1 mutants, PKD-2 is greatly reduced in cilia and forms abnormal aggregates in neuronal cell bodies. Intraflagellar transport (IFT) is not essential for PKD-2 dendritic motility or access to the cilium, but may regulate PKD-2 ciliary abundance. We propose that both general and cell-type-specific factors govern TRPP2/PKD-2 subcellular distribution by forming at least two steps involving somatodendritic and ciliary sorting decisions. PMID:16943275

  1. Characterization of Tetratricopeptide Repeat-Containing Proteins Critical for Cilia Formation and Function

    PubMed Central

    Xu, Yanan; Cao, Jingli; Huang, Shan; Feng, Di; Zhang, Wei; Zhu, Xueliang; Yan, Xiumin

    2015-01-01

    Cilia formation and function require a special set of trafficking machinery termed intraflagellar transport (IFT), consisting mainly of protein complexes IFT-A, IFT-B, BBSome, and microtubule-dependent molecular motors. Tetratricopeptide repeat-containing (TTC) proteins are widely involved in protein complex formation. Nine of them are known to serve as components of the IFT or BBSome complexes. How many TTC proteins are cilia-related and how they function, however, remain unclear. Here we show that twenty TTC genes were upregulated by at least 2-fold during the differentiation of cultured mouse tracheal epithelial cells (MTECs) into multiciliated cells. Our systematic screen in zebrafish identified four novel TTC genes, ttc4, -9c, -36, and -39c, that are critical for cilia formation and motility. Accordingly, their zebrafish morphants displayed typical ciliopathy-related phenotypes, including curved body, abnormal otolith, hydrocephalus, and defective left-right patterning. The morphants of ttc4 and ttc25, a known cilia-related gene, additionally showed pronephric cyst formation. Immunoprecipitation indicated associations of TTC4, -9c, -25, -36, and -39c with components or entire complexes of IFT-A, IFT-B, or BBSome, implying their participations in IFT or IFT-related activities. Our results provide a global view for the relationship between TTC proteins and cilia. PMID:25860617

  2. 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

  3. 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

  4. 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. PMID:23487771

  5. 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.

  6. Swimming like algae: biomimetic soft artificial cilia.

    PubMed

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

    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

  7. 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

  8. 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. PMID:23245658

  9. 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.

  10. 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

  11. HEATR2 Plays a Conserved Role in Assembly of the Ciliary Motile Apparatus

    PubMed Central

    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

    2014-01-01

    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. PMID:25232951

  12. Manipulating Cilia Using the 3DFM

    NASA Astrophysics Data System (ADS)

    Fisher, Jay K.; O'Brien, E. Timothy; Taylor, R. M.; Davis, C. W.; Matsui, H.; Vicci, L.; Matthews, G.; Cribb, J.; Desai, K.; Wilde, B.; Superfine, R.

    2003-11-01

    Mucus flow generated by beating cilia projecting from epithelial cells is responsible for the removal of pathogens in the lungs. When the hydrodynamics of this mucociliary clearance system fails, as happens in the condition of Cystic Fibrosis, ensuing infections can destroy the lungs. A complex phenomenology includes the force generation of the cilia, the coupling of the cilia tips to the overlying mucus during the power stroke, and finally, it is speculated that the cilia act as force sensors to allow the control of the mucus viscosity and volume. We have designed a system that allows the measurement of 3 dimensional forces within an optical microscope using magnetic forces and superparamagnetic beads. We have functionalized the beads with antibodies and attached them to the cilia of human lung cell cultures. A laser is focused onto the bead whose motion is tracked using back focal plane detection of the forward scattered light. We have measured the 3 dimensional trajectory of beads attached to cilia beating up to 15Hz in human lung cell cultures with 10 nm, 1msec resolution. When forces are applied to the bead, we observe changes in the trajectory and velocity, implying a strong mechanoresponse of the cell as sensed by the cilia.

  13. Cilia, Wnt signaling, and the cytoskeleton.

    PubMed

    May-Simera, Helen L; Kelley, Matthew W

    2012-01-01

    Primary cilia have recently been highlighted as key regulators in development and disease. This review focuses on current work demonstrating the broad role of cilia-related proteins in developmental signaling systems. Of particular consideration is the importance of the basal body region, located at the base of the cilium, in its role as a focal point for many signaling pathways and as a microtubule organizing center. As the cilium is effectively a microtubular extension of the cytoskeleton, investigating connections between the cilium and the cytoskeleton provides greater insight into signaling and cell function. Of the many signaling pathways associated with primary cilia, the most extensively studied in association with the cytoskeleton and cytoskeletal rearrangements are both canonical and non-canonical Wnt pathways. One of the key concepts currently emerging is a possible additional role for the traditionally 'cilia-related' proteins in other aspects of cellular processes. In many cases, disruption of such processes manifests at the level of the cilium. While the involvement of cilia and cilia-related proteins in signaling pathways is currently being unraveled, there is a growing body of evidence to support the notion that ciliary proteins are required not only for regulation of Wnt signaling, but also as downstream effectors of Wnt signaling. This review summarizes recent advances in our understanding of the involvement of cilia and basal body proteins in Wnt signaling pathways. PMID:23351924

  14. Immunofluorescent staining of septins in primary cilia.

    PubMed

    Kim, M S; Froese, C D; Xie, H; Trimble, W S

    2016-01-01

    Primary cilia are cellular antennae that receive and transduce extracellular cues. These microtubule-rich structures are comprised of at least three distinct ciliary compartments: basal bodies, transition zone, and axoneme. Septins have been implicated in cilia function at the transition zone, but accumulating evidence suggests that they localize predominantly within the axoneme. Here, we describe three fixation conditions that preserve the substructure of primary cilia and demonstrate known ciliary proteins that localize to these distinct ciliary substructures. Finally, we show immunostaining and live microscopy methods to detect septins within the axoneme. PMID:27473914

  15. 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

  16. Transmembrane protein OSTA-1 shapes sensory cilia morphology via regulation of intracellular membrane trafficking in C. elegans

    PubMed Central

    Olivier-Mason, Anique; Wojtyniak, Martin; Bowie, Rachel V.; Nechipurenko, Inna V.; Blacque, Oliver E.; Sengupta, Piali

    2013-01-01

    The structure and function of primary cilia are critically dependent on intracellular trafficking pathways that transport ciliary membrane and protein components. The mechanisms by which these trafficking pathways are regulated are not fully characterized. Here we identify the transmembrane protein OSTA-1 as a new regulator of the trafficking pathways that shape the morphology and protein composition of sensory cilia in C. elegans. osta-1 encodes an organic solute transporter alpha-like protein, mammalian homologs of which have been implicated in membrane trafficking and solute transport, although a role in regulating cilia structure has not previously been demonstrated. We show that mutations in osta-1 result in altered ciliary membrane volume, branch length and complexity, as well as defects in localization of a subset of ciliary transmembrane proteins in different sensory cilia types. OSTA-1 is associated with transport vesicles, localizes to a ciliary compartment shown to house trafficking proteins, and regulates both retrograde and anterograde flux of the endosome-associated RAB-5 small GTPase. Genetic epistasis experiments with sensory signaling, exocytic and endocytic proteins further implicate OSTA-1 as a crucial regulator of ciliary architecture via regulation of cilia-destined trafficking. Our findings suggest that regulation of transport pathways in a cell type-specific manner contributes to diversity in sensory cilia structure and might allow dynamic remodeling of ciliary architecture via multiple inputs. PMID:23482491

  17. 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

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

    2007-04-24

    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

  18. A Conditional Mutant Having Paralyzed Cilia and a Block in Cytokinesis Is Rescued by Cytoplasmic Exchange in Tetrahymena Thermophila

    PubMed Central

    Pennock, D. G.; Thatcher, T.; Bowen, J.; Bruns, P. J.; Gorovsky, M. A.

    1988-01-01

    Nineteen mutants that are conditional for both the ability to regain motility following deciliation and the ability to grow were isolated. The mutations causing slow growth were placed into five complementation groups. None of the mutations appears to affect energy production as all mutants remained motile at the restrictive temperature. In three complementation groups protein synthesis and the levels of mRNA encoding α-tubulin or actin were largely unaffected at the restrictive temperature, consistent with the hypothesis that mutations in these three groups directly affect the assembly of functional cilia and growth. Complementation group 1 was chosen for further characterization. Both phenotypes were shown to be linked, suggesting they are caused by a single mutation. Group 1 mutants regenerated cilia at the restrictive temperature, but the cilia were nonmotile. This mutation also caused a block in cytokinesis at the restrictive temperature but did not affect nuclear divisions or DNA synthesis. The block in cell division was transiently rescued by wild-type cytoplasm exchanged when mutants were paired with wild-type cells during conjugation (round 1 of genomic exclusion). Thus, at least one mutation has been isolated that affects assembly of some microtubule-based structures in Tetrahymena (cilia during regeneration) but not others (nuclei divide at 38°), and the product of this gene is likely to play a role in both ciliary function and in cytokinesis. PMID:3224807

  19. The Zebrafish Orthologue of the Dyslexia Candidate Gene DYX1C1 Is Essential for Cilia Growth and Function

    PubMed Central

    Chandrasekar, Gayathri; Vesterlund, Liselotte; Hultenby, Kjell; Tapia-Páez, Isabel; Kere, Juha

    2013-01-01

    DYX1C1, a susceptibility gene for dyslexia, encodes a tetratricopeptide repeat domain containing protein that has been implicated in neuronal migration in rodent models. The developmental role of this gene remains unexplored. To understand the biological function(s) of zebrafish dyx1c1 during embryonic development, we cloned the zebrafish dyx1c1 and used morpholino-based knockdown strategy. Quantitative real-time PCR analysis revealed the presence of dyx1c1 transcripts in embryos, early larval stages and in a wide range of adult tissues. Using mRNA in situ hybridization, we show here that dyx1c1 is expressed in many ciliated tissues in zebrafish. Inhibition of dyx1c1 produced pleiotropic phenotypes characteristically associated with cilia defects such as body curvature, hydrocephalus, situs inversus and kidney cysts. We also demonstrate that in dyx1c1 morphants, cilia length is reduced in several organs including Kupffer’s vesicle, pronephros, spinal canal and olfactory placode. Furthermore, electron microscopic analysis of cilia in dyx1c1 morphants revealed loss of both outer (ODA) and inner dynein arms (IDA) that have been shown to be required for cilia motility. Considering all these results, we propose an essential role for dyx1c1 in cilia growth and function. PMID:23650548

  20. Radial Spoke Protein 3 Is a Mammalian Protein Kinase A-anchoring Protein That Binds ERK1/2*

    PubMed Central

    Jivan, Arif; Earnest, Svetlana; Juang, Yu-Chi; Cobb, Melanie H.

    2009-01-01

    Initially identified in Chlamydomonas, RSP3 (radial spoke protein 3) is 1 of more than 20 identified radial spoke structural components of motile cilia and is required for axonemal sliding and flagellar motility. The mammalian orthologs for this and other radial spoke proteins, however, remain to be characterized. We found mammalian RSP3 to bind to the MAPK ERK2 through a yeast two-hybrid screen designed to identify interacting proteins that have a higher affinity for the phosphorylated, active form of the protein kinase. Consistent with the screening result, the human homolog, RSPH3, interacts with and is a substrate for ERK1/2. Moreover, RSPH3 is a protein kinase A-anchoring protein (AKAP) that scaffolds the cAMP-dependent protein kinase holoenzyme. The binding of RSPH3 to the regulatory subunits of cAMP-dependent protein kinase, RIIα and RIIβ, is regulated by ERK1/2 activity and phosphorylation. Here we describe an ERK1/2-interacting AKAP and suggest a mechanism by which cAMP-dependent protein kinase-AKAP binding can be modulated by the activity of other enzymes. PMID:19684019

  1. Detection of primary cilia in human glioblastoma

    PubMed Central

    Sarkisian, Matthew R.; Siebzehnrubl, Dorit; Hoang-Minh, Lan; Deleyrolle, Loic; Silver, Daniel J.; Siebzehnrubl, Florian A.; Guadiana, Sarah M.; Srivinasan, Gayathri; Semple-Rowland, Susan; Harrison, Jeffrey K.; Steindler, Dennis A.; Reynolds, Brent A.

    2015-01-01

    Glioblastoma (GBM) is the most common malignant adult brain tumor and carries a poor prognosis due to primary and acquired resistance. While many cellular features of GBM have been documented, it is unclear if cells within these tumors extend a primary cilium, an organelle whose associated signaling pathways may regulate proliferation, migration, and survival of neural precursor and tumor cells. Using immunohistochemical and electron microscopy (EM) techniques, we screened human GBM tumor biopsies and primary cell lines for cilia. Immunocytochemical staining of five primary GBM cell lines revealed that between 8 and 25 % of the cells in each line possessed gamma tubulin-positive basal bodies from which extended acetylated, alpha-tubulin-positive axonemes. EM analyses confirmed the presence of cilia at the cell surface and revealed that their axonemes contained organized networks of microtubules, a structural feature consistent with our detection of IFT88 and Arl13b, two trafficked cilia proteins, along the lengths of the axonemes. Notably, cilia were detected in each of 23 tumor biopsies (22 primary and 1 recurrent) examined. These cilia were distributed across the tumor landscape including regions proximal to the vasculature and within necrotic areas. Moreover, ciliated cells within these tumors co-stained with Ki67, a marker for actively dividing cells, and ZEB1, a transcription factor that is upregulated in GBM and linked to tumor initiation, invasion, and chemoresistance. Collectively, our data show that subpopulations of cells within human GBM tumors are ciliated. In view of mounting evidence supporting roles of primary cilia in tumor initiation and propagation, it is likely that further study of the effects of cilia on GBM tumor cell function will improve our understanding of GBM pathogenesis and may provide new directions for GBM treatment strategies. PMID:24510433

  2. Developmental Signaling: Does It Bridge the Gap Between Cilia Dysfunction and Renal Cystogenesis?

    PubMed Central

    Tran, Pamela V.; Sharma, Madhulika; Li, Xiaogang; Calvet, James P.

    2015-01-01

    For more than a decade, evidence has accumulated linking dysfunction of primary cilia to renal cystogenesis, yet molecular mechanisms remain undefined. The pathogenesis of renal cysts is complex, involving multiple cellular aberrations and signaling pathways. Adding to this complexity, primary cilia exhibit multiple roles in a context-dependent manner. On renal epithelial cells, primary cilia act as mechanosensors and trigger extracellular Ca2+ influx in response to laminar fluid flow. During mammalian development, primary cilia mediate the Hedgehog (Hh), Wnt, and Notch pathways, which control cell proliferation and differentiation, and tissue morphogenesis. Further, experimental evidence suggests the developmental state of the kidney strongly influences renal cystic disease. Thus, we review evidence for regulation of Ca2+ and cAMP, key molecules in renal cystogenesis, at the primary cilium, the role of Hh, Wnt, and Notch signaling in renal cystic disease, and the interplay between these developmental pathways and Ca2+ signaling. Indeed if these developmental pathways influence renal cystogenesis, these may represent novel therapeutic targets that can be integrated into a combination therapy for renal cystic disease. PMID:24861210

  3. Ciliary motility activity measurement using a dense optical flow algorithm.

    PubMed

    Parrilla, Eduardo; Armengot, Miguel; Mata, Manuel; Cortijo, Julio; Riera, Jaime; Hueso, José L; Moratal, David

    2013-01-01

    Persistent respiratory syncytial virus (RSV) infections have been associated with the exacerbation of chronic inflammatory diseases, including chronic obstructive pulmonary disease (COPD). This virus infects the respiratory epithelium, leading to chronic inflammation, and induces the release of mucins and the loss of cilia activity, two factors that determine mucus clearance and the increase in sputum volume. In this study, an automatic method has been established to determine the ciliary motility activity from cell cultures by means of optical flow computation, and has been applied to 136 control cultures and to 144 RSV-infected cultures. The control group presented an average of cell surface with cilia motility per field of 41 ± 15 % (mean ± standard deviation), while the infected group presented a 11 ± 5 %, t-Student p<0.001. The cutoff value to classify a infected specimen was <17.89 % (sensitivity 0.94, specificity 0.93). This methodology has proved to be a robust technique to evaluate cilia motility in cell cultures. PMID:24110720

  4. Stall Force and Response of Lung Cilia

    NASA Astrophysics Data System (ADS)

    Superfine, Richard; Hill, David; Swaminathan, Vinay; O'Brien, E. Timothy; Boucher, Ric; Button, Brian; Estes, Ashley

    2008-03-01

    We report on the response of lung cilia to applied forces. We have applied magnetic forces to magnetic beads attached to individual human lung cilia in cell cultures. Our magnetic system is capable of generating large forces (˜1nanoNewton on 1 micron beads) with a 3kHz bandwidth. We record the cilia beat motion using video microscopy to record beat frequency and amplitude as a function of applied force. We present three major findings. First, the stall force is approximately 150 pN. Second the frequency is unchanged by the application of forces up to the stall point. Third, the speed of the beat motion slows down according to the diminution of the beat amplitude while maintaining a constant frequency and the speed of the motion is the same whether the beat direction is in the same direction as the applied force or against the applied force.

  5. The roles of evolutionarily conserved functional modules in cilia-related trafficking

    PubMed Central

    Sung, Ching-Hwa; Leroux, Michel R.

    2014-01-01

    Cilia are present across most eukaryotic phyla and have diverse sensory and motility roles in animal physiology, cell signalling and development. Their biogenesis and maintenance depend on vesicular and intraciliary (intraflagellar) trafficking pathways that share conserved structural and functional modules. The functional units of the interconnected pathways, which include proteins involved in membrane coating as well as small GTPases and their accessory factors, were first experimentally associated with canonical vesicular trafficking. These components are, however, ancient, having been co-opted by the ancestral eukaryote to establish the ciliary organelle, and their study can inform us about ciliary biology in higher organisms. PMID:24296415

  6. 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.

  7. 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

  8. Structural defects in cilia of the choroid plexus, subfornical organ and ventricular ependyma are associated with ventriculomegaly

    PubMed Central

    2012-01-01

    Background Hydrocephalus is a heterogeneous disorder with multiple etiologies that are not yet fully understood. Animal models have implicated dysfunctional cilia of the ependyma and choroid plexus in the development of the disorder. In this report, we sought to determine the origin of the ventriculomegaly in four Bardet Biedl syndrome (BBS) mutant mouse strains as models of a ciliopathy. Methods Evans Blue dye was injected into the lateral ventricle of wild- type and BBS mutant mice to determine whether obstruction of intra- or extra-ventricular CSF flow contributed to ventriculomegaly. Transmission electron microscopy (TEM) was used to examine the ultrastructure of the choroid plexus, subfornical organ (SFO), subcommisural organ (SCO), and ventricular ependyma to evaluate their ultrastructure and the morphology of their primary and motile cilia. Results and discussion No obstruction of intra- or extra-ventricular CSF flow was observed, implying a communicating form of hydrocephalus in BBS mutant mice. TEM analyses of the mutants showed no evidence of choroidal papillomas or breakdown of the blood:CSF barrier. In contrast, structural defects were observed in a subpopulation of cilia lining the choroid plexus, SFO, and ventricular ependyma. These included disruptions of the microtubular structure of the axoneme and the presence of electron-dense vesicular-like material along the ciliary shaft and at the tips of cilia. Conclusions Abnormalities in cilia structure and function have the potential to influence ciliary intraflagellar transport (IFT), cilia maintenance, protein trafficking, and regulation of CSF production. Ciliary structural defects are the only consistent pathological features associated with CSF-related structures in BBS mutant mice. These defects are observed from an early age, and may contribute to the underlying pathophysiology of ventriculomegaly. PMID:23046663

  9. 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. PMID:23864713

  10. 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

  11. Bio-inspired artificial cilia with magnetic dynamic properties

    NASA Astrophysics Data System (ADS)

    Sun, Leilei; Zheng, Yongmei

    2015-04-01

    Inspired by the structure and properties of natural cilia, we focused on a facile template-free approach to prepare magnetic artificial cilia grown on the substrate (glass, PDMS, or others). In an applied magnetic field, the cilia formed spontaneously and immediately from magnetic nanoparticles and elastomeric polymer in a liquid solvent by bottom-up self-assembly. The length of prepared cilia could be in the scale of millimeter and reach a high aspect ratio of even over 100. We studied the effect of the magnetic strength applied and the size of nanoparticles to get tunable scale of cilia. The cilia show reversibly bending in an external magnetic field and this bending actuation gave some important functions: to transport macroscopic nonmagnetic materials on the cilia and to mix liquids.

  12. 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

  13. Human follicular fluid adverses hamster spermatozoa motility.

    PubMed

    Wetzels, A; Goverde, H J; Bastiaans, L A; Rolland, R

    1989-01-01

    To determine the optimal conditions for in vitro spermatozoa vitality, human and hamster spermatozoa were incubated at 37 degrees C in T6 medium supplemented with different biologic fluids (10% v/v). The fluids tested were human serum (HUS), hamster serum (HAS), and human follicular fluid (HUF). After incubation the spermatozoa were investigated for their qualitative and quantitative motility. Human spermatozoa maintained a good vitality in all fluids tested (approximately 25% motility after 18-h incubation). The hamster spermatozoa had after an incubation of 4 h a motility of 28.4% in HUS, 14.2% in HAS, and 2.2% in HUF. The quality of the motility was also extremely low in HUF, whereas it was adequate in HUS and in HAS. The presence of species-specific substances in mammalian follicular fluid is discussed. PMID:2589906

  14. 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. PMID:21331419

  15. Localization of a Guanylyl Cyclase to Chemosensory Cilia Requires the Novel Ciliary MYND Domain Protein DAF-25

    PubMed Central

    Jensen, Victor L.; Bialas, Nathan J.; Bishop-Hurley, Sharon L.; Molday, Laurie L.; Kida, Katarzyna; Nguyen, Phuong Anh T.; Blacque, Oliver E.; Molday, Robert S.; Leroux, Michel R.; Riddle, Donald L.

    2010-01-01

    In harsh conditions, Caenorhabditis elegans arrests development to enter a non-aging, resistant diapause state called the dauer larva. Olfactory sensation modulates the TGF-β and insulin signaling pathways to control this developmental decision. Four mutant alleles of daf-25 (abnormal DAuer Formation) were isolated from screens for mutants exhibiting constitutive dauer formation and found to be defective in olfaction. The daf-25 dauer phenotype is suppressed by daf-10/IFT122 mutations (which disrupt ciliogenesis), but not by daf-6/PTCHD3 mutations (which prevent environmental exposure of sensory cilia), implying that DAF-25 functions in the cilia themselves. daf-25 encodes the C. elegans ortholog of mammalian Ankmy2, a MYND domain protein of unknown function. Disruption of DAF-25, which localizes to sensory cilia, produces no apparent cilia structure anomalies, as determined by light and electron microscopy. Hinting at its potential function, the dauer phenotype, epistatic order, and expression profile of daf-25 are similar to daf-11, which encodes a cilium-localized guanylyl cyclase. Indeed, we demonstrate that DAF-25 is required for proper DAF-11 ciliary localization. Furthermore, the functional interaction is evolutionarily conserved, as mouse Ankmy2 interacts with guanylyl cyclase GC1 from ciliary photoreceptors. The interaction may be specific because daf-25 mutants have normally-localized OSM-9/TRPV4, TAX-4/CNGA1, CHE-2/IFT80, CHE-11/IFT140, CHE-13/IFT57, BBS-8, OSM-5/IFT88, and XBX-1/D2LIC in the cilia. Intraflagellar transport (IFT) (required to build cilia) is not defective in daf-25 mutants, although the ciliary localization of DAF-25 itself is influenced in che-11 mutants, which are defective in retrograde IFT. In summary, we have discovered a novel ciliary protein that plays an important role in cGMP signaling by localizing a guanylyl cyclase to the sensory organelle. PMID:21124868

  16. Autophagy and primary cilia: dual interplay.

    PubMed

    Pampliega, Olatz; Cuervo, Ana Maria

    2016-04-01

    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

  17. Drosophila sperm motility in the reproductive tract.

    PubMed

    Yang, Yong; Lu, Xiangyi

    2011-05-01

    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

  18. 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

  19. Methods for Studying Movement of Molecules Within Cilia.

    PubMed

    Lechtreck, Karl F

    2016-01-01

    The assembly of cilia and eukaryotic flagella (interchangeable terms) requires the import of numerous proteins from the cell body into the growing organelle. Proteins move into and inside cilia by diffusion and by motor-based intraflagellar transport (IFT). Many aspects of ciliary protein transport such as the distribution of unloading sites and the frequency of transport can be analyzed using direct in vivo imaging of fluorescently tagged proteins. Here, we will describe how to use total internal reflection fluorescence microcopy (TIRFM) to analyze protein transport in the flagella of the unicellular alga Chlamydomonas reinhardtii, a widely used model for cilia and cilia-related disease. PMID:27514917

  20. Rheotaxis guides mammalian sperm

    PubMed Central

    Miki, Kiyoshi; Clapham, David E

    2013-01-01

    Background In sea urchins, spermatozoan motility is altered by chemotactic peptides, giving rise to the assumption that mammalian eggs also emit chemotactic agents that guide spermatozoa through the female reproductive tract to the mature oocyte. Mammalian spermatozoa indeed undergo complex adaptations within the female (the process of capacitation) that are initiated by agents ranging from pH to progesterone, but these factors are not necessarily taxic. Currently, chemotaxis, thermotaxis, and rheotaxis have not been definitively established in mammals. Results Here, we show that positive rheotaxis, the ability of organisms to orient and swim against the flow of surrounding fluid, is a major taxic factor for mouse and human sperm. This flow is generated within 4 hours of sexual stimulation and coitus in female mice; prolactin-triggered oviductal fluid secretion clears the oviduct of debris, lowers viscosity, and generates the stream that guides sperm migration in the oviduct. Rheotaxic movement is demonstrated in capacitated and uncapacitated spermatozoa in low and high viscosity medium. Finally, we show that a unique sperm motion we quantify using the sperm head's rolling rate reflects sperm rotation that generates essential force for positioning the sperm in the stream. Rotation requires CatSper channels, presumably by enabling Ca2+ influx. Conclusions We propose that rheotaxis is a major determinant of sperm guidance over long distances in the mammalian female reproductive tract. Coitus induces fluid flow to guide sperm in the oviduct. Sperm rheotaxis requires rotational motion during CatSper channel-dependent hyperactivated motility. PMID:23453951

  1. 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

  2. Cilia and Nuclear Pore Proteins: Pore No More?

    PubMed

    Obado, Samson O; Rout, Michael P

    2016-09-12

    Nuclear pore proteins at the base of cilia were thought to regulate transport into cilia. In this issue of Developmental Cell, Del Viso et al. (2016) challenge this view, showing instead that pore proteins localize to ciliary basal bodies and that their perturbation leads to congenital heart disease. PMID:27623377

  3. ATR promotes cilia signalling: links to developmental impacts.

    PubMed

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

    2016-04-15

    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

  4. 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

  5. Visualization of live primary cilia dynamics using fluorescence microscopy

    PubMed Central

    Ott, Carolyn; Lippincott-Schwartz, Jennifer

    2013-01-01

    Here we describe methods that are useful for exploring the formation and functions of primary cilia in living cells. First we describe multiple protocols for visualizing solitary cilia that extend away from the cell body. Primary cilia collect, synthesize, and transmit information about the extracellular space into the cell body to promote critical cellular responses. Problems with cilia formation or function can lead to dramatic changes in cell physiology. These methods can be used to assess cilia formation and length, the location of the cilium relative to other cellular structures, and localization of specific proteins to the cilium. The second protocol describes how to quantify movement of fluorescent molecules within the cilium. The microtubules that form the structural scaffold of the cilium are also critical avenues for kinesin and dynein-mediated movement of proteins within the cilium. Assessing intraflagellar dynamics can provide insight into mechanisms of ciliary-mediated signal perception and transmission. PMID:23208547

  6. 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

  7. Computation of the internal forces in cilia: application to ciliary motion, the effects of viscosity, and cilia interactions.

    PubMed

    Gueron, S; Levit-Gurevich, K

    1998-04-01

    This paper presents a simple and reasonable method for generating a phenomenological model of the internal mechanism of cilia. The model uses a relatively small number of parameters whose values can be obtained by fitting to ciliary beat shapes. Here, we use beat patterns observed in Paramecium. The forces that generate these beats are computed and fit to a simple functional form called the "engine." This engine is incorporated into a recently developed hydrodynamic model that accounts for interactions between neighboring cilia and between the cilia and the surface from which they emerge. The model results are compared to data on ciliary beat patterns of Paramecium obtained under conditions where the beats are two-dimensional. Many essential features of the motion, including several properties that are not built in explicitly, are shown to be captured. In particular, the model displays a realistic change in beat pattern and frequency in response to increased viscosity and to the presence of neighboring cilia in configurations such as rows of cilia and two-dimensional arrays of cilia. We found that when two adjacent model cilia start beating at different phases they become synchronized within several beat periods, as observed in experiments where two flagella are brought into close proximity. Furthermore, examination of various multiciliary configurations shows that an approximately antiplectic wave pattern evolves autonomously. This modeling evidence supports earlier conjectures that metachronism may occur, at least partially, as a self-organized phenomenon due to hydrodynamic interactions between neighboring cilia. PMID:9545031

  8. The effect of cilia and the mucociliary clearance on successful drug delivery.

    PubMed

    Gizurarson, Sveinbjörn

    2015-01-01

    Nasal mucociliary clearance is one of the most important factors affecting nasal delivery of drugs and vaccines. This is also the most important physiological defense mechanism inside the nasal cavity. It removes inhaled (and delivered) particles, microbes and substances trapped in the mucus. Almost all inhaled particles are trapped in the mucus carpet and transported with a rate of 8-10 mm/h toward the pharynx. This transport is conducted by the ciliated cells, which contain about 100-250 motile cellular appendages called cilia, 0.3 µm wide and 5 µm in length that beat about 1000 times every minute or 12-15 Hz. For efficient mucociliary clearance, the interaction between the cilia and the nasal mucus needs to be well structured, where the mucus layer is a tri-layer: an upper gel layer that floats on the lower, more aqueous solution, called the periciliary liquid layer and a third layer of surfactants between these two main layers. Pharmacokinetic calculations of the mucociliary clearance show that this mechanism may account for a substantial difference in bioavailability following nasal delivery. If the formulation irritates the nasal mucosa, this mechanism will cause the irritant to be rapidly diluted, followed by increased clearance, and swallowed. The result is a much shorter duration inside the nasal cavity and therefore less nasal bioavailability. PMID:25739664

  9. Basal foot MTOC organizes pillar MTs required for coordination of beating cilia

    PubMed Central

    Clare, Daniel K.; Magescas, Jérémy; Piolot, Tristan; Dumoux, Maud; Vesque, Christine; Pichard, Evelyne; Dang, Tien; Duvauchelle, Boris; Poirier, Françoise; Delacour, Delphine

    2016-01-01

    Coordination of ciliary beating is essential to ensure mucus clearance in the airway tract. The orientation and synchronization of ciliary motion responds in part to the organization of the underlying cytoskeletal networks. Using electron tomography on mouse trachea, we show that basal bodies are collectively hooked at the cortex by a regular microtubule array composed of 4-5 microtubules. Removal of Galectin-3, one of basal body components, provokes misrecruitment of γ-tubulin, disorganization of this microtubule framework emanating from the basal foot cap, together with loss of basal body alignment and cilium orientation, defects in cilium organization and reduced fluid flow in the tracheal lumen. We conclude that Galectin-3 plays a crucial role in the maintenance of the microtubule organizing center of the cilium and the “pillar” microtubules, and that this network is instrumental for the coordinated orientation and stabilization of motile cilia. PMID:25215410

  10. Planaria as a Model System for the Analysis of Ciliary Assembly and Motility.

    PubMed

    King, Stephen M; Patel-King, Ramila S

    2016-01-01

    Planarian flatworms are carnivorous invertebrates with astounding regenerative properties. They have a ventral surface on which thousands of motile cilia are exposed to the extracellular environment. These beat in a synchronized manner against secreted mucus thereby propelling the animal forward. Similar to the nematode Caenorhabditis elegans, the planarian Schmidtea mediterranea is easy to maintain in the laboratory and is highly amenable to simple RNAi approaches through feeding with dsRNA. The methods are simple and robust, and the level of gene expression reduction that can be obtained is, in many cases, almost total. Moreover, cilia assembly and function is not essential for viability in this organism, as animals readily survive for weeks even with the apparent total absence of this organelle. Both genome and expressed sequence tag databases are available and allow design of vectors to target any desired gene of choice. Combined, these feature make planaria a useful model system in which to examine ciliary assembly and motility, especially in the context of a ciliated epithelium where many organelles beat in a hydrodynamically coupled synchronized manner. In addition, as planaria secrete mucus against which the cilia beat to generate propulsive force, this system may also prove useful for analysis of mucociliary interactions. In this chapter, we provide simple methods to maintain a planarian colony, knockdown gene expression by RNAi, and analyze the resulting animals for whole organism motility as well as ciliary architecture and function. PMID:27514927

  11. Biomimetic cilia arrays generate simultaneous pumping and mixing regimes

    PubMed Central

    Shields, A. R.; Fiser, B. L.; Evans, B. A.; Falvo, M. R.; Washburn, S.; Superfine, R.

    2010-01-01

    Living systems employ cilia to control and to sense the flow of fluids for many purposes, such as pumping, locomotion, feeding, and tissue morphogenesis. Beyond their use in biology, functional arrays of artificial cilia have been envisaged as a potential biomimetic strategy for inducing fluid flow and mixing in lab-on-a-chip devices. Here we report on fluid transport produced by magnetically actuated arrays of biomimetic cilia whose size approaches that of their biological counterparts, a scale at which advection and diffusion compete to determine mass transport. Our biomimetic cilia recreate the beat shape of embryonic nodal cilia, simultaneously generating two sharply segregated regimes of fluid flow: Above the cilia tips their motion causes directed, long-range fluid transport, whereas below the tips we show that the cilia beat generates an enhanced diffusivity capable of producing increased mixing rates. These two distinct types of flow occur simultaneously and are separated in space by less than 5 μm, approximately 20% of the biomimetic cilium length. While this suggests that our system may have applications as a versatile microfluidics device, we also focus on the biological implications of our findings. Our statistical analysis of particle transport identifying an enhanced diffusion regime provides novel evidence for the existence of mixing in ciliated systems, and we demonstrate that the directed transport regime is Poiseuille–Couette flow, the first analytical model consistent with biological measurements of fluid flow in the embryonic node. PMID:20798342

  12. 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

  13. In vivo investigation of cilia structure and function using Xenopus

    PubMed Central

    Brooks, Eric R.; Wallingford, John B.

    2015-01-01

    Cilia are key organelles in development and homeostasis. The ever-expanding complement of cilia associated proteins necessitates rapid and tractable models for in vivo functional investigation. Xenopus laevis provides an attractive model for such studies, having multiple ciliated populations, including primary and multiciliated tissues. The rapid external development of Xenopus and the large cells make it an especially excellent platform for imaging studies. Here we present embryological and cell-biological methods for the investigation of cilia structure and function in Xenopus laevis, with a focus on quantitative live and fixed imaging. PMID:25837389

  14. Serotonin and colonic motility.

    PubMed

    Kendig, D M; Grider, J R

    2015-07-01

    The role of serotonin (5-hydroxytryptamine [5-HT]) in gastrointestinal motility has been studied for over 50 years. Most of the 5-HT in the body resides in the gut wall, where it is located in subsets of mucosal cells (enterochromaffin cells) and neurons (descending interneurons). Many studies suggest that 5-HT is important to normal and dysfunctional gut motility and drugs affecting 5-HT receptors, especially 5-HT3 and 5-HT4 receptors, have been used clinically to treat motility disorders; however, cardiovascular side effects have limited the use of these drugs. Recently studies have questioned the importance and necessity of 5-HT in general and mucosal 5-HT in particular for colonic motility. Recent evidence suggests the importance of 5-HT3 and 5-HT4 receptors for initiation and generation of one of the key colonic motility patterns, the colonic migrating motor complex (CMMC), in rat. The findings suggest that 5-HT3 and 5-HT4 receptors are differentially involved in two different types of rat CMMCs: the long distance contraction (LDC) and the rhythmic propulsive motor complex (RPMC). The understanding of the role of serotonin in colonic motility has been influenced by the specific motility pattern(s) studied, the stimulus used to initiate the motility (spontaneous vs induced), and the route of administration of drugs. All of these considerations contribute to the understanding and the controversy that continues to surround the role of serotonin in the gut. PMID:26095115

  15. 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. PMID:27514923

  16. Caenorhabditis elegans DYF-2, an orthologue of human WDR19, is a component of the intraflagellar transport machinery in sensory cilia.

    PubMed

    Efimenko, Evgeni; Blacque, Oliver E; Ou, Guangshuo; Haycraft, Courtney J; Yoder, Bradley K; Scholey, Jonathan M; Leroux, Michel R; Swoboda, Peter

    2006-11-01

    The intraflagellar transport (IFT) machinery required to build functional cilia consists of a multisubunit complex whose molecular composition, organization, and function are poorly understood. Here, we describe a novel tryptophan-aspartic acid (WD) repeat (WDR) containing IFT protein from Caenorhabditis elegans, DYF-2, that plays a critical role in maintaining the structural and functional integrity of the IFT machinery. We determined the identity of the dyf-2 gene by transgenic rescue of mutant phenotypes and by sequencing of mutant alleles. Loss of DYF-2 function selectively affects the assembly and motility of different IFT components and leads to defects in cilia structure and chemosensation in the nematode. Based on these observations, and the analysis of DYF-2 movement in a Bardet-Biedl syndrome mutant with partially disrupted IFT particles, we conclude that DYF-2 can associate with IFT particle complex B. At the same time, mutations in dyf-2 can interfere with the function of complex A components, suggesting an important role of this protein in the assembly of the IFT particle as a whole. Importantly, the mouse orthologue of DYF-2, WDR19, also localizes to cilia, pointing to an important evolutionarily conserved role for this WDR protein in cilia development and function. PMID:16957054

  17. The significance of ultrastructural abnormalities of human cilia.

    PubMed

    Fox, B; Bull, T B; Makey, A R; Rawbone, R

    1981-12-01

    The electronmicroscopic structure of cilia was studied from the inferior turbinate of the nose in 22 adults, and in 84 biopsies from the bronchial tree of 40 adults. The incidence of compound cilia and abnormal microtubular structures was assessed. There were significant variations in the incidence of abnormalities in different parts of the airways and even within different areas of the same electronmicroscopic section. The focal nature of differences in structure of cilia indicate that abnormalities found in a single biopsy do not necessarily reflect a generalized change in the bronchial tree. Thus, such a finding should not be used as evidence that the abnormalities of cilia are the cause of decrease in mucociliary clearance or that they play a role in the pathogenesis of bronchiectasis and sinusitis. PMID:7307613

  18. 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.

  19. 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

  20. Changes in cell surface primary cilia and microvilli concurrent with measurements of fluid flow across the rabbit corneal endothelium ex vivo.

    PubMed

    Doughty, M J

    1998-12-01

    Primary cilia and microvilli have been reported on the mammalian rabbit corneal endothelium but their relationship to cell function is undefined. Six corneas from healthy 2 kg female albino rabbits were glutaraldehyde-fixed post mortem (15:00 h) or twelve corneal stroma-endothelial preparations incubated at 37 degrees C under an applied hydrostatic pressure of 20 cm H2O for 4 h prior to fixation. The corneal endothelium was assessed by quantitative scanning electron microscopy. Cells fixed immediately post mortem were decorated with small stubby microvilli (average 21 +/- 13/100 micron 2), and only 25% of the cells were decorated with primary cilia having an average length of 2.44 +/- 1.56 microns. Following 4 h ex vivo incubation with a phosphate-buffered Ringer solution, conspicuous microvilli developed to an average density of 40 +/- 19/100 micron 2 and primary cilia were found on 12% of the cells, having on average length of 2.27 +/- 1.38 microns. Following 4 h incubation in a bicarbonate-buffered Ringer solution, small stubby microvilli developed to a density of 49 +/- 18/100 micron 2, and 40% of the cells showed primary cilia with an average length of 4.31 +/- 1.93 microns; the net trans-endothelial fluid flow in the latter set was 60% greater. These studies indicate that the primary cilia on corneal endothelial cells might be responsive to fluid flow, but that mild mechanical and/or chemical stress could also be the cause of the change since the elaboration of primary cilia can be accompanied by microvilli as well. PMID:10036788

  1. Endothelial Cilia Are Essential for Developmental Vascular Integrity in Zebrafish

    PubMed Central

    Kallakuri, Sowjanya; Yu, Jianxin A.; Li, Jade; Li, Yuanyuan; Weinstein, Brant M.; Nicoli, Stefania

    2015-01-01

    The cilium is a signaling platform of the vertebrate cell. It has a critical role in polycystic kidney disease and nephronophthisis. Cilia have been detected on endothelial cells, but the function of these organelles in the vasculature remains incompletely defined. In this study, using genetic and chemical genetic tools in the model organism zebrafish, we reveal an essential role of cilia in developmental vascular integrity. Embryos expressing mutant intraflagellar transport genes, which are essential and specific for cilia biogenesis, displayed increased risk of developmental intracranial hemorrhage, whereas the morphology of the vasculature remained normal. Moreover, cilia were present on endothelial cells in the developing zebrafish vasculature. We further show that the involvement of cilia in vascular integrity is endothelial autonomous, because endothelial-specific re-expression of intraflagellar transport genes in respective mutants rescued the intracranial hemorrhage phenotype. Finally, whereas inhibition of Hedgehog signaling increased the risk of intracranial hemorrhage in ciliary mutants, activation of the pathway rescued this phenotype. In contrast, embryos expressing an inactivating mutation in pkd2, one of two autosomal dominant cystic kidney disease genes, did not show increased risk of developmental intracranial hemorrhage. These results suggest that Hedgehog signaling is a major mechanism for this novel role of endothelial cilia in establishing vascular integrity. PMID:25214579

  2. Pneumatically-actuated artificial cilia array for biomimetic fluid propulsion.

    PubMed

    Gorissen, Benjamin; de Volder, Michaël; Reynaerts, Dominiek

    2015-11-21

    Arrays of beating cilia emerged in nature as one of the most efficient propulsion mechanisms at a small scale, and are omnipresent in microorganisms. Previous attempts at mimicking these systems have foundered against the complexity of fabricating small-scale cilia exhibiting complex beating motions. In this paper, we propose for the first time arrays of pneumatically-actuated artificial cilia that are able to address some of these issues. These artificial cilia arrays consist of six highly flexible silicone rubber actuators with a diameter of 1 mm and a length of 8 mm that can be actuated independently from each other. In an experimental setup, the effects of the driving frequency, phase difference and duty cycle on the net flow in a closed-loop channel have been studied. Net fluid speeds of up to 19 mm s(-1) have been measured. Further, it is possible to invert the flow direction by simply changing the driving frequency or by changing the duty cycle of the driving block pulse pressure wave without changing the bending direction of the cilia. Using PIV measurements, we corroborate for the first time existing mathematical models of cilia arrays to measurements on prototypes. PMID:26439855

  3. Asynchronous beating of cilia enhances particle capture rate

    NASA Astrophysics Data System (ADS)

    Ding, Yang; Kanso, Eva

    2014-11-01

    Many aquatic micro-organisms use beating cilia to generate feeding currents and capture particles in surrounding fluids. One of the capture strategies is to ``catch up'' with particles when a cilium is beating towards the overall flow direction (effective stroke) and intercept particles on the downstream side of the cilium. Here, we developed a 3D computational model of a cilia band with prescribed motion in a viscous fluid and calculated the trajectories of the particles with different sizes in the fluid. We found an optimal particle diameter that maximizes the capture rate. The flow field and particle motion indicate that the low capture rate of smaller particles is due to the laminar flow in the neighbor of the cilia, whereas larger particles have to move above the cilia tips to get advected downstream which decreases their capture rate. We then analyzed the effect of beating coordination between neighboring cilia on the capture rate. Interestingly, we found that asynchrony of the beating of the cilia can enhance the relative motion between a cilium and the particles near it and hence increase the capture rate.

  4. 9 + 0 and 9 + 2 cilia are randomly dispersed in the mouse node.

    PubMed

    Odate, Toru; Takeda, Sen; Narita, Keishi; Kawahara, Toru

    2016-04-01

    The initial determination of left-right asymmetry is an essential process in embryonic development. In mouse embryo, cilia in the node play an important role generating the nodal flow that subsequently triggers left-right determination in the embryo. Although nodal cilia have historically been thought to have a 9 + 0 axonemal configuration, the existence of 9 + 2 cilia has been reported so far. Because the distribution of those two types of cilia within the node has not yet been reported, we assessed the arrangement of 9 + 0 and 9 + 2 cilia in the node. In this study, we concluded that most of the nodal cilia were 9 + 0 in structure and there were much fewer 9 + 2 cilia than 9 + 0 cilia. Furthermore, the two types of cilia were randomly distributed in the node with no regularity. In addition, we studied the embryonic origin of the crown cells surrounding the node to better understand their identity. PMID:26520785

  5. Pediatric intestinal motility disorders

    PubMed Central

    Gfroerer, Stefan; Rolle, Udo

    2015-01-01

    Pediatric intestinal motility disorders affect many children and thus not only impose a significant impact on pediatric health care in general but also on the quality of life of the affected patient. Furthermore, some of these conditions might also have implications for adulthood. Pediatric intestinal motility disorders frequently present as chronic constipation in toddler age children. Most of these conditions are functional, meaning that constipation does not have an organic etiology, but in 5% of the cases, an underlying, clearly organic disorder can be identified. Patients with organic causes for intestinal motility disorders usually present in early infancy or even right after birth. The most striking clinical feature of children with severe intestinal motility disorders is the delayed passage of meconium in the newborn period. This sign is highly indicative of the presence of Hirschsprung disease (HD), which is the most frequent congenital disorder of intestinal motility. HD is a rare but important congenital disease and the most significant entity of pediatric intestinal motility disorders. The etiology and pathogenesis of HD have been extensively studied over the last several decades. A defect in neural crest derived cell migration has been proven as an underlying cause of HD, leading to an aganglionic distal end of the gut. Numerous basic science and clinical research related studies have been conducted to better diagnose and treat HD. Resection of the aganglionic bowel remains the gold standard for treatment of HD. Most recent studies show, at least experimentally, the possibility of a stem cell based therapy for HD. This editorial also includes rare causes of pediatric intestinal motility disorders such as hypoganglionosis, dysganglionosis, chronic intestinal pseudo-obstruction and ganglioneuromatosis in multiple endocrine metaplasia. Underlying organic pathologies are rare in pediatric intestinal motility disorders but must be recognized as early as

  6. Pediatric intestinal motility disorders.

    PubMed

    Gfroerer, Stefan; Rolle, Udo

    2015-09-01

    Pediatric intestinal motility disorders affect many children and thus not only impose a significant impact on pediatric health care in general but also on the quality of life of the affected patient. Furthermore, some of these conditions might also have implications for adulthood. Pediatric intestinal motility disorders frequently present as chronic constipation in toddler age children. Most of these conditions are functional, meaning that constipation does not have an organic etiology, but in 5% of the cases, an underlying, clearly organic disorder can be identified. Patients with organic causes for intestinal motility disorders usually present in early infancy or even right after birth. The most striking clinical feature of children with severe intestinal motility disorders is the delayed passage of meconium in the newborn period. This sign is highly indicative of the presence of Hirschsprung disease (HD), which is the most frequent congenital disorder of intestinal motility. HD is a rare but important congenital disease and the most significant entity of pediatric intestinal motility disorders. The etiology and pathogenesis of HD have been extensively studied over the last several decades. A defect in neural crest derived cell migration has been proven as an underlying cause of HD, leading to an aganglionic distal end of the gut. Numerous basic science and clinical research related studies have been conducted to better diagnose and treat HD. Resection of the aganglionic bowel remains the gold standard for treatment of HD. Most recent studies show, at least experimentally, the possibility of a stem cell based therapy for HD. This editorial also includes rare causes of pediatric intestinal motility disorders such as hypoganglionosis, dysganglionosis, chronic intestinal pseudo-obstruction and ganglioneuromatosis in multiple endocrine metaplasia. Underlying organic pathologies are rare in pediatric intestinal motility disorders but must be recognized as early as

  7. Oscillations of Eukaryotic Cilia and Flagella

    NASA Astrophysics Data System (ADS)

    Gopinath, Arvind; Mahadevan, Lakshminarayanan

    2006-11-01

    The undulating beat of eukaryotic flagella and cilia produces forces that move cells and cause locomotion. The timing mechanisms that generate these periodic undulations are still mysterious and the question of how these oscillations arise is still a subject of much research - both experimental and theoretical. Recent experimental results on paralyzed and reconstituted flagella offer new insight into the dynamical mechanisms that could result in sustained waveform generation. Motivated by these recent experimental results we propose a model that mimics the flagellar structure as motor driven elastic, inextensible filaments. We hypothesize that the oscillations arise due to motor (dynein) driven, constrained, relative sliding of parts of the flagella. The dynamical equations describing the evolution of the populations of attached and detached motors is actively coupled to the local configuration as well as local sliding velocities via strain and configuration dependent kinetic reaction rates. At the same time, the filament configuration is actively coupled to the motor densities via the dependence of the active internal torque densities on the motor populations as well as their internal state. Appropriate ensemble averaged force-velocity relationships for the motors completes the set of equations. Numerical solutions reveal onset of dynamical instabilities via Hopf-bifurcations with oscillatory waveforms emerging from a trivial base state corresponding to a straight, non-moving flagellum.

  8. Caught Nek-ing: cilia and centrioles.

    PubMed

    Quarmby, Lynne M; Mahjoub, Moe R

    2005-11-15

    The Nek family of cell-cycle kinases is widely represented in eukaryotes and includes numerous proteins that were described only recently and remain poorly characterized. Comparing Neks in the context of clades allows us to examine the question of whether microbial eukaryotic Neks, although not strictly orthologs of their vertebrate counterparts, can provide clues to ancestral functions that might be retained in the vertebrate Neks. Relatives of the Nek2/NIMA proteins play important roles at the G2-M transition in nuclear envelope breakdown and centromere separation. Nek6, Nek7 and Nek9 also seem to regulate mitosis. By contrast, Nek1 and Nek8 have been linked with polycystic kidney disease. Results of statistical analysis indicate that the family coevolved with centrioles that function as both microtubule-organizing centers and the basal bodies of cilia. This evolutionary perspective, taken together with functional studies of microbial Neks, provides new insights into the cellular roles of the proteins and disease with which some of them have been linked. PMID:16280549

  9. 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. PMID:21614349

  10. The role of QseC quorum-sensing sensor kinase in colonization and norepinephrine-enhanced motility of Salmonella enterica serovar Typhirmurium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transcriptional analysis of Salmonella enterica serovar Typhimurium in the presence of the mammalian hormone Norepinephrine (NE) revealed up-regulation of chemotaxis and motility genes. Motility assays confirmed enhanced motility of wild-type S. Typhimurium in the presence of NE that could be block...

  11. 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. PMID:26045730

  12. 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

  13. Cell motility on nanotopography

    NASA Astrophysics Data System (ADS)

    Kimura, Masahiro; Tsai, Irene; Green, Angelo; Jacobson, Bruce; Russell, Thomas

    2003-03-01

    Cell motility is strongly influenced by the structure of the substratum. Understanding cells motility on a surface has significant applications both in vivo and in vitro applications, such as biological sensors and hip replacement. A gradient surface is used to study the effect of the lateral nanotopography on cell motility. A gradient surface is generated by block copolymer and homopolymer blends, where the concentration of the components varies uniformly across the surface. The two homopolymers phase separate on the micron scale and this length scale gradually decrease to the nanoscopic, i.e. microphase separation of the diblock, as the copolymer concentration increases. Quantitative analysis of the speed of cell migration is correlated to the lateral length scale of the surface.

  14. [Obesity and gastrointestinal motility].

    PubMed

    Lee, Joon Seong

    2006-08-01

    Gastrointestinal (GI) motility has a crucial role in the food consumption, digestion and absorption, and also controls the appetite and satiety. In obese patients, various alterations of GI motility have been investigated. The prevalence of GERD and esophageal motor disorders in obese patients are higher than those of general population. Gastric emptying of solid food is generally accelerated and fasting gastric volume especially in distal stomach is larger in obese patients without change in accommodation. Contractile activity of small intestine in fasting period is more prominent, but orocecal transit is delayed. Autonomic dysfunction is frequently demonstrated in obese patients. These findings correspond with increased appetite and delayed satiety in obese patients, but causes or results have not been confirmed. Therapeutic interventions of these altered GI motility have been developed using botulinum toxin, gastric electrical stimulation in obese patients. Novel agents targeted for GI hormone modulation (such as ghrelin and leptin) need to be developed in the near future. PMID:16929152

  15. 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

  16. Effectiveness of Hair Bundle Motility as the Cochlear Amplifier

    PubMed Central

    Sul, Bora; Iwasa, Kuni H.

    2009-01-01

    Abstract The effectiveness of hair bundle motility in mammalian and avian ears is studied by examining energy balance for a small sinusoidal displacement of the hair bundle. The condition that the energy generated by a hair bundle must be greater than energy loss due to the shear in the subtectorial gap per hair bundle leads to a limiting frequency that can be supported by hair-bundle motility. Limiting frequencies are obtained for two motile mechanisms for fast adaptation, the channel re-closure model and a model that assumes that fast adaptation is an interplay between gating of the channel and the myosin motor. The limiting frequency obtained for each of these models is an increasing function of a factor that is determined by the morphology of hair bundles and the cochlea. Primarily due to the higher density of hair cells in the avian inner ear, this factor is ∼10-fold greater for the avian ear than the mammalian ear, which has much higher auditory frequency limit. This result is consistent with a much greater significance of hair bundle motility in the avian ear than that in the mammalian ear. PMID:19917218

  17. 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

  18. 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.

  19. 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.

  20. Ins and outs of GPCR signaling in primary cilia.

    PubMed

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

    2015-09-01

    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

  1. Primary Cilia Regulate Branching Morphogenesis During Mammary Gland Development

    PubMed Central

    McDermott, Kimberly M.; Liu, Bob Y.; Tlsty, Thea D.; Pazour, Gregory J.

    2010-01-01

    Summary During mammary gland development an epithelial bud undergoes branching morphogenesis to expand into a continuous tree-like network of branched ducts [1]. The process involves multiple cell types that are coordinated by hormones and growth factors coupled with signaling events including Wnt and Hedgehog [2-5]. Primary cilia play key roles in the development of many organs by coordinating extracellular signaling (Wnt, Hedgehog) with cellular physiology [6-8]. During mammary development, we find cilia on luminal epithelial, myoepithelial and stromal cells during early branching morphogenesis when epithelial ducts extend into the fat pad and undergo branching morphogenesis. When branching is complete, cilia disappear from luminal epithelial cells but remain on myoepithelial and stromal cells. Ciliary dysfunction caused by intraflagellar transport (IFT) defects results in branching defects. These include decreased ductal extension and decreased secondary and tertiary branching along with reduced lobular-alveolar development during pregnancy and lactation. We find increased canonical Wnt and decreased Hedgehog signaling in the mutant glands, which is consistent with the role of cilia in regulating these pathways [6-11]. In mammary gland and other organs, increased canonical Wnt [12-14] and decreased Hedgehog [15, 16] signaling decreases branching morphogenesis suggesting that Wnt and Hedgehog signaling connect ciliary dysfunction to branching defects. PMID:20381354

  2. Primary cilia and Gli3 activity regulate cerebral cortical size

    PubMed Central

    Wilson, Sandra L.; Wilson, John P.; Wang, Chengbing; Wang, Baolin; McConnell, Susan K.

    2012-01-01

    During neural development, patterning, neurogenesis and overall growth are highly regulated and coordinated between different brain regions. Here, we show that primary cilia and the regulation of Gli activity, are necessary for the normal expansion of the cerebral cortex. We show that loss of Kif3a, an important functional component of primary cilia, leads to the degeneration of primary cilia, marked overgrowth of the cortex, and altered cell cycle kinetics within cortical progenitors. The G1 phase of the cell cycle is shortened through a mechanism likely involving reduced Gli3 activity and a resulting increase in expression of cyclin D1 and Fgf15. The defects in Gli3 activity alone are sufficient to accelerate cell cycle kinetics and cause the molecular changes seen in brains that lack cilia. Finally, we show that levels of full-length and repressor Gli3 proteins are tightly regulated during normal development and correlate with changes in expression of two known Shh-target genes, CyclinD1 and Fgf15, and with the normal lengthening of the cell cycle during corticogenesis. These data suggest that Gli3 activity is regulated through the primary cilium to control cell cycle length in the cortex and thus determine cortical size. PMID:21976438

  3. 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. PMID:23685964

  4. 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

  5. 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

  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. 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. PMID:23988175

  9. 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

  10. An ocular motility conundrum.

    PubMed

    McElnea, Elizabeth Margaret; Stephenson, Kirk; Lanigan, Bernie; Flitcroft, Ian

    2014-01-01

    Two siblings, an 11-year-old boy and a 7-year-old girl presented with bilateral symmetrical ptosis and limited eye movements. Having already been reviewed on a number of occasions by a variety of specialists in multiple hospital settings a diagnosis of their ocular motility disorder had remained elusive. We describe their cases, outline the differential diagnosis and review the investigations performed which were influential in finally making a diagnosis. PMID:25349186

  11. 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

  12. The Ciliary Rootlet Maintains Long-Term Stability of Sensory Cilia

    PubMed Central

    Yang, Jun; Gao, Jiangang; Adamian, Michael; Wen, Xiao-Hong; Pawlyk, Basil; Zhang, Luo; Sanderson, Michael J.; Zuo, Jian; Makino, Clint L.; Li, Tiansen

    2005-01-01

    The striated ciliary rootlet is a prominent cytoskeleton originating from basal bodies of ciliated cells. Although a familiar structure in cell biology, its function has remained unresolved. In this study, we carried out targeted disruption in mice of the gene for rootletin, a component of the rootlet. In the mutant, ciliated cells are devoid of rootlets. Phototransduction and ciliary beating in sensory and motile cilia initially exhibit no apparent functional deficits. However, photoreceptors degenerate over time, and mutant lungs appear prone to pathological changes consistent with insufficient mucociliary clearance. Further analyses revealed a striking fragility at the ciliary base in photoreceptors lacking rootlets. In vitro assays suggest that the rootlet is among the least dynamic of all cytoskeletons and interacts with actin filaments. Thus, a primary function of the rootlet is to provide structural support for the cilium. Inasmuch as photoreceptors elaborate an exceptionally enlarged sensory cilium, they are especially dependent on the rootlet for structural integrity and long-term survival. PMID:15870283

  13. 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. PMID:22935613

  14. 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

  15. Cilia-based flow network in the brain ventricles.

    PubMed

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

    2016-07-01

    Cerebrospinal fluid conveys many physiologically important signaling factors through the ventricular cavities of the brain. We investigated the transport of cerebrospinal fluid in the third ventricle of the mouse brain and discovered a highly organized pattern of cilia modules, which collectively give rise to a network of fluid flows that allows for precise transport within this ventricle. We also discovered a cilia-based switch that reliably and periodically alters the flow pattern so as to create a dynamic subdivision that may control substance distribution in the third ventricle. Complex flow patterns were also present in the third ventricles of rats and pigs. Our work suggests that ciliated epithelia can generate and maintain complex, spatiotemporally regulated flow networks. PMID:27387952

  16. Genetic Ablation of Type III Adenylyl Cyclase Exerts Region-Specific Effects on Cilia Architecture in the Mouse Nose

    PubMed Central

    Challis, Rosemary C.; Tian, Huikai; Yin, Wenbin; Ma, Minghong

    2016-01-01

    We recently reported that olfactory sensory neurons in the dorsal zone of the mouse olfactory epithelium exhibit drastic location-dependent differences in cilia length. Furthermore, genetic ablation of type III adenylyl cyclase (ACIII), a key olfactory signaling protein and ubiquitous marker for primary cilia, disrupts the cilia length pattern and results in considerably shorter cilia, independent of odor-induced activity. Given the significant impact of ACIII on cilia length in the dorsal zone, we sought to further investigate the relationship between cilia length and ACIII level in various regions throughout the mouse olfactory epithelium. We employed whole-mount immunohistochemical staining to examine olfactory cilia morphology in phosphodiesterase (PDE) 1C-/-;PDE4A-/- (simplified as PDEs-/- hereafter) and ACIII-/- mice in which ACIII levels are reduced and ablated, respectively. As expected, PDEs-/- animals exhibit dramatically shorter cilia in the dorsal zone (i.e., where the cilia pattern is found), similar to our previous observation in ACIII-/- mice. Remarkably, in a region not included in our previous study, ACIII-/- animals (but not PDEs-/- mice) have dramatically elongated, comet-shaped cilia, as opposed to characteristic star-shaped olfactory cilia. Here, we reveal that genetic ablation of ACIII has drastic, location-dependent effects on cilia architecture in the mouse nose. These results add a new dimension to our current understanding of olfactory cilia structure and regional organization of the olfactory epithelium. Together, these findings have significant implications for both cilia and sensory biology. PMID:26942602

  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. IFT46 plays crucial roles in craniofacial and cilia development.

    PubMed

    Park, Inji; Lee, Hyun-Kyung; Kim, Chowon; Ismail, Tayaba; Kim, Yoo-Kyung; Park, Jeen-Woo; Kwon, Oh-Shin; Kang, Beom Sik; Lee, Dong-Seok; Park, Tae-Joo; Park, Mae-Ja; Choi, Sun-Cheol; Lee, Hyun-Shik

    2016-08-26

    The intraflagellar transport (IFT) system is essential for bidirectional movement of ciliary components from the basal body to the tip beneath the ciliary sheath and is conserved for cilia and flagella formation in most vertebrates. IFT complex A is involved in anterograde trafficking, whereas complex B is involved in retrograde trafficking. IFT46 is well known as a crucial component of IFT complex B, however, its developmental functions are poorly understood. In this study, we investigated the novel functions of IFT46 during vertebrate development, especially, ciliogenesis and neurogenesis, because IFT46 is strongly expressed in both multiciliated cells of epithelial and neural tissues. Knockdown of IFT46 using morpholino microinjections caused shortening of the body axis as well as the formation of fewer and shorter cilia. Furthermore, loss of IFT46 down-regulated the expression of the neural plate and neural tube markers, thus may influence Wnt/planar cell polarity and the sonic hedgehog signaling pathway during neurogenesis. In addition, loss of IFT46 caused craniofacial defects by interfering with cartilage formation. In conclusion, our results depict that IFT46 plays important roles in cilia as well as in neural and craniofacial development. PMID:27320864

  19. 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.

  20. 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

  1. 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.

  2. Cyclic AMP induces maturation of trout sperm axoneme to initiate motility

    NASA Astrophysics Data System (ADS)

    Morisawa, Masaaki

    1982-02-01

    Cyclic AMP has long been implicated as an activator of sperm motility1-5. From more recent experiments using demembranated mammalian and sea urchin spermatozoa6,7, it was concluded that cyclic AMP only increases the motility of the axoneme after it has been initiated by MgATP2-. We have now carried out similar experiments using spermatozoa collected from the rainbow trout and demembranated by treatment with the detergent Triton X-100. Our results suggest that in this species, cyclic AMP is required before MgATP2- to trigger maturation of the nonmotile axoneme. Subsequent addition of an energy source then induces motility.

  3. Motile responses in outer hair cells.

    PubMed

    Zenner, H P

    1986-01-01

    Motile responses of cochlear hair cells open new perspectives for the understanding of cochlear hearing mechanisms and hearing disorders located in hair cells. Direct visualization of hair cell motility was achieved by a method for the study of living isolated mammalian outer hair cells (OHCs) which has overcome some of the complexities in dealing with the heterogeneous organ of Corti. Electrophysiological giga-seal whole-cell recordings of single OHC prepared by this approach had revealed negative cell potentials ranging from -32 mV to -70 mV (Gitter et al. (1986) Oto-Rhino-Laryngol. in press). Elucidation of HC motility has come from two lines of experiments. One follows from the observation that exposure of the lateral and basal membrane parts of living OHCs to increasing bath K+ concentrations resulted in a sustained reversible depolarization of the cell. Here, we report that by depolarization of the cell membrane in the presence of 25-125 mM K+/Cl- a sustained contraction of OHC was induced. This was followed by relaxation in the presence of artificial perilymph containing 5.4 mM K+/Cl-. By alternating these procedures OHCs were made to undergo as many as five cycles of contraction and relaxation. External Ca2+ was not required for the initial contraction but was essential for relaxation. Following repeated contraction/relaxation cycles the cytoplasm of individual OHCs exhibited a filamentous network, correlating with a new infracuticular anti-actin binding capacity. The second series of experiments originates in the observation that permeabilized OHCs contracted in the presence of ATP. No response was seen in the presence of control nucleotides.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3733547

  4. 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. PMID:20695442

  5. 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

  6. Biochemical studies of olfaction: isolation, characterization, and odorant binding activity of cilia from rainbow trout olfactory rosettes.

    PubMed

    Rhein, L D; Cagan, R H

    1980-08-01

    The role of cilia in recognition of olfactory stimuli has been controversial. Cilia from the intact olfactory rosettes of the rainbow trout Salmo gairdneri were isolated, characterized biochemically, and examined by electron microscopy. The markers studied are those associated with cilia in other organisms. Dynein arms contain Mg2+-AtPase; this enzyme was enriched in the isolated cilia preparation. Guanine nucleotides are associated with the outer microtubule doublets of cilia but adenine nucleotides are not; a substantial enrichment in guanine, relative to adenine, was found in the cilia preparation. Tubulin, the structural protein component of microtubules, occurs in large amounts in cilia. Disc gel electrophoresis indicated tubulin in the cilia preparation. Electron microscopy confirmed the presence of cilia in the isolated preparation. Rainbow trout have an acute sense of smell and many amino acids are odorants to this species. Functional activity of the cilia preparation relevant to odorant recognition was assessed by using binding of radioactively labeled odorant amino acids. L-Alanine, L-serine, L-threonine, L-lysine, and D-alanine bound to the cilia preparation. This study provides direct biochemical evidence that olfactory cilia bind odorant molecules and supports the hypothesis that odorant recognition sites are integral parts of the cilia. PMID:6449006

  7. 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.

  8. Mathematical embryology: the fluid mechanics of nodal cilia

    NASA Astrophysics Data System (ADS)

    Smith, D. J.; Smith, A. A.; Blake, J. R.

    2011-07-01

    Left-right symmetry breaking is critical to vertebrate embryonic development; in many species this process begins with cilia-driven flow in a structure termed the `node'. Primary `whirling' cilia, tilted towards the posterior, transport morphogen-containing vesicles towards the left, initiating left-right asymmetric development. We review recent theoretical models based on the point-force stokeslet and point-torque rotlet singularities, explaining how rotation and surface-tilt produce directional flow. Analysis of image singularity systems enforcing the no-slip condition shows how tilted rotation produces a far-field `stresslet' directional flow, and how time-dependent point-force and time-independent point-torque models are in this respect equivalent. Associated slender body theory analysis is reviewed; this approach enables efficient and accurate simulation of three-dimensional time-dependent flow, time-dependence being essential in predicting features of the flow such as chaotic advection, which have subsequently been determined experimentally. A new model for the nodal flow utilising the regularized stokeslet method is developed, to model the effect of the overlying Reichert's membrane. Velocity fields and particle paths within the enclosed domain are computed and compared with the flow profiles predicted by previous `membrane-less' models. Computations confirm that the presence of the membrane produces flow-reversal in the upper region, but no continuous region of reverse flow close to the epithelium. The stresslet far-field is no longer evident in the membrane model, due to the depth of the cavity being of similar magnitude to the cilium length. Simulations predict that vesicles released within one cilium length of the epithelium are generally transported to the left via a `loopy drift' motion, sometimes involving highly unpredictable detours around leftward cilia [truncated

  9. Mammalian aromatases.

    PubMed

    Conley, A; Hinshelwood, M

    2001-05-01

    Aromatase is the enzyme complex that catalyses the synthesis of oestrogens from androgens, and therefore it has unique potential to influence the physiological balance between the sex steroid hormones. Both aromatase cytochrome P450 (P450arom) and NADPH-cytochrome P450 reductase (reductase), the two essential components of the enzyme complex, are highly conserved among mammals and vertebrates. Aromatase expression occurs in the gonads and brain, and is essential for reproductive development and fertility. Of interest are the complex mechanisms involving alternative promoter utilization that have evolved to control tissue-specific expression in these tissues. In addition, in a number of species, including humans, expression of aromatase has a broader tissue distribution, including placenta, adipose and bone. The relevance of oestrogen synthesis and possibly androgen metabolism in these peripheral sites of expression is now becoming clear from studies in P450arom knockout (ArKO) mice and from genetic defects recognized recently in both men and women. Important species differences in the physiological roles of aromatase expression are also likely to emerge, despite the highly conserved nature of the enzyme system. The identification of functionally distinct, tissue-specific isozymes of P450arom in at least one mammal, pigs, and several species of fish indicates that there are additional subtle, but physiologically significant, species-specific roles for aromatase. Comparative studies of mammalian and other vertebrate aromatases will expand understanding of the role played by this ancient enzyme system in the evolution of reproduction and the adaptive influence of oestrogen synthesis on general health and well being. PMID:11427156

  10. 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

  11. 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

  12. Primary cilia are highly oriented with respect to collagen direction and long axis of extensor tendon.

    PubMed

    Donnelly, Eve; Ascenzi, Maria-Grazia; Farnum, Cornelia

    2010-01-01

    Skeletal tissues adapt to their mechanical environments by modulating gene expression, cell metabolism, and extracellular matrix (ECM) architecture; however, the mechanosensory mechanisms for these processes are incompletely understood. Primary cilia have emerged as critical components of the cellular mechanosensory apparatus and have been hypothesized to participate in establishment of cellular and ECM orientation, but their function in skeletal tissues is just beginning to be examined. Here we focused on tendon, a tissue with an oriented matrix that is ideal for analysis of spatial relationships between primary cilia and the ECM. The objective of this study was to characterize the incidence and orientation of tenocyte primary cilia in their native ECM. Primary cilia, nuclei, and collagen were analyzed three-dimensionally in immunofluorescently labeled rat extensor tendon using multiphoton microscopy and semiautomated morphometry. Primary cilia were observed in 64% of tenocytes. The cilia were highly oriented with respect to the ECM: cilia were aligned parallel to the collagen fibers and the long axis of the tendon. This study represents the first quantification of the in situ incidence and orientation of primary cilia in tendon. PMID:19603516

  13. 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

  14. Developmental changes in primary cilia in the mouse tooth germ and oral cavity.

    PubMed

    Hisamoto, Meri; Goto, Marie; Muto, Mami; Nio-Kobayashi, Junko; Iwanaga, Toshihiko; Yokoyama, Atsuro

    2016-01-01

    The primary cilium, a sensory apparatus, functions as both a chemical and mechanical sensor to receive environmental stimuli. The present study focused on the primary cilia in the epithelialmesenchymal interaction during tooth development. We examined the localization and direction of projection of primary cilia in the tooth germ and oral cavity of mice by immunohistochemical observation. Adenylyl cyclase 3 (ACIII)-immunolabeled cilia were visible in the inner/outer enamel epithelium of molars at the fetal stage and then conspicuously developed in the odontoblast layer postnatally. The primary cilia in ameloblasts and odontoblasts-shown by the double staining of acetylated tubulin and γ-tubulin-were regularly arranged from postnatal Day12, projecting apart from each other. The periodontal ligament possessed ACIII-positive cilia, which gathered on both sides of the dentin/cement and alveolar bone in postnatal days. In the oral cavity, numerous long primary cilia immunoreactive for ACIII were condensed at subepithelial stromal cells in the oral processes in fetuses, while postnatally a small number of short cilia were dispersed throughout the stroma of the oral cavity. These findings suggest that the primary cilia showing stage- and regionspecific morphology are involved in the epithelial-mesenchymal interaction during tooth development via mechano- and/or chemoreception for growth factors. PMID:27356608

  15. 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

  16. 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.

  17. 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.

  18. 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

  19. 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. PMID:22922713

  20. 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

  1. Regulation of cilia assembly, disassembly, and length by protein phosphorylation.

    PubMed

    Cao, Muqing; Li, Guihua; Pan, Junmin

    2009-01-01

    The exact mechanism by which cells are able to assemble, regulate, and disassemble cilia or flagella is not yet completely understood. Recent studies in several model systems, including Chlamydomonas, Tetrahymena, Leishmania, Caenorhabditis elegans, and mammals, provide increasing biochemical and genetic evidence that phosphorylation of multiple protein kinases plays a key role in cilia assembly, disassembly, and length regulation. Members of several protein kinase families--including aurora kinases, never in mitosis A (NIMA)-related protein kinases, mitogen-activated protein (MAP) kinases, and a novel cyclin-dependent protein kinase--are involved in the ciliary regulation process. Among the newly identified protein kinase substrates are Chlamydomonas kinesin-13 (CrKinesin13), a microtubule depolymerizer, and histone deacetylase 6 (HDAC6), a microtubule deacetylase. Chlamydomonas aurora/Ipl1p-like protein kinase (CALK) and CrKinesin13 are two proteins that undergo phosphorylation changes correlated with flagellar assembly or disassembly. CALK becomes phosphorylated when flagella are lost, whereas CrKinesin13 is phosphorylated when new flagella are assembled. Conversely, suppressing CrKinesin13 expression results in cells with shorter flagella. PMID:20362099

  2. 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

  3. Motility mutants of Dictyostelium discoideum

    PubMed Central

    1982-01-01

    We describe six motility mutants of Dictyostelium discoideum in this report. They were identified among a group of temperature-sensitive growth (Tsg) mutants that had been previously isolated using an enrichment for phagocytosis-defective cells. The Tsg mutants were screened for their ability to produce tracks on gold-coated cover slips, and several strains were found that were temperature-sensitive for migration in this assay. Analysis of spontaneous Tsg+ revertants of 10 migration-defective strains identified six strains that co-reverted the Tsg and track formation phenotypes. Characterization of these six strains indicated that they were defective at restrictive temperature in track formation, phagocytosis of bacteria, and pseudopodial and filopodial activity, while retaining normal rates of oxygen consumption and viability. Because they had lost this group of motile capabilities, these strains were designated motility mutants. The Tsg+ revertants of these mutants, which coordinately recovered all of the motile activities, were found at frequencies consistent with single genetic events. Analysis of the motility mutants and their revertants suggests a relationship between the motility mutations in some of these strains and genes affecting axenic growth. PMID:7118999

  4. Actin-based phagosome motility.

    PubMed

    Zhang, Fangliang; Southwick, Frederick S; Purich, Daniel L

    2002-10-01

    Despite abundant evidence of actin's involvement at the particle internalization stage of phagocytosis, little is known about whether phagosomes undergo the same type of actin-based motility as observed with endocytic vesicles or such intracellular pathogens as Listeria and Shigella. By employing video microscopy to follow the fate of latex bead-containing phagosomes within the cytoplasm of bone marrow macrophages, we have made the novel observation of actin-based phagosome motility. Immunofluorescence microscopy confirmed that phagosomes containing IgG-opsonized, bovine serum albumin (or BSA) -coated or uncoated latex beads all formed actin-rich rocket tails that persisted only during a brief, 1-2 min period of actin-based motility. Average speeds of actin-based phagosome motility were 0.13 +/- 0.06 microm/s for IgG-coated beads, 0.14 +/- 0.04 microm/s for BSA-coated beads, and 0.11+/- 0.03 microm/s for uncoated beads. Moreover, the speeds and motile-phase duration of each type of phagosome were comparable to the behavior of pinosomes [Merrifield et al., 1999: Nat. Cell Biol. 1:72-74.]. Determination of optimal conditions for observing and analyzing actin-based phagosome motility should facilitate future investigations of phagocytosis and phagosome maturation. PMID:12211106

  5. 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

  6. Longitudinal analysis of Plasmodium sporozoite motility in the dermis reveals component of blood vessel recognition.

    PubMed

    Hopp, Christine S; Chiou, Kevin; Ragheb, Daniel R T; Salman, Ahmed; 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. PMID:26271010

  7. 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

  8. Hyperglycemia in the absence of cilia accelerates cystogenesis and induces renal damage.

    PubMed

    Sas, Kelli M; Yin, Hong; Fitzgibbon, Wayne R; Baicu, Catalin F; Zile, Michael R; Steele, Stacy L; Amria, May; Saigusa, Takamitsu; Funk, Jason; Bunni, Marlene A; Siegal, Gene P; Siroky, Brian J; Bissler, John J; Bell, P Darwin

    2015-07-01

    In polycystic kidney disease (PKD), the rate of cyst formation and disease progression is highly variable. The lack of predictability in disease progression may be due to additional environmental factors or pathophysiological processes called "third hits." Diabetes is a growing epidemic, and recent studies suggest that PKD patients may be at an increased risk for this disease. We sought to determine if hyperglycemia enhances the initiation and rate of cystogenesis. Tamoxifen was administered to adult Ift88 conditional floxed allele mice to induce cilia loss in the presence of Cre. Subsequent administration of streptozotocin resulted in equivalent hyperglycemia in cilia(+) and cilia(-) mice. Hyperglycemia with loss of cilia increased the rate of cyst formation and cell proliferation. Structural and functional alterations in the kidney, including focal glomerular foot process effacement, interstitial inflammation, formation of primitive renal tubules, polyuria, and increased proteinuria, were also observed in hyperglycemic cilia(-) mice. Gene array analysis indicated enhanced Wnt and epithelial-to-mesenchymal transition signaling in the kidney of hyperglycemic cilia(-) mice. These data show that hyperglycemia, in the absence of cilia, results in renal structural and functional damage and accelerates cystogenesis, suggesting that diabetes is a risk factor in the progression of PKD. PMID:25904703

  9. Modulation of primary cilia length by melanin-concentrating hormone receptor 1.

    PubMed

    Hamamoto, Akie; Yamato, Shogo; Katoh, Yohei; Nakayama, Kazuhisa; Yoshimura, Kentaro; Takeda, Sen; Kobayashi, Yuki; Saito, Yumiko

    2016-06-01

    Melanin-concentrating hormone (MCH) receptor 1 (MCHR1) is a class A G-protein-coupled receptor (GPCR). The MCH-MCHR1 system has been implicated in the regulation of feeding, emotional processing, and sleep in rodents. Recent work revealed that MCHR1 is selectively expressed in neuronal primary cilia of the central nervous system. Cilia have various chemosensory functions in many types of cell, and ciliary dysfunction is associated with ciliopathies such as polycystic kidney disease and obesity. Although dynamic modulation of neuronal cilia length is observed in obese mice, the functional interaction of neuronal ciliary GPCR and its endogenous ligand has not yet been elucidated. We report here that MCH treatment significantly reduced cilia length in hTERT-RPE1 cells (hRPE1 cells) transfected with MCHR1. Quantitative analyses indicated that MCH-induced cilia shortening progressed in a dose-dependent manner with an EC50 lower than 1nM when cells were treated for 6h. Although the assembly and disassembly of primary cilia are tightly coupled to the cell cycle, cell cycle reentry was not a determinant of MCH-induced cilia shortening. We confirmed that MCH elicited receptor internalization, Ca(2+) mobilization, ERK and Akt phosphorylation, and inhibition of cyclic AMP accumulation in MCHR1-expressing hRPE1 cells. Among these diverse pathways, we revealed that Gi/o-dependent Akt phosphorylation was an important component in the initial stage of MCH-induced cilia length shortening. Furthermore, induction of fewer cilia by Kif3A siRNA treatment significantly decreased the MCH-mediated phosphorylation of Akt, indicating the functional importance of the MCHR1-Akt pathway in primary cilia. Taken together, the present data suggest that the MCH-MCHR1 axis may modulate the sensitivity of cells to external environments by controlling the cilia length. Therefore, further characterization of MCHR1 as a ciliary GPCR will provide a potential molecular mechanism to link cilia length

  10. 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.

  11. 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. PMID:26231314

  12. 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.

  13. [Mechanism of bacterial gliding motility].

    PubMed

    Nakane, Daisuke

    2015-01-01

    Bacteria have various way to move over solid surfaces, such as glass, agar, and host cell. These movements involve surface appendages including flagella, type IV pili and other "mysterious" nano-machineries. Gliding motility was a term used various surface movements by several mechanisms that have not been well understood in past few decades. However, development of visualization techniques allowed us to make much progress on their dynamics of machineries. It also provided us better understanding how bacteria move over surfaces and why bacteria move in natural environments. In this review, I will introduce recent studies on the gliding motility of Flavobacteium and Mycoplasma based on the detail observation of single cell and its motility machinery with micro-nano scales. PMID:26632217

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

    PubMed Central

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

    2014-01-01

    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. PMID:24853502

  15. Metachronal wave of artificial cilia array actuated by applied magnetic field

    NASA Astrophysics Data System (ADS)

    Tsumori, Fujio; Marume, Ryuma; Saijou, Akinori; Kudo, Kentaro; Osada, Toshiko; Miura, Hideshi

    2016-06-01

    In this paper, a biomimetic microstructure related to cilia, which are effective fluidic and conveying systems in nature, is described. Authors have already reported that a magnetic elastomer pillar actuated by a rotating magnetic field can work like a natural cilium. In the present work, we show examples of a cilia array with a metachronal wave as the next step. A metachronal wave is a sequential action of a number of cilia. It is theoretically known that a metachronal wave gives a higher fluidic efficiency; however, there has been no report on a metachronal wave by artificial cilia. We prepared magnetic elastomer pillars that contain chainlike clusters of magnetic particles. The orientation of chains was set to be different in each pillar so that each pillar will deform with a different phase.

  16. 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

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

    PubMed

    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

    2014-10-01

    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

  18. 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

  19. Reduced cilia frequencies in human renal cell carcinomas versus neighboring parenchymal tissue

    PubMed Central

    2013-01-01

    Background Cilia are essential organelles in multiple organ systems, including the kidney where they serve as important regulators of renal homeostasis. Renal nephron cilia emanate from the apical membrane of epithelia, extending into the lumen where they function in flow-sensing and ligand-dependent signaling cascades. Ciliary dysfunction underlies renal cyst formation that is in part caused by deregulation of planar cell polarity and canonical Wnt signaling. Renal cancer pathologies occur sporadically or in heritable syndromes caused by germline mutations in tumor suppressor genes including VHL. Importantly, Von Hippel-Lindau (VHL) patients frequently develop complex renal cysts that can be considered a premalignant stage. One of the well-characterized molecular functions of VHL is its requirement for the maintenance of cilia. In this study, tissue from 110 renal cancer patients who underwent nephrectomy was analyzed to determine if lower ciliary frequency is a common hallmark of renal tumorigenesis by comparing cilia frequencies in both tumor and adjacent parenchymal tissue biopsies from the same kidney. Methods We stained sections of human renal material using markers for cilia. Preliminary staining was performed using an immunofluorescent approach and a combination of acetylated-α-tubulin and pericentrin antibodies and DAPI. After validation of an alternative, higher throughput approach using acetylated-α-tubulin immunohistochemistry, we continued to manually quantify cilia in all tissues. Nuclei were separately counted in an automated fashion in order to determine ciliary frequencies. Similar staining and scoring for Ki67 positive cells was performed to exclude that proliferation obscures cilia formation potential. Results Samples from renal cell carcinoma patients deposited in our hospital tissue bank were previously used to compose a tissue microarray containing three cores of both tumor and parenchymal tissue per patient. Cilia frequencies in a total of

  20. Colonic motility in ulcerative colitis

    PubMed Central

    Antonelli, Elisabetta; Villanacci, Vincenzo; Baldoni, Monia; Dore, Maria Pina

    2014-01-01

    Background Inflammatory conditions affecting the gut may cause motility disturbances, and ulcerative colitis – one of the main disorders among the inflammatory bowel diseases – may display abnormal colonic motility. Aim To review the abnormalities of the large bowel in ulcerative colitis, by considering the motility, laboratory (in vitro) and pathological studies dealing with this topic. Methods A comprehensive online search of Medline and the Science Citation Index was carried out. Results Patients with ulcerative colitis frequently display colonic motor abnormalities, including lack of contractility, an increase of propulsive contractile waves, an excessive production of nitric oxide, vasoactive intestinal polypeptide nerves, interleukin 1 beta, neurotensin, tachykinins levels and the weaker action of substance P, likely related to a neuromuscular dysfunction due to the inflammatory process. Conclusions A better understanding of the pathophysiological grounds of altered colonic motility in ulcerative colitis may lead to a more in-depth knowledge of the accompanying symptoms and to better and more targeted therapeutic approaches. PMID:25452840

  1. 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

  2. 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.

  3. 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.

  4. 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

  5. An in vitro assay for entry into cilia reveals unique properties of the soluble diffusion barrier.

    PubMed

    Breslow, David K; Koslover, Elena F; Seydel, Federica; Spakowitz, Andrew J; Nachury, Maxence V

    2013-10-14

    Specific proteins are concentrated within primary cilia, whereas others remain excluded. To understand the mechanistic basis of entry into cilia, we developed an in vitro assay using cells in which the plasma membrane was permeabilized, but the ciliary membrane was left intact. Using a diffusion-to-capture system and quantitative analysis, we find that proteins >9 nm in diameter (∼100 kD) are restricted from entering cilia, and we confirm these findings in vivo. Interference with the nuclear pore complex (NPC) or the actin cytoskeleton in permeabilized cells demonstrated that the ciliary diffusion barrier is mechanistically distinct from those of the NPC or the axon initial segment. Moreover, applying a mass transport model to this system revealed diffusion coefficients for soluble and membrane proteins within cilia that are compatible with rapid exploration of the ciliary space in the absence of active transport. Our results indicate that large proteins require active transport for entry into cilia but not necessarily for movement inside cilia. PMID:24100294

  6. An in vitro assay for entry into cilia reveals unique properties of the soluble diffusion barrier

    PubMed Central

    Breslow, David K.; Koslover, Elena F.; Seydel, Federica; Spakowitz, Andrew J.

    2013-01-01

    Specific proteins are concentrated within primary cilia, whereas others remain excluded. To understand the mechanistic basis of entry into cilia, we developed an in vitro assay using cells in which the plasma membrane was permeabilized, but the ciliary membrane was left intact. Using a diffusion-to-capture system and quantitative analysis, we find that proteins >9 nm in diameter (∼100 kD) are restricted from entering cilia, and we confirm these findings in vivo. Interference with the nuclear pore complex (NPC) or the actin cytoskeleton in permeabilized cells demonstrated that the ciliary diffusion barrier is mechanistically distinct from those of the NPC or the axon initial segment. Moreover, applying a mass transport model to this system revealed diffusion coefficients for soluble and membrane proteins within cilia that are compatible with rapid exploration of the ciliary space in the absence of active transport. Our results indicate that large proteins require active transport for entry into cilia but not necessarily for movement inside cilia. PMID:24100294

  7. 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

  8. Osmotic damage as a predictor of motility loss during convective desiccation of bovine sperm.

    PubMed

    Sitaula, Ranjan; Jimenez, Jorge; Bhowmick, Sankha

    2013-12-01

    Current state-of-the art technologies are lagging in the application of desiccation storage to mammalian cells using nonreducing sugars. For bovine sperm, motility is irreversibly lost before reaching a sufficiently low moisture content necessary for preservation. It is hypothesized that much of the damage during drying is related to the osmotic stress encountered due to increased osmolarity of the extracellular environment. To test this hypothesis, we subjected sperm to liquid hyperosmotic environments for varying time-periods and measured their motility. We then extracted parameters for two models for motility loss based on these experiments: a first-order rate injury model (Fast or Slow) and a multi-modal (MM) injury model. The MM injury model incorporated an additional function accounting for damage induced by a time-independent osmotic change. Based on these models, we predicted sperm motility loss measured from natural and forced convective desiccation experiments. The MM injury model was able to closely bracket motility loss for desiccation as an osmotic change event with time-independent and time-dependent components. While the mechanistic basis of osmotic damage requires further exploration, the model can serve as a bracketing tool for predicting motility loss during desiccation based on excipients designed to minimize osmotic damage. PMID:24835367

  9. A mechanical model for guided motion of mammalian cells

    NASA Astrophysics Data System (ADS)

    Bitter, P.; Beck, K. L.; Lenz, P.

    2015-12-01

    We introduce a generic, purely mechanical model for environment-sensitive motion of mammalian cells that is applicable to chemotaxis, haptotaxis, and durotaxis as modes of motility. It is able to theoretically explain all relevant experimental observations, in particular, the high efficiency of motion, the behavior on inhomogeneous substrates, and the fixation of the lagging pole during motion. Furthermore, our model predicts that efficiency of motion in following a gradient depends on cell geometry (with more elongated cells being more efficient).

  10. Analysis of PCP Defects in Mammalian Eye Lens

    PubMed Central

    Sugiyama, Yuki; McAvoy, John W.

    2013-01-01

    Multicellular tissues and organs often show planar cell polarity (PCP) where the constituent cells align along an axis to form coordinated patterns. Mammalian eye lenses are mainly comprised of epithelial-derived fiber cells which exhibit highly ordered alignment that is regulated by PCP signaling. Each fiber cell has an apically situated primary cilium and in most cases this is polarized towards the lens anterior pole. Here we describe how to visualize the global cellular alignment of lens fiber cells by examining the suture pattern that is formed by the tips of fibers meeting at the anterior pole. We also describe a method for whole mount preparation which allows observation of the polarized distribution of primary cilia at the apical surface of lens fibers. Given its relative simplicity, at least in cellular terms, and its requirement for a high degree of precision in cellular alignment and orientation, we predict that the lens will be an excellent model system to help elucidate the role of cilia and PCP components in development of three-dimensional organization in tissues and organs. PMID:22218899

  11. 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

  12. The Shape of Motile Cells

    PubMed Central

    Mogilner, Alex; Keren, Kinneret

    2010-01-01

    Motile cells — fan-like keratocytes, hand-shaped nerve growth cones, polygonal fibroblasts, to name but a few — come in different shapes and sizes. We discuss the origins of this diversity as well as what shape tells us about the physics and biochemistry underlying cell movement. We start with geometric rules describing cell-edge kinetics that govern cell shape, followed by a discussion of the underlying biophysics; we consider actin treadmilling, actin–myosin contraction, cell-membrane deformations, adhesion, and the complex interactions between these modules, as well as their regulation by microtubules and Rho GTPases. Focusing on several different cell types, including keratocytes and fibroblasts, we discuss how dynamic cell morphology emerges from the interplay between the different motility modules and the environment. PMID:19906578

  13. 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

    2016-01-01

    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. PMID:27434268

  14. 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. PMID:25978533

  15. 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. PMID:26909801

  16. Secreted frizzled-related protein disrupts PCP in eye lens fiber cells that have polarised primary cilia.

    PubMed

    Sugiyama, Yuki; Stump, Richard J W; Nguyen, Anke; Wen, Li; Chen, Yongjuan; Wang, Yanshu; Murdoch, Jennifer N; Lovicu, Frank J; McAvoy, John W

    2010-02-15

    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 an 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

  17. 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

  18. Opposing microtubule motors control motility, morphology and cargo segregation during ER-to-Golgi transport.

    PubMed

    Brown, Anna K; Hunt, Sylvie D; Stephens, David J

    2014-01-01

    We recently demonstrated that dynein and kinesin motors drive multiple aspects of endosomal function in mammalian cells. These functions include driving motility, maintaining morphology (notably through providing longitudinal tension to support vesicle fission), and driving cargo sorting. Microtubule motors drive bidirectional motility during traffic between the endoplasmic reticulum (ER) and Golgi. Here, we have examined the role of microtubule motors in transport carrier motility, morphology, and domain organization during ER-to-Golgi transport. We show that, consistent with our findings for endosomal dynamics, microtubule motor function during ER-to-Golgi transport of secretory cargo is required for motility, morphology, and cargo sorting within vesicular tubular carriers en route to the Golgi. Our data are consistent with previous findings that defined roles for dynein-1, kinesin-1 (KIF5B) and kinesin-2 in this trafficking step. Our high resolution tracking data identify some intriguing aspects. Depletion of kinesin-1 reduces the number of motile structures seen, which is in line with other findings relating to the role of kinesin-1 in ER export. However, those transport carriers that were produced had a much greater run length suggesting that this motor can act as a brake on anterograde motility. Kinesin-2 depletion did not significantly reduce the number of motile transport carriers but did cause a similar increase in run length. These data suggest that kinesins act as negative regulators of ER-to-Golgi transport. Depletion of dynein not only reduced the number of motile carriers formed but also caused tubulation of carriers similar to that seen for sorting nexin-coated early endosomes. Our data indicated that the previously observed anterograde-retrograde polarity of transport carriers in transit to the Golgi from the ER is maintained by microtubule motor function. PMID:24705013

  19. 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

  20. Primary cilia regulate mTORC1 activity and cell size through Lkb1.

    PubMed

    Boehlke, Christopher; Kotsis, Fruzsina; Patel, Vishal; Braeg, Simone; Voelker, Henriette; Bredt, Saskia; Beyer, Theresa; Janusch, Heike; Hamann, Christoph; Gödel, Markus; Müller, Klaus; Herbst, Martin; Hornung, Miriam; Doerken, Mara; Köttgen, Michael; Nitschke, Roland; Igarashi, Peter; Walz, Gerd; Kuehn, E Wolfgang

    2010-11-01

    The mTOR pathway is the central regulator of cell size. External signals from growth factors and nutrients converge on the mTORC1 multi-protein complex to modulate downstream targets, but how the different inputs are integrated and translated into specific cellular responses is incompletely understood. Deregulation of the mTOR pathway occurs in polycystic kidney disease (PKD), where cilia (filiform sensory organelles) fail to sense urine flow because of inherited mutations in ciliary proteins. We therefore investigated if cilia have a role in mTOR regulation. Here, we show that ablation of cilia in transgenic mice results in enlarged cells when compared with control animals. In vitro analysis demonstrated that bending of the cilia by flow is required for mTOR downregulation and cell-size control. Surprisingly, regulation of cell size by cilia is independent of flow-induced calcium transients, or Akt. However, the tumour-suppressor protein Lkb1 localises in the cilium, and flow results in increased AMPK phosphorylation at the basal body. Conversely, knockdown of Lkb1 prevents normal cell-size regulation under flow conditions. Our results demonstrate that the cilium regulates mTOR signalling and cell size, and identify the cilium-basal body compartment as a spatially restricted activation site for Lkb1 signalling. PMID:20972424

  1. 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

  2. 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.

  3. 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.

  4. Proximal tubule proliferation is insufficient to induce rapid cyst formation after cilia disruption.

    PubMed

    Sharma, Neeraj; Malarkey, Erik B; Berbari, Nicolas F; O'Connor, Amber K; Vanden Heuvel, Gregory B; Mrug, Michal; Yoder, Bradley K

    2013-02-01

    Disrupting the function of cilia in mouse kidneys results in rapid or slow progression of cystic disease depending on whether the animals are juveniles or adults, respectively. Renal injury can also markedly accelerate the renal cyst formation that occurs after disruption of cilia in adult mice. Rates of cell proliferation are markedly higher in juvenile than adult kidneys and increase after renal injury, suggesting that cell proliferation may enhance the development of cysts. Here, we induced cilia loss in the kidneys of adult mice in the presence or absence of a Cux-1 transgene, which maintains cell proliferation. By using this model, we were able to avoid additional factors such as inflammation and dedifferentiation, which associate with renal injury and may also influence the rate of cystogenesis. After induction of cilia loss, cystic disease was not more pronounced in adult mice with the Cux-1 transgene compared with those without the transgene. In conclusion, these data suggest that proliferation is unlikely to be the sole mechanism underlying the rapid cystogenesis observed after injury in mice that lose cilia function in adulthood. PMID:23411784

  5. 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

  6. 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.

  7. Motility is crucial for the infectious life cycle of Borrelia burgdorferi.

    PubMed

    Sultan, Syed Z; Manne, Akarsh; Stewart, Philip E; Bestor, Aaron; Rosa, Patricia A; Charon, Nyles W; Motaleb, M A

    2013-06-01

    The Lyme disease spirochete, Borrelia burgdorferi, exists in a zoonotic cycle involving an arthropod tick and mammalian host. Dissemination of the organism within and between these hosts depends upon the spirochete's ability to traverse through complex tissues. Additionally, the spirochete outruns the host immune cells while migrating through the dermis, suggesting the importance of B. burgdorferi motility in evading host clearance. B. burgdorferi's periplasmic flagellar filaments are composed primarily of a major protein, FlaB, and minor protein, FlaA. By constructing a flaB mutant that is nonmotile, we investigated for the first time the absolute requirement for motility in the mouse-tick life cycle of B. burgdorferi. We found that whereas wild-type cells are motile and have a flat-wave morphology, mutant cells were nonmotile and rod shaped. These mutants were unable to establish infection in C3H/HeN mice via either needle injection or tick bite. In addition, these mutants had decreased viability in fed ticks. Our studies provide substantial evidence that the periplasmic flagella, and consequently motility, are critical not only for optimal survival in ticks but also for infection of the mammalian host by the arthropod tick vector. PMID:23529620

  8. Elastic mismatch enhances cell motility

    NASA Astrophysics Data System (ADS)

    Bresler, Yony; Palmieri, Benoit; Grant, Martin

    In recent years, the study of physics phenomena in cancer has drawn considerable attention. In cancer metastasis, a soft cancer cell leaves the tumor, and must pass through the endothelium before reaching the bloodstream. Using a phase-field model we have shown that the elasticity mismatch between cells alone is sufficient to enhance the motility of thesofter cancer cell by means of bursty migration, in agreement with experiment. We will present further characterization of these behaviour, as well as new possible applications for this model.

  9. The essential roles of transition fibers in the context of cilia

    PubMed Central

    Wei, Qing; Ling, Kun; Hu, Jinghua

    2015-01-01

    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. PMID:25988548

  10. Mutations in the Cilia Gene ARL13B Lead to the Classical Form of Joubert Syndrome

    PubMed Central

    Cantagrel, Vincent; Silhavy, Jennifer L.; Bielas, Stephanie L.; Swistun, Dominika; Marsh, Sarah E.; Bertrand, Julien Y.; Audollent, Sophie; Attié-Bitach, Tania; Holden, Kenton R.; Dobyns, William B.; Traver, David; Al-Gazali, Lihadh; Ali, Bassam R.; Lindner, Tom H.; Caspary, Tamara; Otto, Edgar A.; Hildebrandt, Friedhelm; Glass, Ian A.; Logan, Clare V.; Johnson, Colin A.; Bennett, Christopher; Brancati, Francesco; Valente, Enza Maria; Woods, C. Geoffrey; Gleeson, Joseph G.

    2008-01-01

    Joubert syndrome (JS) and related disorders are a group of autosomal-recessive conditions sharing the “molar tooth sign” on axial brain MRI, together with cerebellar vermis hypoplasia, ataxia, and psychomotor delay. JS is suggested to be a disorder of cilia function and is part of a spectrum of disorders involving retinal, renal, digital, oral, hepatic, and cerebral organs. We identified mutations in ARL13B in two families with the classical form of JS. ARL13B belongs to the Ras GTPase family, and in other species is required for ciliogenesis, body axis formation, and renal function. The encoded Arl13b protein was expressed in developing murine cerebellum and localized to the cilia in primary neurons. Overexpression of human wild-type but not patient mutant ARL13B rescued the Arl13b scorpion zebrafish mutant. Thus, ARL13B has an evolutionarily conserved role mediating cilia function in multiple organs. PMID:18674751

  11. A Group of ent-Kaurane Diterpenoids Inhibit Hedgehog Signaling and Induce Cilia Elongation

    PubMed Central

    Jiang, Shiyou; Du, Jiacheng; Kong, Qinghua; Li, Chaocui; Li, Yan; Sun, Handong; Pu, Jianxin; Mao, Bingyu

    2015-01-01

    The Hedgehog (Hh) signaling pathway plays important roles in the tumorigenesis of multiple cancers and is a key target for drug discovery. In a screen of natural products extracted from Chinese herbs, we identified eight ent-Kaurane diterpenoids and two triterpene dilactones as novel Hh pathway antagonists. Epistatic analyses suggest that these compounds likely act at the level or downstream of Smoothened (Smo) and upstream of Suppressor of Fused (Sufu). The ent-Kauranoid-treated cells showed elongated cilia, suppressed Smo trafficking to cilia, and mitotic defects, while the triterpene dilactones had no effect on the cilia and ciliary Smo. These ent-Kaurane diterpenoids provide new prototypes of Hh inhibitors, and are valuable probes for deciphering the mechanisms of Smo ciliary transport and ciliogenesis. PMID:26439749

  12. Cycling of the signaling protein phospholipase D through cilia requires the BBSome only for the export phase

    PubMed Central

    Brown, Jason M.; Sampaio, Julio L.; Craft, Julie M.; Shevchenko, Andrej; Evans, James E.; Witman, George B.

    2013-01-01

    The BBSome is a complex of seven proteins, including BBS4, that is cycled through cilia by intraflagellar transport (IFT). Previous work has shown that the membrane-associated signaling protein phospholipase D (PLD) accumulates abnormally in cilia of Chlamydomonas reinhardtii bbs mutants. Here we show that PLD is a component of wild-type cilia but is enriched ∼150-fold in bbs4 cilia; this accumulation occurs progressively over time and results in altered ciliary lipid composition. When wild-type BBSomes were introduced into bbs cells, PLD was rapidly removed from the mutant cilia, indicating the presence of an efficient BBSome-dependent mechanism for exporting ciliary PLD. This export requires retrograde IFT. Importantly, entry of PLD into cilia is BBSome and IFT independent. Therefore, the BBSome is required only for the export phase of a process that continuously cycles PLD through cilia. Another protein, carbonic anhydrase 6, is initially imported normally into bbs4 cilia but lost with time, suggesting that its loss is a secondary effect of BBSome deficiency. PMID:23589493

  13. MNS1 Is Essential for Spermiogenesis and Motile Ciliary Functions in Mice

    PubMed Central

    Zhou, Jian; Yang, Fang; Leu, N. Adrian; Wang, P. Jeremy

    2012-01-01

    During spermiogenesis, haploid round spermatids undergo dramatic cell differentiation and morphogenesis to give rise to mature spermatozoa for fertilization, including nuclear elongation, chromatin remodeling, acrosome formation, and development of flagella. The molecular mechanisms underlining these fundamental processes remain poorly understood. Here, we report that MNS1, a coiled-coil protein of unknown function, is essential for spermiogenesis. We find that MNS1 is expressed in the germ cells in the testes and localizes to sperm flagella in a detergent-resistant manner, indicating that it is an integral component of flagella. MNS1–deficient males are sterile, as they exhibit a sharp reduction in sperm production and the remnant sperm are immotile with abnormal short tails. In MNS1–deficient sperm flagella, the characteristic arrangement of “9+2” microtubules and outer dense fibers are completely disrupted. In addition, MNS1–deficient mice display situs inversus and hydrocephalus. MNS1–deficient tracheal motile cilia lack some outer dynein arms in the axoneme. Moreover, MNS1 monomers interact with each other and are able to form polymers in cultured somatic cells. These results demonstrate that MNS1 is essential for spermiogenesis, the assembly of sperm flagella, and motile ciliary functions. PMID:22396656

  14. CCDC65 Mutation Causes Primary Ciliary Dyskinesia with Normal Ultrastructure and Hyperkinetic Cilia

    PubMed Central

    Horani, Amjad; Brody, Steven L.; Ferkol, Thomas W.; Shoseyov, David; Wasserman, Mollie G.; Ta-shma, Asaf; Wilson, Kate S.; Bayly, Philip V.; Amirav, Israel; Cohen-Cymberknoh, Malena; Dutcher, Susan K.; Elpeleg, Orly; Kerem, Eitan

    2013-01-01

    Background Primary ciliary dyskinesia (PCD) is a genetic disorder characterized by impaired ciliary function, leading to chronic sinopulmonary disease. The genetic causes of PCD are still evolving, while the diagnosis is often dependent on finding a ciliary ultrastructural abnormality and immotile cilia. Here we report a novel gene associated with PCD but without ciliary ultrastructural abnormalities evident by transmission electron microscopy, but with dyskinetic cilia beating. Methods Genetic linkage analysis was performed in a family with a PCD subject. Gene expression was studied in Chlamydomonas reinhardtii and human airway epithelial cells, using RNA assays and immunostaining. The phenotypic effects of candidate gene mutations were determined in primary culture human tracheobronchial epithelial cells transduced with gene targeted shRNA sequences. Video-microscopy was used to evaluate cilia motion. Results A single novel mutation in CCDC65, which created a termination codon at position 293, was identified in a subject with typical clinical features of PCD. CCDC65, an orthologue of the Chlamydomonas nexin-dynein regulatory complex protein DRC2, was localized to the cilia of normal nasal epithelial cells but was absent in those from the proband. CCDC65 expression was up-regulated during ciliogenesis in cultured airway epithelial cells, as was DRC2 in C. reinhardtii following deflagellation. Nasal epithelial cells from the affected individual and CCDC65-specific shRNA transduced normal airway epithelial cells had stiff and dyskinetic cilia beating patterns compared to control cells. Moreover, Gas8, a nexin-dynein regulatory complex component previously identified to associate with CCDC65, was absent in airway cells from the PCD subject and CCDC65-silenced cells. Conclusion Mutation in CCDC65, a nexin-dynein regulatory complex member, resulted in a frameshift mutation and PCD. The affected individual had altered cilia beating patterns, and no detectable

  15. 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.

  16. Metachronal beating of cilia under the influence of Casson fluid and magnetic field

    NASA Astrophysics Data System (ADS)

    Akbar, Noreen Sher; Khan, Zafar Hayat

    2015-03-01

    Metachronal beating of cilia under the influence of Casson fluid and magnetic field is considered. The model for cilia literature is modelled for the first time. The governing coupled equations are constructed under long wavelength and low Reynold's number approximation. Exact solutions are evaluated for stream function and pressure gradient. The important results in this study are the variation of the Hartmann number M, Casson fluid parameter ζ. The velocity field increases due to the increase in Hartmann number M near the channel walls while velocity field decreases at the center of the channel. Comparative study is also made for Casson fluid with Newtonian fluid.

  17. Immotile cilia syndrome: A recombinant family at HLA-linked gene locus

    SciTech Connect

    Gasparini, P.; Grifa, A.; Oggiano, N.; Fabbrizzi, E.; Giorgi, P.L.

    1994-02-15

    The immotile-cilia syndrome (ICS) is an autosomal recessive trait of congenital dismobility or even complete immobility of cilia in the ciliated epithelia (MIM 244400). Recurrent upper respiratory infections in early childhood are the most common clinical findings. Recently a disease locus was mapped by sib pair analysis in two unrelated families on 6p tightly linked to HLA class II loci, such as DR and DQ. In order to confirm this assignment and to test the presence of possible heterogeneity, the authors analyzed several ICS families utilizing DNA makers of HLA class II region. Here they report the identification of a recombinant family at this locus. 3 refs., 1 fig.

  18. Autonomic control of gut motility: a comparative view.

    PubMed

    Olsson, Catharina; Holmgren, Susanne

    2011-11-16

    Gut motility is regulated to optimize food transport and processing. The autonomic innervation of the gut generally includes extrinsic cranial and spinal autonomic nerves. It also comprises the nerves contained entirely within the gut wall, i.e. the enteric nervous system. The extrinsic and enteric nervous control follows a similar pattern throughout the vertebrate groups. However, differences are common and may occur between groups and families as well as between closely related species. In this review, we give an overview of the distribution and effects of common neurotransmitters in the vertebrate gut. While the focus is on birds, reptiles, amphibians and fish, mammalian data are included to form the background for comparisons. While some transmitters, like acetylcholine and nitric oxide, show similar distribution patterns and effects in most species investigated, the role of others is more varying. The significance for these differences is not yet fully understood, emphasizing the need for continued comparative studies of autonomic control. PMID:20724224

  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. Innervation of Gill Lateral Cells in the Bivalve Mollusc Crassostrea virginica Affects Cellular Membrane Potential and Cilia Activity

    PubMed Central

    Catapane, Edward J; Nelson, Michael; Adams, Trevon; Carroll, Margaret A

    2016-01-01

    Gill lateral cells of Crassostrea virginica are innervated by the branchial nerve, which contains serotonergic and dopaminergic fibers that regulate cilia beating rate. Terminal release of serotonin or dopamine results in an increase or decrease, respectively, of cilia beating rate in lateral gill cells. In this study we used the voltage sensitive fluorescent probe DiBAC4(3) to quantify changes in gill lateral cell membrane potential in response to electrical stimulation of the branchial nerve or to applications of serotonin and dopamine, and correlate these changes to cilia beating rates. Application of serotonin to gill lateral cells caused prolonged membrane depolarization, similar to plateau potentials, while increasing cilia beating rate. Application of dopamine hyperpolarized the resting membrane while decreasing cilia beating rate. Low frequency (5 Hz) electrical stimulations of the branchial nerve, which cause terminal release of endogenous serotonin, or high frequency (20 Hz) stimulations, which cause terminal release of endogenous dopamine, had the same effects on gill lateral cell membrane potentials and cilia beating rate as the respective applications of serotonin or dopamine. The study shows that innervation of gill lateral cells by the branchial nerve affects membrane potential as well as cilia beating rate, and demonstrates a strong correlation between changes in membrane potential and regulation of cilia beating rate. The study furthers the understanding of serotonin and dopamine signaling in the innervation and regulation of gill cilia in bivalves. The study also shows that voltage sensitive fluorescent probes like DiBAC 4(3) can be successfully used as an alternative to microelectrodes to measure changes in membrane potential of ciliated gill cells and other small cells with fast moving cilia. PMID:27489887

  1. GPR88 Reveals a Discrete Function of Primary Cilia as Selective Insulators of GPCR Cross-Talk

    PubMed Central

    Marley, Aaron; Choy, Regina Wai-Yan; von Zastrow, Mark

    2013-01-01

    A number of G protein-coupled receptors (GPCRs) localize to primary cilia but the functional significance of cilia to GPCR signaling remains incompletely understood. We investigated this question by focusing on the D1 dopamine receptor (D1R) and beta-2 adrenergic receptor (B2AR), closely related catecholamine receptors that signal by stimulating production of the diffusible second messenger cyclic AMP (cAMP) but differ in localization relative to cilia. D1Rs robustly concentrate on cilia of IMCD3 cells, as shown previously in other ciliated cell types, but disrupting cilia did not affect D1R surface expression or ability to mediate a concentration-dependent cAMP response. By developing a FRET-based biosensor suitable for resolving intra- from extra- ciliary cAMP changes, we found that the D1R-mediated cAMP response is not restricted to cilia and extends into the extra-ciliary cytoplasm. Conversely the B2AR, which we show here is effectively excluded from cilia, also generated a cAMP response in both ciliary and extra-ciliary compartments. We identified a distinct signaling effect of primary cilia through investigating GPR88, an orphan GPCR that is co-expressed with the D1R in brain, and which we show here is targeted to cilia similarly to the D1R. In ciliated cells, mutational activation of GPR88 strongly reduced the D1R-mediated cAMP response but did not affect the B2AR-mediated response. In marked contrast, in non-ciliated cells, GPR88 was distributed throughout the plasma membrane and inhibited the B2AR response. These results identify a discrete ‘insulating’ function of primary cilia in conferring selectivity on integrated catecholamine signaling through lateral segregation of receptors, and suggest a cellular activity of GPR88 that might underlie its effects on dopamine-dependent behaviors. PMID:23936473

  2. A novel serotonin-secreting cell type regulates ciliary motility in the mucociliary epidermis of Xenopus tadpoles.

    PubMed

    Walentek, Peter; Bogusch, Susanne; Thumberger, Thomas; Vick, Philipp; Dubaissi, Eamon; Beyer, Tina; Blum, Martin; Schweickert, Axel

    2014-04-01

    The embryonic skin of Xenopus tadpoles serves as an experimental model system for mucociliary epithelia (MCE) such as the human airway epithelium. MCEs are characterized by the presence of mucus-secreting goblet and multiciliated cells (MCCs). A third cell type, ion-secreting cells (ISCs), is present in the larval skin as well. Synchronized beating of MCC cilia is required for directional transport of mucus. Here we describe a novel cell type in the Xenopus laevis larval epidermis, characterized by serotonin synthesis and secretion. It is termed small secretory cell (SSC). SSCs are detectable at early tadpole stages, unlike MCCs and ISCs, which are specified at early neurulation. Subcellularly, serotonin was found in large, apically localized vesicle-like structures, which were entirely shed into the surrounding medium. Pharmacological inhibition of serotonin synthesis decreased the velocity of cilia-driven fluid flow across the skin epithelium. This effect was mediated by serotonin type 3 receptor (Htr3), which was expressed in ciliated cells. Knockdown of Htr3 compromised flow velocity by reducing the ciliary motility of MCCs. SSCs thus represent a distinct and novel entity of the frog tadpole MCE, required for ciliary beating and mucus transport across the larval skin. The identification and characterization of SSCs consolidates the value of the Xenopus embryonic skin as a model system for human MCEs, which have been known for serotonin-dependent regulation of ciliary beat frequency. PMID:24598162

  3. 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

  4. Exact solution of cilia induced flow of a Jeffrey fluid in an inclined tube.

    PubMed

    Maqbool, K; Shaheen, S; Mann, A B

    2016-01-01

    The present study investigated the cilia induced flow of MHD Jeffrey fluid through an inclined tube. This study is carried out under the assumptions of long wavelength and low Reynolds number approximations. Exact solutions for the velocity profile, pressure rise, pressure gradient, volume flow rate and stream function are obtained. Effects of pertinent physical parameters on the computational results are presented graphically. PMID:27610298

  5. Microtubule modifications and stability are altered by cilia perturbation and in cystic kidney disease

    PubMed Central

    Berbari, Nicolas F.; Sharma, Neeraj; Malarkey, Erik B.; Pieczynski, Jay N.; Boddu, Ravindra; Gaertig, Jacek; Guay-Woodford, Lisa; Yoder, Bradley K.

    2013-01-01

    Summary Disruption of the primary cilium is associated with a growing number of human diseases collectively termed ciliopathies. Ciliopathies present with a broad range of clinical features consistent with the near ubiquitous nature of the organelle and its role in diverse signaling pathways throughout development and adult homeostasis. The clinical features associated with cilia dysfunction can include such phenotypes as polycystic kidneys, skeletal abnormalities, blindness, anosmia, and obesity. Although the clinical relevance of the primary cilium is evident, the effects that cilia dysfunction has on the cell and how this contributes to disease remains poorly understood. Here, we show that loss of ciliogenesis genes such as Ift88 and Kif3a lead to increases in post-translational modifications on cytosolic microtubules. This effect was observed in cilia mutant kidney cells grown in vitro and in vivo in cystic kidneys. The hyper-acetylation of microtubules resulting from cilia loss is associated with both altered microtubule stability and increased α-tubulin acetyl-transferase activity. Intriguingly, the effect on microtubules was also evident in renal samples from patients with autosomal recessive polycystic kidneys. These findings indicate that altered microtubule post-translational modifications may influence some of the phenotypes observed in ciliopathies. PMID:23124988

  6. 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. PMID:25399777

  7. 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.

  8. 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

  9. 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. PMID:494232

  10. Asymmetric Distribution of Primary Cilia Allocates Satellite Cells for Self-Renewal.

    PubMed

    Jaafar Marican, Nur Hayati; Cruz-Migoni, Sara B; Borycki, Anne-Gaëlle

    2016-06-14

    Regeneration of vertebrate skeletal muscles requires satellite cells, a population of stem cells that are quiescent in normal conditions and divide, differentiate, and self-renew upon activation triggered by exercise, injury, and degenerative diseases. Satellite cell self-renewal is essential for long-term tissue homeostasis, and previous work has identified a number of external cues that control this process. However, little is known of the possible intrinsic control mechanisms of satellite cell self-renewal. Here, we show that quiescent satellite cells harbor a primary cilium, which is rapidly disassembled upon entry into the cell cycle. Contrasting with a commonly accepted belief, cilia reassembly does not occur uniformly in cells exiting the cell cycle. We found that primary cilia reassemble preferentially in cells committed to self-renew, and disruption of cilia reassembly causes a specific deficit in self-renewing satellite cells. These observations indicate that primary cilia provide an intrinsic cue essential for satellite cell self-renewal. PMID:27161363

  11. Cyst growth, polycystins, and primary cilia in autosomal dominant polycystic kidney disease.

    PubMed

    Lee, Seung Hun; Somlo, Stefan

    2014-06-01

    The primary cilium of renal epithelia acts as a transducer of extracellular stimuli. Polycystin (PC)1 is the protein encoded by the PKD1 gene that is responsible for the most common and severe form of autosomal dominant polycystic kidney disease (ADPKD). PC1 forms a complex with PC2 via their respective carboxy-terminal tails. Both proteins are expressed in the primary cilia. Mutations in either gene affect the normal architecture of renal tubules, giving rise to ADPKD. PC1 has been proposed as a receptor that modulates calcium signals via the PC2 channel protein. The effect of PC1 dosage has been described as the rate-limiting modulator of cystic disease. Reduced levels of PC1 or disruption of the balance in PC1/PC2 level can lead to the clinical features of ADPKD, without complete inactivation. Recent data show that ADPKD resulting from inactivation of polycystins can be markedly slowed if structurally intact cilia are also disrupted at the same time. Despite the fact that no single model or mechanism from these has been able to describe exclusively the pathogenesis of cystic kidney disease, these findings suggest the existence of a novel cilia-dependent, cyst-promoting pathway that is normally repressed by polycystin function. The results enable us to rethink our current understanding of genetics and cilia signaling pathways of ADPKD. PMID:26877954

  12. Loss of primary cilia upregulates renal hypertrophic signaling and promotes cystogenesis.

    PubMed

    Bell, P Darwin; 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

    2011-05-01

    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

  13. Prickle3 synergizes with Wtip to regulate basal body organization and cilia growth

    PubMed Central

    Chu, Chih-Wen; Ossipova, Olga; Ioannou, Andriani; Sokol, Sergei Y.

    2016-01-01

    PCP proteins maintain planar polarity in many epithelial tissues and have been implicated in cilia development in vertebrate embryos. In this study we examine Prickle3 (Pk3), a vertebrate homologue of Drosophila Prickle, in Xenopus gastrocoel roof plate (GRP). GRP is a tissue equivalent to the mouse node, in which cilia-generated flow promotes left-right patterning. We show that Pk3 is enriched at the basal body of GRP cells but is recruited by Vangl2 to anterior cell borders. Interference with Pk3 function disrupted the anterior polarization of endogenous Vangl2 and the posterior localization of cilia in GRP cells, demonstrating its role in PCP. Strikingly, in cells with reduced Pk3 activity, cilia growth was inhibited and γ-tubulin and Nedd1 no longer associated with the basal body, suggesting that Pk3 has a novel function in basal body organization. Mechanistically, this function of Pk3 may involve Wilms tumor protein 1-interacting protein (Wtip), which physically associates with and cooperates with Pk3 to regulate ciliogenesis. We propose that, in addition to cell polarity, PCP components control basal body organization and function. PMID:27062996

  14. Hedgehog signaling regulates myelination in the peripheral nervous system through primary cilia.

    PubMed

    Yoshimura, Kentaro; Takeda, Sen

    2012-02-01

    Myelination is an essential prerequisite for the nervous system to transmit an impulse efficiently by a saltatory conduction. In the peripheral nervous system (PNS), Schwann cells (SCs) engage in myelination. However, a detailed molecular mechanism underlying myelination still remains unclear. In this study, we hypothesized that the primary cilia of SCs are the regulators of Hedgehog (Hh) signaling-mediated myelination. To confirm our hypothesis, we used mouse dorsal root ganglion (DRG)/SC co-cultures, wherein the behavior of SCs could be analyzed by maintaining the interaction of SCs with DRG neurons. Under these conditions, SCs had primary cilia, and Hh signaling molecules accumulated on the primary cilia. When the SCs were stimulated by the addition of desert hedgehog or smoothened agonist, formation of myelin segments on the DRG axons was facilitated. On the contrary, upon administration of cyclopamine, an inhibitor of Hh signaling, myelin segments became comparable to those of controls. Of note, the ratio of SCs harboring primary cilium reached the highest point during the early phase of myelination. Furthermore, the strongest effects of Hh on myelination were encountered during the same stage. These results collectively indicate that Hh signaling regulates myelin formation through primary cilia in the PNS. PMID:22101064

  15. 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

  16. 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. PMID:27272319

  17. Identification of G Protein-Coupled Receptors (GPCRs) in Primary Cilia and Their Possible Involvement in Body Weight Control.

    PubMed

    Omori, Yoshihiro; Chaya, Taro; Yoshida, Satoyo; Irie, Shoichi; Tsujii, Toshinori; Furukawa, Takahisa

    2015-01-01

    Primary cilia are sensory organelles that harbor various receptors such as G protein-coupled receptors (GPCRs). We analyzed subcellular localization of 138 non-odorant GPCRs. We transfected GPCR expression vectors into NIH3T3 cells, induced ciliogenesis by serum starvation, and observed subcellular localization of GPCRs by immunofluorescent staining. We found that several GPCRs whose ligands are involved in feeding behavior, including prolactin-releasing hormone receptor (PRLHR), neuropeptide FF receptor 1 (NPFFR1), and neuromedin U receptor 1 (NMUR1), localized to the primary cilia. In addition, we found that a short form of dopamine receptor D2 (DRD2S) is efficiently transported to the primary cilia, while a long form of dopamine receptor D2 (DRD2L) is rarely transported to the primary cilia. Using an anti-Prlhr antibody, we found that Prlhr localized to the cilia on the surface of the third ventricle in the vicinity of the hypothalamic periventricular nucleus. We generated the Npy2r-Cre transgenic mouse line in which Cre-recombinase is expressed under the control of the promoter of Npy2r encoding a ciliary GPCR. By mating Npy2r-Cre mice with Ift80 flox mice, we generated Ift80 conditional knockout (CKO) mice in which Npy2r-positive cilia were diminished in number. We found that Ift80 CKO mice exhibited a body weight increase. Our results suggest that Npy2r-positive cilia are important for body weight control. PMID:26053317

  18. Galectin-7 modulates the length of the primary cilia and wound repair in polarized kidney epithelial cells.

    PubMed

    Rondanino, Christine; Poland, Paul A; Kinlough, Carol L; Li, Hui; Rbaibi, Youssef; Myerburg, Michael M; Al-bataineh, Mohammad M; Kashlan, Ossama B; Pastor-Soler, Nuria M; Hallows, Kenneth R; Weisz, Ora A; Apodaca, Gerard; Hughey, Rebecca P

    2011-09-01

    Galectins (Gal) are β-galactoside-binding proteins that function in epithelial development and homeostasis. An overlapping role for Gal-3 and Gal-7 in wound repair was reported in stratified epithelia. Although Gal-7 was thought absent in simple epithelia, it was reported in a proteomic analysis of cilia isolated from cultured human airway, and we recently identified Gal-7 transcripts in Madin-Darby canine kidney (MDCK) cells (Poland PA, Rondanino C, Kinlough CL, Heimburg-Molinaro J, Arthur CM, Stowell SR, Smith DF, Hughey RP. J Biol Chem 286: 6780-6790, 2011). We now report that Gal-7 is localized exclusively on the primary cilium of MDCK, LLC-PK(1) (pig kidney), and mpkCCD(c14) (mouse kidney) cells as well as on cilia in the rat renal proximal tubule. Gal-7 is also present on most cilia of multiciliated cells in human airway epithelia primary cultures. Interestingly, exogenous glutathione S-transferase (GST)-Gal-7 bound the MDCK apical plasma membrane as well as the cilium, while the lectin Ulex europeaus agglutinin, with glycan preferences similar to Gal-7, bound the basolateral plasma membrane as well as the cilium. In pull-down assays, β1-integrin isolated from either the basolateral or apical/cilia membranes of MDCK cells was similarly bound by GST-Gal-7. Selective localization of Gal-7 to cilia despite the presence of binding sites on all cell surfaces suggests that intracellular Gal-7 is specifically delivered to cilia rather than simply binding to surface glycoconjugates after generalized secretion. Moreover, depletion of Gal-7 using tetracycline-induced short-hairpin RNA in mpkCCD(c14) cells significantly reduced cilia length and slowed wound healing in a scratch assay. We conclude that Gal-7 is selectively targeted to cilia and plays a key role in surface stabilization of glycoconjugates responsible for integrating cilia function with epithelial repair. PMID:21677144

  19. Mechanisms of cilia-driven transport in the airways in the absence of mucus.

    PubMed

    Bermbach, Saskia; Weinhold, Karina; Roeder, Thomas; Petersen, Frank; Kugler, Christian; Goldmann, Torsten; Rupp, Jan; König, Peter

    2014-07-01

    Airway mucus is thought to be required for the clearance of inhaled particles by mucociliary transport, but this view has recently been challenged. To test if mucus is necessary for cilia-driven particle transport, we removed mucus from murine and human ex vivo airway preparations by thorough rinsing with buffer with or without additional dithiothreitol washing. The transport of particles with diameters of 4.5 μm, 200 nm, and 40 nm and of bacteria was analyzed by video microscopy. Complete removal of mucus was verified by wheat germ agglutinin staining and by scanning electron microscopy. In the absence of mucus, we observed efficient transport of particles and bacteria by direct cilia-mediated propulsion or via fluid flow generated by ciliary beating. Virus-sized particles had the tendency to attach to cilia. Because direct contact of particles with ciliated cells occurs in the absence of mucus, we examined if this direct interaction changes epithelial function. Neither bacteria- nor LPS-induced nuclear translocation of NF-κB p65 in ciliated cells occurred, indicating that mere contact between ciliated cells and bacteria during transport does not activate the epithelium. Attachment of virus-sized particles to cilia could induce mucus release and/or increase the ciliary beat frequency. Our results indicate that cilia-driven transport of particles with various sizes is possible in murine and human airways without the presence of mucus. If mucus-free transport fails, the epithelium can react by releasing mucus or increasing the ciliary beat frequency to maintain particle transport. PMID:24467665

  20. Protonmotive force and motility of Bacillus subtilis.

    PubMed Central

    Shioi, J I; Imae, Y; Oosawa, F

    1978-01-01

    Motility of Bacillus subtilis was inhibited within a few minutes by a combination of valinomycin and a high concentration of potassium ions in the medium at neutral pH. Motility was restored by lowering the concentration of valinomycin or potassium ions. The valinomycin concentration necessary for motility inhibition was determined at various concentrations of potassium ions and various pH's. At pH 7.5, valinomycin of any concentration did not inhibit the motility, when the potassium ion concentration was lower than 9 mM. In the presence of 230 mM potassium ion, the motility inhibition by valinomycin was not detected at pH lower than 6.1. These results are easily explained by the idea that the motility of B. subtilis is supported by the electrochemical potential difference of the proton across the membrane, or the protonmotive force. The electrochemical potential difference necessary for motility was estimated to be about -90 mV. PMID:25261

  1. 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

  2. Auxin transport inhibitors impair vesicle motility and actin cytoskeleton dynamics in diverse eukaryotes

    PubMed Central

    Dhonukshe, Pankaj; Grigoriev, Ilya; Fischer, Rainer; Tominaga, Motoki; Robinson, David G.; Hašek, Jiří; Paciorek, Tomasz; Petrášek, Jan; Seifertová, Daniela; Tejos, Ricardo; Meisel, Lee A.; Zažímalová, Eva; Gadella, Theodorus W. J.; Stierhof, York-Dieter; Ueda, Takashi; Oiwa, Kazuhiro; Akhmanova, Anna; Brock, Roland; Spang, Anne; Friml, Jiří

    2008-01-01

    Many aspects of plant development, including patterning and tropisms, are largely dependent on the asymmetric distribution of the plant signaling molecule auxin. Auxin transport inhibitors (ATIs), which interfere with directional auxin transport, have been essential tools in formulating this concept. However, despite the use of ATIs in plant research for many decades, the mechanism of ATI action has remained largely elusive. Using real-time live-cell microscopy, we show here that prominent ATIs such as 2,3,5-triiodobenzoic acid (TIBA) and 2-(1-pyrenoyl) benzoic acid (PBA) inhibit vesicle trafficking in plant, yeast, and mammalian cells. Effects on micropinocytosis, rab5-labeled endosomal motility at the periphery of HeLa cells and on fibroblast mobility indicate that ATIs influence actin cytoskeleton. Visualization of actin cytoskeleton dynamics in plants, yeast, and mammalian cells show that ATIs stabilize actin. Conversely, stabilizing actin by chemical or genetic means interferes with endocytosis, vesicle motility, auxin transport, and plant development, including auxin transport-dependent processes. Our results show that a class of ATIs act as actin stabilizers and advocate that actin-dependent trafficking of auxin transport components participates in the mechanism of auxin transport. These studies also provide an example of how the common eukaryotic process of actin-based vesicle motility can fulfill a plant-specific physiological role. PMID:18337510

  3. 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

  4. Mammalian cardiolipin biosynthesis.

    PubMed

    Mejia, Edgard M; Nguyen, Hieu; Hatch, Grant M

    2014-04-01

    Cardiolipin is a major phospholipid in mitochondria and is involved in the generation of cellular energy in the form of ATP. In mammalian and eukaryotic cells it is synthesized via the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol phosphate pathway. This brief review will describe some of the more recent studies on mammalian cardiolipin biosynthesis and provide an overview of regulation of cardiolipin biosynthesis. In addition, the important role that this key phospholipid plays in disease processes including heart failure, diabetes, thyroid hormone disease and the genetic disease Barth Syndrome will be discussed. PMID:24144810

  5. Extracellular Regulation of Sperm Transmembrane Adenylyl Cyclase by a Forward Motility Stimulating Protein

    PubMed Central

    Dey, Souvik; Roy, Debarun; Majumder, Gopal C.; Bhattacharyya, Debdas

    2014-01-01

    of progressive motility by a physiologic extracellular factor in a mammalian species. PMID:25350397

  6. [Effect of drugs on granulocyte motility].

    PubMed

    Schmidt, D; Morenz, J

    1985-01-01

    The in-vitro influence of drugs on the chemokinesis and chemotaxis of neutrophils was investigated in order to prevent additional drug-induced motility impairment of cells in cases of already existing host defense disorders and for an eventual specific treatment of motility defects. Granulocyte motility is unimpaired by penicillin, ampicillin, carbenicillin, streptomycin, nystatin, and cyclophosphamide. The chemokinesis and chemotaxis of neutrophils are inhibited by erythromycin, oxytetracycline, doxycycline, chloramphenicol, hydrocortisone, g-strophanthin, digoxin, and digitoxin and in higher concentrations also by sulfonamides, gentamycin, prednisolone, methylprednisolone, dexamethasone, and phenylbutazone. Chemotaxis is selectively or rather more inhibited than chemokinesis by amphotericin B, griseofulvin, vinblastine++, trifluoperazine, and promethazine. Granulocyte motility is, however, stimulated by ascorbic acid, potassium thiocyanate, levamisole, lithium, and metofenazate. PMID:3161313

  7. Motility in the epsilon-proteobacteria.

    PubMed

    Beeby, Morgan

    2015-12-01

    The epsilon-proteobacteria are a widespread group of flagellated bacteria frequently associated with either animal digestive tracts or hydrothermal vents, with well-studied examples in the human pathogens of Helicobacter and Campylobacter genera. Flagellated motility is important to both pathogens and hydrothermal vent members, and a number of curious differences between the epsilon-proteobacterial and enteric bacterial motility paradigms make them worthy of further study. The epsilon-proteobacteria have evolved to swim at high speed and through viscous media that immobilize enterics, a phenotype that may be accounted for by the molecular architecture of the unusually large epsilon-proteobacterial flagellar motor. This review summarizes what is known about epsilon-proteobacterial motility and focuses on a number of recent discoveries that rationalize the differences with enteric flagellar motility. PMID:26590774

  8. Implications of altered gastrointestinal motility in obesity.

    PubMed

    Gallagher, T K; Geoghegan, J G; Baird, A W; Winter, D C

    2007-10-01

    The onset of obesity occurs as a result of an imbalance between nutrient consumption/absorption and energy expenditure. Gastrointestinal (GI) motility plays a critical role in the rate of consumption of foods, digestion, and absorption of nutrients. Various segments of the GI tract coordinate in a complex yet precise way, to control the process of food consumption, digestion, and absorption of nutrients. GI motility not only regulates the rates at which nutrients are processed and absorbed in the gut, but also, via mechanical and neurohormonal methods, participates in the control of appetite and satiety. Altered GI motility has frequently been observed in obese patients, the significance of which is incompletely understood. However, these alterations can be considered as potential contributing factors in the development and maintenance of obesity and changed eating behavior. Therapies aimed at regulating or counteracting the observed changes in GI motility are being actively explored and applied clinically in the management of obese patients. PMID:18098402

  9. Flagellum Density Regulates Proteus mirabilis Swarmer Cell Motility in Viscous Environments

    PubMed Central

    Tuson, Hannah H.; Copeland, Matthew F.; Carey, Sonia; Sacotte, Ryan

    2013-01-01

    Proteus mirabilis is an opportunistic pathogen that is frequently associated with urinary tract infections. In the lab, P. mirabilis cells become long and multinucleate and increase their number of flagella as they colonize agar surfaces during swarming. Swarming has been implicated in pathogenesis; however, it is unclear how energetically costly changes in P. mirabilis cell morphology translate into an advantage for adapting to environmental changes. We investigated two morphological changes that occur during swarming—increases in cell length and flagellum density—and discovered that an increase in the surface density of flagella enabled cells to translate rapidly through fluids of increasing viscosity; in contrast, cell length had a small effect on motility. We found that swarm cells had a surface density of flagella that was ∼5 times larger than that of vegetative cells and were motile in fluids with a viscosity that inhibits vegetative cell motility. To test the relationship between flagellum density and velocity, we overexpressed FlhD4C2, the master regulator of the flagellar operon, in vegetative cells of P. mirabilis and found that increased flagellum density produced an increase in cell velocity. Our results establish a relationship between P. mirabilis flagellum density and cell motility in viscous environments that may be relevant to its adaptation during the infection of mammalian urinary tracts and movement in contact with indwelling catheters. PMID:23144253

  10. Neonatal seizures induced by pentylenetetrazol or kainic acid disrupt primary cilia growth on developing mouse cortical neurons.

    PubMed

    Parker, Alexander K; Le, Megan M; Smith, Tyler S; Hoang-Minh, Lan B; Atkinson, Eric W; Ugartemendia, George; Semple-Rowland, Susan; Coleman, Jason E; Sarkisian, Matthew R

    2016-08-01

    Neonatal or early-life seizures (ELS) are often associated with life-long neurophysiological, cognitive and behavioral deficits, but the underlying mechanisms contributing to these deficits remain poorly understood. Newborn, post-migratory cortical neurons sprout ciliary buds (procilia) that mature into primary cilia. Disruption of the growth or signaling capabilities of these cilia has been linked to atypical neurite outgrowth from neurons and abnormalities in neuronal circuitry. Here, we tested the hypothesis that generalized seizures induced by pentylenetetrazol (PTZ) or kainic acid (KA) during early postnatal development impair neuronal and/or glial ciliogenesis. Mice received PTZ (50 or 100mg/kg), KA (2mg/kg), or saline either once at birth (P0), or once daily from P0 to P4. Using immunohistochemistry and electron microscopy, the cilia of neurons and glia were examined at P7, P14, and P42. A total of 83 regions were analyzed, representing 13 unique neocortical and hippocampal regions. Neuronal cilia were identified by co-expression of NeuN and type 3 adenylyl cyclase (ACIII) or somatostatin receptor 3 (SSTR3), while glial cilia were identified by co-expression of GFAP, Arl13b, and gamma-tubulin. We found that PTZ exposure at either P0 or from P0 to P4 induced convulsive behavior, followed by acute and lasting effects on neuronal cilia lengths that varied depending on the cortical region, PTZ dose, injection frequency, and time post-PTZ. Both increases and decreases in neuronal cilia length were observed. No changes in the length of glial cilia were observed under any of the test conditions. Lastly, we found that a single KA seizure at P0 led to similar abnormalities in neuronal cilia lengths. Our results suggest that seizure(s) occurring during early stages of cortical development induce persistent and widespread changes in neuronal cilia length. Given the impact neuronal cilia have on neuronal differentiation, ELS-induced changes in ciliogenesis may

  11. ATPases, ion exchangers and human sperm motility.

    PubMed

    Peralta-Arias, Rubén D; Vívenes, Carmen Y; Camejo, María I; Piñero, Sandy; Proverbio, Teresa; Martínez, Elizabeth; Marín, Reinaldo; Proverbio, Fulgencio

    2015-05-01

    Human sperm has several mechanisms to control its ionic milieu, such as the Na,K-ATPase (NKA), the Ca-ATPase of the plasma membrane (PMCA), the Na(+)/Ca(2) (+)-exchanger (NCX) and the Na(+)/H(+)-exchanger (NHE). On the other hand, the dynein-ATPase is the intracellular motor for sperm motility. In this work, we evaluated NKA, PMCA, NHE, NCX and dynein-ATPase activities in human sperm and investigated their correlation with sperm motility. Sperm motility was measured by Computer Assisted Semen Analysis. It was found that the NKA activity is inhibited by ouabain with two Ki (7.9 × 10(-9) and 9.8 × 10(-5) M), which is consistent with the presence of two isoforms of α subunit of the NKA in the sperm plasma membranes (α1 and α4), being α4 more sensitive to ouabain. The decrease in NKA activity is associated with a reduction in sperm motility. In addition, sperm motility was evaluated in the presence of known inhibitors of NHE, PMCA and NCX, such as amiloride, eosin, and KB-R7943, respectively, as well as in the presence of nigericin after incubation with ouabain. Amiloride, eosin and KB-R7943 significantly reduced sperm motility. Nigericin reversed the effect of ouabain and amiloride on sperm motility. Dynein-ATPase activity was inhibited by acidic pH and micromolar concentrations of Ca(2) (+). We explain our results in terms of inhibition of the dynein-ATPase in the presence of higher cytosolic H(+) and Ca(2) (+), and therefore inhibition of sperm motility. PMID:25820902

  12. 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

  13. A continuous roll-pulling approach for the fabrication of magnetic artificial cilia with microfluidic pumping capability.

    PubMed

    Wang, Ye; den Toonder, Jaap; Cardinaels, Ruth; Anderson, Patrick

    2016-06-21

    Magnetic artificial cilia are micro-hairs covering a surface that can be actuated using a time-dependent magnetic field to pump or mix fluids in microfluidic devices. This paper presents a novel fabrication method to realize magnetic artificial cilia using a roll-pulling process, in which a cylinder decorated with micro-pillars rolls over a liquid precursor film that contains magnetic particles at a speed up to 1 m s(-1), while a magnetic field is applied. Due to the interaction between the pillars and the liquid film, micro-hairs are pulled out of the film. In this way, surfaces with slender cone-shaped magnetic artificial cilia were produced. When integrated in a closed-loop channel, the artificial cilia were shown to be capable of generating substantial microfluidic pumping using external magnetic actuation. The spatial arrangement of the cilia can be varied by altering the layout of the micro-pillars on the roll surface. In addition, the final geometry of the individual cilia depends on the rheological properties of the precursor material in combination with the processing parameters of the roll-pulling process. A rheological study and fabrication tests were carried out for a range of precursor material compositions to obtain insight into the relation between precursor rheology and processing conditions on the one hand, and cilia geometry on the other hand. The development of this cleanroom-free, high speed and potentially large area method of production of artificial cilia is another step towards their implementation in real-life applications. PMID:27210071

  14. Novel mechanisms power bacterial gliding motility.

    PubMed

    Nan, Beiyan; Zusman, David R

    2016-07-01

    For many bacteria, motility is essential for survival, growth, virulence, biofilm formation and intra/interspecies interactions. Since natural environments differ, bacteria have evolved remarkable motility systems to adapt, including swimming in aqueous media, and swarming, twitching and gliding on solid and semi-solid surfaces. Although tremendous advances have been achieved in understanding swimming and swarming motilities powered by flagella, and twitching motility powered by Type IV pili, little is known about gliding motility. Bacterial gliders are a heterogeneous group containing diverse bacteria that utilize surface motilities that do not depend on traditional flagella or pili, but are powered by mechanisms that are less well understood. Recently, advances in our understanding of the molecular machineries for several gliding bacteria revealed the roles of modified ion channels, secretion systems and unique machinery for surface movements. These novel mechanisms provide rich source materials for studying the function and evolution of complex microbial nanomachines. In this review, we summarize recent findings made on the gliding mechanisms of the myxobacteria, flavobacteria and mycoplasmas. PMID:27028358

  15. Ghrelin family of peptides and gut motility.

    PubMed

    Asakawa, Akihiro; Ataka, Koji; Fujino, Kazunori; Chen, Chih-Yen; Kato, Ikuo; Fujimiya, Mineko; Inui, Akio

    2011-04-01

    Acyl ghrelin, des-acyl ghrelin, and obestatin are three peptides isolated from the gastrointestinal tract and encoded by the same preproghrelin gene. Three ghrelin gene products participate in modulating appetite, adipogenesis, glucose metabolism, cell proliferation, immune, sleep, memory, anxiety, cognition, and stress. We have investigated the effects of ghrelin family of peptides on fed and fasted motor activities in the stomach and duodenum of freely moving conscious rats by manometric method. Intracerebroventricular (ICV) and intravenous (IV) administration of acyl ghrelin induced fasted motor activity in the duodenum in fed rats. ICV and IV administration of des-acyl ghrelin disrupted fasted motor activity in the antrum. Changes in gastric motility induced by IV administration of des-acyl ghrelin were antagonized by ICV administration of a corticotropin-releasing factor (CRF) 2 receptor antagonist. IV administration of obestatin decreased the percentage motor index in the antrum and prolonged the time taken to return to fasted motility in the duodenum in fed rats. ICV administration of CRF 1 and 2 receptor antagonists prevented the effects of obestatin on gastroduodenal motility. Ghrelin gene products regulate feeding-associated gastroduodenal motility. Stomach may regulate various functions including gastrointestinal motility via acyl ghrelin, des-acyl ghrelin and obestatin as an endocrine organ. Increasing knowledge of the effects of ghrelin family of peptides on gastrointestinal motility could lead to innovative new therapies for functional gastrointestinal disorders. PMID:21443714

  16. Active stochastic stress fluctuations in the cell cytoskeleton stir the cell and activate primary cilia

    NASA Astrophysics Data System (ADS)

    Schmidt, Christoph F.; Fakhri, Nikta; Battle, Christopher; Ott, Carolyn M.; Wessel, Alok D.; Lippincott-Schwartz, Jennifer; Mackintosh, Frederick C.

    2015-03-01

    Cells are active systems with molecular force generation that drives complex dynamics at the supramolecular scale. Much of cellular dynamics is driven by myosin motors interacting with the actin cytoskeleton. We discovered active random ``stirring'' driven by cytoplasmic myosin as an intermediate mode of transport, different from both thermal diffusion and directed motor activity. We found a further manifestation of cytoskeletal dynamics in the active motion patterns of primary cilia generated by epithelial cells. These cilia were thought to be immotile due to the absence of dynein motors, but it turns out that their anchoring deeper inside the cell in combination with the strongly fluctuating cortex results in clearly measurable non-equilibrium fluctuations.

  17. Ca2+-dependent regulation of beat frequency of cilia in Paramecium.

    PubMed

    Nakaoka, Y; Tanaka, H; Oosawa, F

    1984-01-01

    Triton-extracted models of Paramecium cells prepared in the presence of Mg2+ and EGTA showed Ca2+ sensitivity not only in the direction of beat but also in the beat frequency of cilia. The beat frequency increased over a range of concentration of Ca2+ from 2 X 10(-7) to 4 X 10(-7) M, in which the model swam forwards. The frequency increased also above 10(-6)M-Ca2+, in which the model swam backwards. The increase in frequency was inhibited by calmodulin antagonists. Intracellular injection of a Ca2+ buffer giving a free Ca2+ concentration of 2 X 10(-7) to 5 X 10(-7) M in intact cells induced an increase in the beat frequency. Therefore, it is very likely that the beat frequency of cilia is regulated by the intracellular concentration of Ca2+. PMID:6715425

  18. 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

  19. Cilia-associated bacteria in fatal Bordetella bronchiseptica pneumonia of dogs and cats.

    PubMed

    Taha-Abdelaziz, Khaled; Bassel, Laura L; Harness, Melanie L; Clark, Mary Ellen; Register, Karen B; Caswell, Jeff L

    2016-07-01

    Bordetella bronchiseptica frequently causes nonfatal tracheobronchitis, but its role in fatal pneumonia is less recognized. Our study evaluated histologic identification of cilia-associated bacteria as a method for diagnosis of B. bronchiseptica pneumonia. Cases of fatal bronchopneumonia were studied retrospectively, excluding neonates and cases of aspiration pneumonia, minor lung lesions, or autolysis. The study population comprised 36 canine and 31 feline cases of bronchopneumonia. B. bronchiseptica was identified in 8 of 36 canine and 14 of 31 feline cases based on immunohistochemistry (IHC) using serum from a rabbit hyperimmunized with pertactin, PCR testing (Fla2/Fla12), and/or bacterial culture data when available. Of these, IHC was positive in 4 canine and 7 feline cases, PCR was positive in 8 canine and 14 feline cases, and B. bronchiseptica was isolated in 2 of 5 canine and 3 of 9 feline cases tested. Examination of histologic sections stained with hematoxylin and eosin revealed bronchial cilia-associated bacteria in 4 of 36 canine and 5 of 31 feline cases; these were all positive by IHC and PCR. The presence of cilia-associated bacteria had been noted in the pathology report for only 2 of these 9 cases. Thus, the presence of cilia-associated bacteria seems frequently overlooked by pathologists, but is a diagnostically significant feature of B. bronchiseptica pneumonia. A specific diagnosis of B. bronchiseptica pneumonia is important because it suggests primary or opportunistic bacterial pneumonia rather than aspiration pneumonia, and because of the risk of animal-to-animal transmission of B. bronchiseptica, the availability of vaccines for disease prevention, and the potential zoonotic risk to immunocompromised pet owners. PMID:27178716

  20. Hsp40 is involved in cilia regeneration in sea urchin embryos.

    PubMed

    Casano, Caterina; Gianguzza, Fabrizio; Roccheri, Maria C; Di Giorgi, Rossana; Maenza, Luigia; Ragusa, Maria A

    2003-12-01

    In a previous paper we demonstrated that, in Paracentrotus lividus embryos, deciliation represents a specific kind of stress that induces an increase in the levels of an acidic protein of about 40 kD (p40). Here we report that deciliation also induces an increase in Hsp40 chaperone levels and enhancement of its ectodermal localization. We suggest that Hsp40 might play a chaperoning role in cilia regeneration. PMID:14623926

  1. Global genetic analysis in mice unveils central role for cilia in congenital heart disease.

    PubMed

    Li, You; Klena, Nikolai T; Gabriel, George C; Liu, Xiaoqin; Kim, Andrew J; Lemke, Kristi; Chen, Yu; Chatterjee, Bishwanath; Devine, William; Damerla, Rama Rao; Chang, Chienfu; Yagi, Hisato; San Agustin, Jovenal T; Thahir, Mohamed; Anderton, Shane; Lawhead, Caroline; Vescovi, Anita; Pratt, Herbert; Morgan, Judy; Haynes, Leslie; Smith, Cynthia L; Eppig, Janan T; Reinholdt, Laura; Francis, Richard; Leatherbury, Linda; Ganapathiraju, Madhavi K; Tobita, Kimimasa; Pazour, Gregory J; Lo, Cecilia W

    2015-05-28

    Congenital heart disease (CHD) is the most prevalent birth defect, affecting nearly 1% of live births; the incidence of CHD is up to tenfold higher in human fetuses. A genetic contribution is strongly suggested by the association of CHD with chromosome abnormalities and high recurrence risk. Here we report findings from a recessive forward genetic screen in fetal mice, showing that cilia and cilia-transduced cell signalling have important roles in the pathogenesis of CHD. The cilium is an evolutionarily conserved organelle projecting from the cell surface with essential roles in diverse cellular processes. Using echocardiography, we ultrasound scanned 87,355 chemically mutagenized C57BL/6J fetal mice and recovered 218 CHD mouse models. Whole-exome sequencing identified 91 recessive CHD mutations in 61 genes. This included 34 cilia-related genes, 16 genes involved in cilia-transduced cell signalling, and 10 genes regulating vesicular trafficking, a pathway important for ciliogenesis and cell signalling. Surprisingly, many CHD genes encoded interacting proteins, suggesting that an interactome protein network may provide a larger genomic context for CHD pathogenesis. These findings provide novel insights into the potential Mendelian genetic contribution to CHD in the fetal population, a segment of the human population not well studied. We note that the pathways identified show overlap with CHD candidate genes recovered in CHD patients, suggesting that they may have relevance to the more complex genetics of CHD overall. These CHD mouse models and >8,000 incidental mutations have been sperm archived, creating a rich public resource for human disease modelling. PMID:25807483

  2. Primary cilia in stem cells and neural progenitors are regulated by neutral sphingomyelinase 2 and ceramide

    PubMed Central

    He, Qian; Wang, Guanghu; Wakade, Sushama; Dasgupta, Somsankar; Dinkins, Michael; Kong, Ji Na; Spassieva, Stefka D.; Bieberich, Erhard

    2014-01-01

    We show here that human embryonic stem (ES) and induced pluripotent stem cell–derived neuroprogenitors (NPs) develop primary cilia. Ciliogenesis depends on the sphingolipid ceramide and its interaction with atypical PKC (aPKC), both of which distribute to the primary cilium and the apicolateral cell membrane in NP rosettes. Neural differentiation of human ES cells to NPs is concurrent with a threefold elevation of ceramide—in particular, saturated, long-chain C16:0 ceramide (N-palmitoyl sphingosine) and nonsaturated, very long chain C24:1 ceramide (N-nervonoyl sphingosine). Decreasing ceramide levels by inhibiting ceramide synthase or neutral sphingomyelinase 2 leads to translocation of membrane-bound aPKC to the cytosol, concurrent with its activation and the phosphorylation of its substrate Aurora kinase A (AurA). Inhibition of aPKC, AurA, or a downstream target of AurA, HDAC6, restores ciliogenesis in ceramide-depleted cells. Of importance, addition of exogenous C24:1 ceramide reestablishes membrane association of aPKC, restores primary cilia, and accelerates neural process formation. Taken together, these results suggest that ceramide prevents activation of HDAC6 by cytosolic aPKC and AurA, which promotes acetylation of tubulin in primary cilia and, potentially, neural processes. This is the first report on the critical role of ceramide generated by nSMase2 in stem cell ciliogenesis and differentiation. PMID:24694597

  3. Cilia-Associated Genes Play Differing Roles in Aminoglycoside-Induced Hair Cell Death in Zebrafish

    PubMed Central

    Stawicki, Tamara M.; Hernandez, Liana; Esterberg, Robert; Linbo, Tor; Owens, Kelly N.; Shah, Arish N.; Thapa, Nihal; Roberts, Brock; Moens, Cecilia B.; Rubel, Edwin W.; Raible, David W.

    2016-01-01

    Hair cells possess a single primary cilium, called the kinocilium, early in development. While the kinocilium is lost in auditory hair cells of most species it is maintained in vestibular hair cells. It has generally been believed that the primary role of the kinocilium and cilia-associated genes in hair cells is in the establishment of the polarity of actin-based stereocilia, the hair cell mechanotransduction apparatus. Through genetic screening and testing of candidate genes in zebrafish (Danio rerio) we have found that mutations in multiple cilia genes implicated in intraflagellar transport (dync2h1, wdr35, ift88, and traf3ip), and the ciliary transition zone (cc2d2a, mks1, and cep290) lead to resistance to aminoglycoside-induced hair cell death. These genes appear to have differing roles in hair cells, as mutations in intraflagellar transport genes, but not transition zone genes, lead to defects in kinocilia formation and processes dependent upon hair cell mechanotransduction activity. These mutants highlight a novel role of cilia-associated genes in hair cells, and provide powerful tools for further study. PMID:27207957

  4. A Computational Model of Dynein Activation Patterns that Can Explain Nodal Cilia Rotation

    PubMed Central

    Chen, Duanduan; Zhong, Yi

    2015-01-01

    Normal left-right patterning in vertebrates depends on the rotational movement of nodal cilia. In order to produce this ciliary motion, the activity of axonemal dyneins must be tightly regulated in a temporal and spatial manner; the specific activation pattern of the dynein motors in the nodal cilia has not been reported. Contemporary imaging techniques cannot directly assess dynein activity in a living cilium. In this study, we establish a three-dimensional model to mimic the ciliary ultrastructure and assume that the activation of dynein proteins is related to the interdoublet distance. By employing finite-element analysis and grid deformation techniques, we simulate the mechanical function of dyneins by pairs of point loads, investigate the time-variant interdoublet distance, and simulate the dynein-triggered ciliary motion. The computational results indicate that, to produce the rotational movement of nodal cilia, the dynein activity is transferred clockwise (looking from the tip) between the nine doublet microtubules, and along each microtubule, the dynein activation should occur faster at the basal region and slower when it is close to the ciliary tip. Moreover, the time cost by all the dyneins along one microtubule to be activated can be used to deduce the dynein activation pattern; it implies that, as an alternative method, measuring this time can indirectly reveal the dynein activity. The proposed protein-structure model can simulate the ciliary motion triggered by various dynein activation patterns explicitly and may contribute to furthering the studies on axonemal dynein activity. PMID:26153700

  5. Endocytic recycling protein EHD1 regulates primary cilia morphogenesis and SHH signaling during neural tube development

    PubMed Central

    Bhattacharyya, Sohinee; Rainey, Mark A; Arya, Priyanka; Dutta, Samikshan; George, Manju; Storck, Matthew D.; McComb, Rodney D.; Muirhead, David; Todd, Gordon L.; Gould, Karen; Datta, Kaustubh; Waes, Janee Gelineau-van; Band, Vimla; Band, Hamid

    2016-01-01

    Members of the four-member C-terminal EPS15-Homology Domain-containing (EHD) protein family play crucial roles in endocytic recycling of cell surface receptors from endosomes to the plasma membrane. In this study, we show that Ehd1 gene knockout in mice on a predominantly B6 background is embryonic lethal. Ehd1-null embryos die at mid-gestation with a failure to complete key developmental processes including neural tube closure, axial turning and patterning of the neural tube. We found that Ehd1-null embryos display short and stubby cilia on the developing neuroepithelium at embryonic day 9.5 (E9.5). Loss of EHD1 also deregulates the ciliary SHH signaling with Ehd1-null embryos displaying features indicative of increased SHH signaling, including a significant downregulation in the formation of the GLI3 repressor and increase in the ventral neuronal markers specified by SHH. Using Ehd1-null MEFS we found that EHD1 protein co-localizes with the SHH receptor Smoothened in the primary cilia upon ligand stimulation. Under the same conditions, EHD1 was shown to co-traffic with Smoothened into the developing primary cilia and we identify EHD1 as a direct binding partner of Smoothened. Overall, our studies identify the endocytic recycling regulator EHD1 as a novel regulator of the primary cilium-associated trafficking of Smoothened and Hedgehog signaling. PMID:26884322

  6. Congenital Heart Disease Genetics Uncovers Context-Dependent Organization and Function of Nucleoporins at Cilia.

    PubMed

    Del Viso, Florencia; Huang, Fang; Myers, Jordan; Chalfant, Madeleine; Zhang, Yongdeng; Reza, Nooreen; Bewersdorf, Joerg; Lusk, C Patrick; Khokha, Mustafa K

    2016-09-12

    Human genomics is identifying candidate genes for congenital heart disease (CHD), but discovering the underlying mechanisms remains challenging. In a patient with CHD and heterotaxy (Htx), a disorder of left-right patterning, we previously identified a duplication in Nup188. However, a mechanism to explain how a component of the nuclear pore complex (NPC) could cause Htx/CHD was undefined. Here, we show that knockdown of Nup188 or its binding partner Nup93 leads to a loss of cilia during embryonic development while leaving NPC function largely intact. Many data, including the localization of endogenous Nup188/93 at cilia bases, support their direct role at cilia. Super-resolution imaging of Nup188 shows two barrel-like structures with dimensions and organization incompatible with an NPC-like ring, arguing against a proposed "ciliary pore complex." We suggest that the nanoscale organization and function of nucleoporins are context dependent in a way that is required for the structure of the heart. PMID:27593162

  7. Pulsed electromagnetic fields promote osteoblast mineralization and maturation needing the existence of primary cilia.

    PubMed

    Yan, Juan-Li; Zhou, Jian; Ma, Hui-Ping; Ma, Xiao-Ni; Gao, Yu-Hai; Shi, Wen-Gui; Fang, Qing-Qing; Ren, Qian; Xian, Cory J; Chen, Ke-Ming

    2015-03-15

    Although pulsed electromagnetic fields (PEMFs) have been approved as a therapy for osteoporosis, action mechanisms and optimal parameters are elusive. To determine the optimal intensity, exposure effects of 50 Hz PEMFs of 0.6-3.6 mT (0.6 interval at 90 min/day) were investigated on proliferation and osteogenic differentiation of cultured calvarial osteoblasts. All intensity groups stimulated proliferation significantly with the highest effect at 0.6 mT. The 0.6 mT group also obtained the optimal osteogenic effect as demonstrated by the highest ALP activity, ALP(+) CFU-f colony formation, nodule mineralization, and expression of COL-1 and BMP-2. To verify our hypothesis that the primary cilia are the cellular sensors for PEMFs, osteoblasts were also transfected with IFT88 siRNA or scrambled control, and osteogenesis-promoting effects of 0.6 mT PEMFs were found abrogated when primary cilia were inhibited by IFT88 siRNA. Thus primary cilia of osteoblasts play an indispensable role in mediating PEMF osteogenic effect in vitro. PMID:25661534

  8. Cilia-Associated Genes Play Differing Roles in Aminoglycoside-Induced Hair Cell Death in Zebrafish.

    PubMed

    Stawicki, Tamara M; Hernandez, Liana; Esterberg, Robert; Linbo, Tor; Owens, Kelly N; Shah, Arish N; Thapa, Nihal; Roberts, Brock; Moens, Cecilia B; Rubel, Edwin W; Raible, David W

    2016-01-01

    Hair cells possess a single primary cilium, called the kinocilium, early in development. While the kinocilium is lost in auditory hair cells of most species it is maintained in vestibular hair cells. It has generally been believed that the primary role of the kinocilium and cilia-associated genes in hair cells is in the establishment of the polarity of actin-based stereocilia, the hair cell mechanotransduction apparatus. Through genetic screening and testing of candidate genes in zebrafish (Danio rerio) we have found that mutations in multiple cilia genes implicated in intraflagellar transport (dync2h1, wdr35, ift88, and traf3ip), and the ciliary transition zone (cc2d2a, mks1, and cep290) lead to resistance to aminoglycoside-induced hair cell death. These genes appear to have differing roles in hair cells, as mutations in intraflagellar transport genes, but not transition zone genes, lead to defects in kinocilia formation and processes dependent upon hair cell mechanotransduction activity. These mutants highlight a novel role of cilia-associated genes in hair cells, and provide powerful tools for further study. PMID:27207957

  9. The intraflagellar transport protein IFT80 is required for cilia formation and osteogenesis

    PubMed Central

    Yang, Shuying; Wang, Changdong

    2012-01-01

    Intraflagellar transport (IFT) proteins are essential for the assembly and maintenance of cilia, which play important roles in development and homeostasis. IFT80 is a newly defined IFT protein. Partial mutation of IFT80 in humans causes diseases such as Jeune asphyxiating thoracic dystrophy (JATD) and short rib polydactyly (SRP) type III with abnormal skeletal development. However, the role and mechanism of IFT80 in osteogenesis is unknown. Here, we first detected IFT80 expression pattern and found that IFT80 was highly expressed in mouse long bone, skull, and during osteoblast differentiation. By using lentivirus-mediated RNA interference (RNAi) technology to silence IFT80 in murine mesenchymal progenitor cell line-C3H10T1/2 and bone marrow derived stromal cells, we found that silencing IFT80 led to either shortening or loss of cilia and the decrease of Arl13b expression - a small GTPase that is localized in cilia. Additionally, silencing IFT80 blocked the expression of osteoblast markers and significantly inhibited ALP activity and cell mineralization. We further found that IFT80 silencing inhibited the expression of Gli2, a critical transcriptional factor in the hedgehog signaling pathway. Overexpression of Gli2 rescued the deficiency of osteoblast differentiation from IFT80-silenced cells, and dramatically promoted osteoblast differentiation. Moreover, introduction of Smo agonist (SAG) promotes osteoblast differentiation, which was partially inhibited by IFT80 silencing. Thus, these results suggested that IFT80 plays an important role in osteogenesis through regulating Hedgehog/Gli signal pathways. PMID:22771375

  10. Statistical physical models of cellular motility

    NASA Astrophysics Data System (ADS)

    Banigan, Edward J.

    Cellular motility is required for a wide range of biological behaviors and functions, and the topic poses a number of interesting physical questions. In this work, we construct and analyze models of various aspects of cellular motility using tools and ideas from statistical physics. We begin with a Brownian dynamics model for actin-polymerization-driven motility, which is responsible for cell crawling and "rocketing" motility of pathogens. Within this model, we explore the robustness of self-diffusiophoresis, which is a general mechanism of motility. Using this mechanism, an object such as a cell catalyzes a reaction that generates a steady-state concentration gradient that propels the object in a particular direction. We then apply these ideas to a model for depolymerization-driven motility during bacterial chromosome segregation. We find that depolymerization and protein-protein binding interactions alone are sufficient to robustly pull a chromosome, even against large loads. Next, we investigate how forces and kinetics interact during eukaryotic mitosis with a many-microtubule model. Microtubules exert forces on chromosomes, but since individual microtubules grow and shrink in a force-dependent way, these forces lead to bistable collective microtubule dynamics, which provides a mechanism for chromosome oscillations and microtubule-based tension sensing. Finally, we explore kinematic aspects of cell motility in the context of the immune system. We develop quantitative methods for analyzing cell migration statistics collected during imaging experiments. We find that during chronic infection in the brain, T cells run and pause stochastically, following the statistics of a generalized Levy walk. These statistics may contribute to immune function by mimicking an evolutionarily conserved efficient search strategy. Additionally, we find that naive T cells migrating in lymph nodes also obey non-Gaussian statistics. Altogether, our work demonstrates how physical

  11. Mammalian Septins Nomenclature

    PubMed Central

    Macara, Ian G.; Baldarelli, Richard; Field, Christine M.; Glotzer, Michael; Hayashi, Yasuhide; Hsu, Shu-Chan; Kennedy, Mary B.; Kinoshita, Makoto; Longtine, Mark; Low, Claudia; Maltais, Lois J.; McKenzie, Louise; Mitchison, Timothy J.; Nishikawa, Toru; Noda, Makoto; Petty, Elizabeth M.; Peifer, Mark; Pringle, John R.; Robinson, Phillip J.; Roth, Dagmar; Russell, S.E. Hilary; Stuhlmann, Heidi; Tanaka, Manami; Tanaka, Tomoo; Trimble, William S.; Ware, Jerry; Zeleznik-Le, Nancy J.; Zieger, Barbara

    2002-01-01

    There are 10 known mammalian septin genes, some of which produce multiple splice variants. The current nomenclature for the genes and gene products is very confusing, with several different names having been given to the same gene product and distinct names given to splice variants of the same gene. Moreover, some names are based on those of yeast or Drosophila septins that are not the closest homologues. Therefore, we suggest that the mammalian septin field adopt a common nomenclature system, based on that adopted by the Mouse Genomic Nomenclature Committee and accepted by the Human Genome Organization Gene Nomenclature Committee. The human and mouse septin genes will be named SEPT1–SEPT10 and Sept1–Sept10, respectively. Splice variants will be designated by an underscore followed by a lowercase “v” and a number, e.g., SEPT4_v1. PMID:12475938

  12. Mammalian sweet taste receptors.

    PubMed

    Nelson, G; Hoon, M A; Chandrashekar, J; Zhang, Y; Ryba, N J; Zuker, C S

    2001-08-10

    The sense of taste provides animals with valuable information about the quality and nutritional value of food. Previously, we identified a large family of mammalian taste receptors involved in bitter taste perception (the T2Rs). We now report the characterization of mammalian sweet taste receptors. First, transgenic rescue experiments prove that the Sac locus encodes T1R3, a member of the T1R family of candidate taste receptors. Second, using a heterologous expression system, we demonstrate that T1R2 and T1R3 combine to function as a sweet receptor, recognizing sweet-tasting molecules as diverse as sucrose, saccharin, dulcin, and acesulfame-K. Finally, we present a detailed analysis of the patterns of expression of T1Rs and T2Rs, thus providing a view of the representation of sweet and bitter taste at the periphery. PMID:11509186

  13. Cellular Motility--Experiments on Contractile and Motile Mechanisms in the Slime Mould, Physarum Polycephalum

    ERIC Educational Resources Information Center

    Holmes, R. P.; Stewart, P. R.

    1977-01-01

    Actin and myosin have now been demonstrated to be important constituents of many eukaryotic cells. Their role is primarily that of a contractile system underlying all aspects of cellular motility. Described here is a simple experimental system to demonstrate quantitatively aspects of motility and its regulation in a slime mold. (Author/MA)

  14. Non-essential role for cilia in coordinating precise alignment of lens fibres.

    PubMed

    Sugiyama, Yuki; Shelley, Elizabeth J; Yoder, Bradley K; Kozmik, Zbynek; May-Simera, Helen L; Beales, Philip L; Lovicu, Frank J; McAvoy, John W

    2016-02-01

    The primary cilium, a microtubule-based organelle found in most cells, is a centre for mechano-sensing fluid movement and cellular signalling, notably through the Hedgehog pathway. We recently found that each lens fibre cell has an apically situated primary cilium that is polarised to the side of the cell facing the anterior pole of the lens. The direction of polarity is similar in neighbouring cells so that in the global view, lens fibres exhibit planar cell polarity (PCP) along the equatorial-anterior polar axis. Ciliogenesis has been associated with the establishment of PCP, although the exact relationship between PCP and the role of cilia is still controversial. To test the hypothesis that the primary cilia have a role in coordinating the precise alignment/orientation of the fibre cells, IFT88, a key component of the intraflagellar transport (IFT) complex, was removed specifically from the lens at different developmental stages using several lens-specific Cre-expressing mouse lines (MLR10- and LR-Cre). Irrespective of which Cre-line was adopted, both demonstrated that in IFT88-depleted cells, the ciliary axoneme was absent or substantially shortened, confirming the disruption of primary cilia formation. However no obvious histological defects were detected even when IFT88 was removed from the lens placode as early as E9.5. Specifically, the lens fibres aligned/oriented towards the poles to form the characteristic Y-shaped sutures as normal. Consistent with this, in primary lens epithelial explants prepared from these conditional knockout mouse lenses, the basal bodies still showed polarised localisation at the apical surface of elongating cells upon FGF-induced fibre differentiation. We further investigated the lens phenotype in knockouts of Bardet-Biedl Syndrome (BBS) proteins 4 and 8, the components of the BBSome complex which modulate ciliary function. In these BBS4 and 8 knockout lenses, again we found the pattern of the anterior sutures formed by the

  15. Study of human sperm motility post cryopreservation

    PubMed Central

    Oberoi, Bhavni; Kumar, Sushil; Talwar, Pankaj

    2014-01-01

    Background Cryopreservation of spermatozoa is a widely used technique to preserve the fertility of males. It can also benefit the armed forces personnel who are to be sent for long recruitments, while leaving their families behind. This study, apart from studying the effects of freezing and thawing, reveals the effect of the post thaw interval on the motility of the human spermatozoa and thus widens the insemination window period. Methods A detailed semen analysis was carried out as per the WHO guidelines for 25 samples. The samples were then washed, analysed and frozen in liquid nitrogen. The semen samples were subsequently thawed and similarly analysed after 20 min and 40 min of thawing. This was then followed by statistical analysis of the comparative motilities. Results Motility of sperms is found to decrease after cryopreservation. However, the study revealed that after thawing a significant increase in the motility of the sperms was noted with the progression of time (p < 0.05). Conclusion By simulating conditions similar to the in vivo conditions for the post thaw semen samples, we can safely wait, confirm the parameters like motility and count, and then inseminate the samples instead of blindly inseminating them immediately after thawing. PMID:25382909

  16. Motility modes of the parasite Trypanosoma brucei

    NASA Astrophysics Data System (ADS)

    Temel, Fatma Zeynep; Qu, Zijie; McAllaster, Michael; de Graffenried, Christopher; Breuer, Kenneth

    2015-11-01

    The parasitic single-celled protozoan Trypanosoma brucei causes African Sleeping Sickness, which is a fatal disease in humans and animals that threatens more than 60 million people in 36 African countries. Cell motility plays a critical role in the developmental phases and dissemination of the parasite. Unlike many other motile cells such as bacteria Escherichia coli or Caulobacter crescentus, the flagellum of T. brucei is attached along the length of its awl-like body, producing a unique mode of motility that is not fully understood or characterized. Here, we report on the motility of T. brucei, which swims using its single flagellum employing both rotating and undulating propulsion modes. We tracked cells in real-time in three dimensions using fluorescent microscopy. Data obtained from experiments using both short-term tracking within the field of view and long-term tracking using a tracking microscope were analyzed. Motility modes and swimming speed were analyzed as functions of cell size, rotation rate and undulation pattern. Research supported by NSF.

  17. Local anesthetics inhibit kinesin motility and microtentacle protrusions in human epithelial and breast tumor cells.

    PubMed

    Yoon, Jennifer R; Whipple, Rebecca A; Balzer, Eric M; Cho, Edward H; Matrone, Michael A; Peckham, Michelle; Martin, Stuart S

    2011-10-01

    Detached breast tumor cells produce dynamic microtubule protrusions that promote reattachment of cells and are termed tubulin microtentacles (McTNs) due to their mechanistic distinctions from actin-based filopodia/invadopodia and tubulin-based cilia. McTNs are enriched with vimentin and detyrosinated α-tubulin, (Glu-tubulin). Evidence suggests that vimentin and Glu-tubulin are cross-linked by kinesin motor proteins. Using known kinesin inhibitors, Lidocaine and Tetracaine, the roles of kinesins in McTN formation and function were tested. Live-cell McTN counts, adhesion assays, immunofluorescence, and video microscopy were performed to visualize inhibitor effects on McTNs. Viability and apoptosis assays were used to confirm the non-toxicity of the inhibitors. Treatments of human non-tumorigenic mammary epithelial and breast tumor cells with Lidocaine or Tetracaine caused rapid collapse of vimentin filaments. Live-cell video microscopy demonstrated that Tetracaine reduces motility of intracellular GFP-kinesin and causes centripetal collapse of McTNs. Treatment with Tetracaine inhibited the extension of McTNs and their ability to promote tumor cell aggregation and reattachment. Lidocaine showed similar effects but to a lesser degree. Our current data support a model in which the inhibition of kinesin motor proteins by Tetracaine leads to the reductions in McTNs, and provides a novel mechanism for the ability of this anesthetic to decrease metastatic progression. PMID:21069453

  18. Optical trapping reveals propulsion forces, power generation and motility efficiency of the unicellular parasites Trypanosoma brucei brucei

    PubMed Central

    Stellamanns, Eric; Uppaluri, Sravanti; Hochstetter, Axel; Heddergott, Niko; Engstler, Markus; Pfohl, Thomas

    2014-01-01

    Unicellular parasites have developed sophisticated swimming mechanisms to survive in a wide range of environments. Cell motility of African trypanosomes, parasites responsible for fatal illness in humans and animals, is crucial both in the insect vector and the mammalian host. Using millisecond-scale imaging in a microfluidics platform along with a custom made optical trap, we are able to confine single cells to study trypanosome motility. From the trapping characteristics of the cells, we determine the propulsion force generated by cells with a single flagellum as well as of dividing trypanosomes with two fully developed flagella. Estimates of the dissipative energy and the power generation of single cells obtained from the motility patterns of the trypanosomes within the optical trap indicate that specific motility characteristics, in addition to locomotion, may be required for antibody clearance. Introducing a steerable second optical trap we could further measure the force, which is generated at the flagellar tip. Differences in the cellular structure of the trypanosomes are correlated with the trapping and motility characteristics and in consequence with their propulsion force, dissipative energy and power generation. PMID:25269514

  19. Optical trapping reveals propulsion forces, power generation and motility efficiency of the unicellular parasites Trypanosoma brucei brucei

    NASA Astrophysics Data System (ADS)

    Stellamanns, Eric; Uppaluri, Sravanti; Hochstetter, Axel; Heddergott, Niko; Engstler, Markus; Pfohl, Thomas

    2014-10-01

    Unicellular parasites have developed sophisticated swimming mechanisms to survive in a wide range of environments. Cell motility of African trypanosomes, parasites responsible for fatal illness in humans and animals, is crucial both in the insect vector and the mammalian host. Using millisecond-scale imaging in a microfluidics platform along with a custom made optical trap, we are able to confine single cells to study trypanosome motility. From the trapping characteristics of the cells, we determine the propulsion force generated by cells with a single flagellum as well as of dividing trypanosomes with two fully developed flagella. Estimates of the dissipative energy and the power generation of single cells obtained from the motility patterns of the trypanosomes within the optical trap indicate that specific motility characteristics, in addition to locomotion, may be required for antibody clearance. Introducing a steerable second optical trap we could further measure the force, which is generated at the flagellar tip. Differences in the cellular structure of the trypanosomes are correlated with the trapping and motility characteristics and in consequence with their propulsion force, dissipative energy and power generation.

  20. Trimebutine as a modulator of gastrointestinal motility.

    PubMed

    Lee, Hyun-Tai; Kim, Byung Joo

    2011-06-01

    Trimebutine has been used for treatment of both hypermotility and hypomotility disorders of the gastrointestinal (GI) tract, such as irritable bowel syndrome. In this issue, Tan et al. (2011) examined the concentration-dependent dual effects of trimebutine on colonic motility in guinea pig. The authors suggested that trimebutine attenuated colonic motility mainly through the inhibition of L-type Ca(2+) channels at higher concentrations, whereas, at lower concentrations, it depolarized membrane potentials by reducing BK(ca) currents, resulting in the enhancement of the muscle contractions. Trimebutine might be a plausible modulator of GI motility, which gives an insight in developing new prokinetic agents. Further studies to elucidate the effects of trimebutine on the interstitial cells of Cajal, the pacemaker in GI muscles would promote the therapeutic benefits as a GI modulator. PMID:21725804

  1. Direct Upstream Motility in Escherichia coli

    PubMed Central

    Kaya, Tolga; Koser, Hur

    2012-01-01

    We provide an experimental demonstration of positive rheotaxis (rapid and continuous upstream motility) in wild-type Escherichia coli freely swimming over a surface. This hydrodynamic phenomenon is dominant below a critical shear rate and robust against Brownian motion and cell tumbling. We deduce that individual bacteria entering a flow system can rapidly migrate upstream (>20 μm/s) much faster than a gradually advancing biofilm. Given a bacterial population with a distribution of sizes and swim speeds, local shear rate near the surface determines the dominant hydrodynamic mode for motility, i.e., circular or random trajectories for low shear rates, positive rheotaxis for moderate flow, and sideways swimming at higher shear rates. Faster swimmers can move upstream more rapidly and at higher shear rates, as expected. Interestingly, we also find on average that both swim speed and upstream motility are independent of cell aspect ratio. PMID:22500751

  2. Mechanism of shape determination in motile cells

    PubMed Central

    Keren, Kinneret; Pincus, Zachary; Allen, Greg M.; Barnhart, Erin L.; Marriott, Gerard; Mogilner, Alex; Theriot, Julie A.

    2010-01-01

    The shape of motile cells is determined by many dynamic processes spanning several orders of magnitude in space and time, from local polymerization of actin monomers at subsecond timescales to global, cell-scale geometry that may persist for hours. Understanding the mechanism of shape determination in cells has proved to be extremely challenging due to the numerous components involved and the complexity of their interactions. Here we harness the natural phenotypic variability in a large population of motile epithelial keratocytes from fish (Hypsophrys nicaraguensis) to reveal mechanisms of shape determination. We find that the cells inhabit a low-dimensional, highly correlated spectrum of possible functional states. We further show that a model of actin network treadmilling in an inextensible membrane bag can quantitatively recapitulate this spectrum and predict both cell shape and speed. Our model provides a simple biochemical and biophysical basis for the observed morphology and behaviour of motile cells. PMID:18497816

  3. Incidence of motile Aeromonas spp. in foods.

    PubMed

    Pin, C; Marín, M L; García, M L; Tormo, J; Selgas, M D; Casas, C

    1994-09-01

    A total of 80 food samples were purchased from local retail consumer shops and examined for the presence of motile Aeromonas spp. Of the food categories tested, poultry had the highest incidence, with 100% positive. This was followed by lamb samples, with 60% positive. Raw milk and cheese samples had very low incidence (20%). No motile Aeromonas spp. were found in pre-prepared salads. Shellfish, fish, pork and beef samples had incidences of 40%. Most of the strains isolated were Aeromonas hydrophila, and for most of the food categories, no Aeromonas caviae isolates were obtained. PMID:7873101

  4. Response of a Motile/Non-Motile Escherichia coli Front to Hydrodynamic excitations

    NASA Astrophysics Data System (ADS)

    Baabour, Magali; Douarche, Carine; Salin, Dominique

    2014-11-01

    In a recent study (Douarche et al. PRL 102, 198101 (2009)), it has been shown that the motility of Escherichia coli (E. coli) is highly correlated to the oxygen level in a minimal medium: bacteria swim as long as they are provided with oxygen but reversibly transit to a non-motile state when they lack of it. Hence, when oxygen diffuses into an anaerobic sample of non-motile bacteria, a propagating front delineates a region of motile bacteria where oxygen is present from a region of non-motile ones where the oxygen is still not present. To study the response of this front to hydrodynamics excitation, we use the same fluorescent E. coli bacterial solution in rectangular cross section glass cells open to air (oxygen) at one inlet. After bacteria have consumed the oxygen of the solution, the presence of the oxygen only at the open edge of the sample leads to the formation of an analogous stationary front between motile and non-motile bacteria. We follow the response of this front to hydrodynamics flows such as an oscillating Poiseuille flow or natural convection. We analyze both the macroscopic behavior (shape and width) of the front as well as the microscopic displacements of individual bacteria. The dispersive behavior of this bacterial front is compared to the one of equivalent. Collaboration between Laboratories FAST and LPS, Univ Paris Sud and CNRS.

  5. The Intraflagellar Transport Protein IFT27 Promotes BBSome Exit from Cilia through the GTPase ARL6/BBS3

    PubMed Central

    Liew, Gerald M.; Ye, Fan; Nager, Andrew R.; Murphy, J. Patrick; Lee, Jaclyn S.; Aguiar, Mike; Breslow, David K.; Gygi, Steven P.; Nachury, Maxence V.

    2014-01-01

    SUMMARY The sorting of signaling receptors into and out of cilia relies on the BBSome, a complex of Bardet-Biedl syndrome (BBS) proteins, and on the intraflagellar transport (IFT) machinery. GTP loading onto the Arf-like GTPase ARL6/BBS3 drives assembly of a membrane-apposed BBSome coat that promotes cargo entry into cilia, yet how and where ARL6 is activated remains elusive. Here, we show that the Rab-like GTPase IFT27/RABL4, a known component of IFT complex B, promotes the exit of BBSome and associated cargoes from cilia. Unbiased proteomics and biochemical reconstitution assays show that, upon disengagement from the rest of IFT-B, IFT27 directly interacts with the nucleotide-free form of ARL6. Furthermore, IFT27 prevents aggregation of nucleotide-free ARL6 in solution. Thus, we propose that IFT27 separates from IFT-B inside cilia to promote ARL6 activation, BBSome coat assembly and subsequent ciliary exit, mirroring the process by which BBSome mediates cargo entry into cilia. PMID:25443296

  6. Silibinin negatively contributes to primary cilia length via autophagy regulated by histone deacetylase 6 in confluent mouse embryo fibroblast 3T3-L1 cells.

    PubMed

    Xu, Qian; Liu, Wei; Liu, Xiaoling; Liu, Weiwei; Wang, Hongju; Yao, Guodong; Zang, Linghe; Hayashi, Toshihiko; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi

    2016-09-01

    Primary cilium is a cellular antenna, signalling as a sensory organelle. Numerous pathological manifestation is associated with change of its length. Although the interaction between autophagy and primary cilia has been suggested, the role of autophagy in primary cilia length is largely unknown. In this study the primary cilia were immunostained and observed by using confocal fluorescence microscopy, and we found that silibinin, a natural flavonoid, shortened the length of primary cilia, meanwhile it also induced autophagy in 3T3-L1 cells. This study was designed to investigate the significance of silibinin-induced autophagy in primary ciliary structure in confluent mouse embryo fibroblast 3T3-L1 cells. Either blocking the autophagic flux with pre-treatment with the autophagy inhibitor, 3-methyladenine (3-MA), or transfection of siRNA targeting LC3 inhibited the reduction of cilia length caused by silibinin exposure. Autophagy induced by silibinin decreased expressions of the cilia-associated proteins, such as IFT88, KIF3a and Ac-tubulin, while 3-MA restored it, indicating that autophagy induced by silibinin led to a reduction of primary cilia length. Histone deacetylase 6 (HDAC6), which was suggested as a mediator of autophagy, was up-regulated by silibinin in a time-dependent manner. In addition, 3T3-L1 cells treated with siRNA against HDAC6 had a reduced autophagic level and were protected from silibinin-induced cilia shortening. Taken together, we conclude that the HDAC6-mediated autophagy negatively regulates primary cilia length during silibinin treatment and has the potential to serve as a therapeutic target for primary cilia-associated ciliopathies. These findings thus provide new information about the potential link between autophagy and primary cilia. PMID:27435857

  7. Determination of motility forces on isolated chromosomes with laser tweezers

    PubMed Central

    Khatibzadeh, Nima; Stilgoe, Alexander B.; Bui, Ann A. M.; Rocha, Yesenia; Cruz, Gladys M.; Loke, Vince; Shi, Linda Z.; Nieminen, Timo A.; Rubinsztein-Dunlop, Halina; Berns, Michael W.

    2014-01-01

    Quantitative determination of the motility forces of chromosomes during cell division is fundamental to understanding a process that is universal among eukaryotic organisms. Using an optical tweezers system, isolated mammalian chromosomes were held in a 1064 nm laser trap. The minimum force required to move a single chromosome was determined to be ≈0.8–5 pN. The maximum transverse trapping efficiency of the isolated chromosomes was calculated as ≈0.01–0.02. These results confirm theoretical force calculations of ≈0.1–12 pN to move a chromosome on the mitotic or meiotic spindle. The verification of these results was carried out by calibration of the optical tweezers when trapping microspheres with a diameter of 4.5–15 µm in media with 1–7 cP viscosity. The results of the chromosome and microsphere trapping experiments agree with optical models developed to simulate trapping of cylindrical and spherical specimens. PMID:25359514

  8. Mammalian Endogenous Retroviruses.

    PubMed

    Mager, Dixie L; Stoye, Jonathan P

    2015-02-01

    Over 40% of mammalian genomes comprise the products of reverse transcription. Among such retrotransposed sequences are those characterized by the presence of long terminal repeats (LTRs), including the endogenous retroviruses (ERVs), which are inherited genetic elements closely resembling the proviruses formed following exogenous retrovirus infection. Sequences derived from ERVs make up at least 8 to 10% of the human and mouse genomes and range from ancient sequences that predate mammalian divergence to elements that are currently still active. In this chapter we describe the discovery, classification and origins of ERVs in mammals and consider cellular mechanisms that have evolved to control their expression. We also discuss the negative effects of ERVs as agents of genetic disease and cancer and review examples of ERV protein domestication to serve host functions, as in placental development. Finally, we address growing evidence that the gene regulatory potential of ERV LTRs has been exploited multiple times during evolution to regulate genes and gene networks. Thus, although recently endogenized retroviral elements are often pathogenic, those that survive the forces of negative selection become neutral components of the host genome or can be harnessed to serve beneficial roles. PMID:26104559

  9. Synthetic control of mammalian-cell motility by engineering chemotaxis to an orthogonal bioinert chemical signal

    PubMed Central

    Park, Jason S.; Rhau, Benjamin; Hermann, Aynur; McNally, Krista A.; Zhou, Carmen; Gong, Delquin; Weiner, Orion D.; Conklin, Bruce R.; Onuffer, James; Lim, Wendell A.

    2014-01-01

    Directed migration of diverse cell types plays a critical role in biological processes ranging from development and morphogenesis to immune response, wound healing, and regeneration. However, techniques to direct, manipulate, and study cell migration in vitro and in vivo in a specific and facile manner are currently limited. We conceived of a strategy to achieve direct control over cell migration to arbitrary user-defined locations, independent of native chemotaxis receptors. Here, we show that genetic modification of cells with an engineered G protein-coupled receptor allows us to redirect their migration to a bioinert drug-like small molecule, clozapine-N-oxide (CNO). The engineered receptor and small-molecule ligand form an orthogonal pair: The receptor does not respond to native ligands, and the inert drug does not bind to native cells. CNO-responsive migration can be engineered into a variety of cell types, including neutrophils, T lymphocytes, keratinocytes, and endothelial cells. The engineered cells migrate up a gradient of the drug CNO and transmigrate through endothelial monolayers. Finally, we demonstrate that T lymphocytes modified with the engineered receptor can specifically migrate in vivo to CNO-releasing beads implanted in a live mouse. This technology provides a generalizable genetic tool to systematically perturb and control cell migration both in vitro and in vivo. In the future, this type of migration control could be a valuable module for engineering therapeutic cellular devices. PMID:24711398

  10. Esophageal motility abnormalities in gastroesophageal reflux disease.

    PubMed

    Martinucci, Irene; de Bortoli, Nicola; Giacchino, Maria; Bodini, Giorgia; Marabotto, Elisa; Marchi, Santino; Savarino, Vincenzo; Savarino, Edoardo

    2014-05-01

    Esophageal motility abnormalities are among the main factors implicated in the pathogenesis of gastroesophageal reflux disease. The recent introduction in clinical and research practice of novel esophageal testing has markedly improved our understanding of the mechanisms contributing to the development of gastroesophageal reflux disease, allowing a better management of patients with this disorder. In this context, the present article intends to provide an overview of the current literature about esophageal motility dysfunctions in patients with gastroesophageal reflux disease. Esophageal manometry, by recording intraluminal pressure, represents the gold standard to diagnose esophageal motility abnormalities. In particular, using novel techniques, such as high resolution manometry with or without concurrent intraluminal impedance monitoring, transient lower esophageal sphincter (LES) relaxations, hypotensive LES, ineffective esophageal peristalsis and bolus transit abnormalities have been better defined and strongly implicated in gastroesophageal reflux disease development. Overall, recent findings suggest that esophageal motility abnormalities are increasingly prevalent with increasing severity of reflux disease, from non-erosive reflux disease to erosive reflux disease and Barrett's esophagus. Characterizing esophageal dysmotility among different subgroups of patients with reflux disease may represent a fundamental approach to properly diagnose these patients and, thus, to set up the best therapeutic management. Currently, surgery represents the only reliable way to restore the esophagogastric junction integrity and to reduce transient LES relaxations that are considered to be the predominant mechanism by which gastric contents can enter the esophagus. On that ground, more in depth future studies assessing the pathogenetic role of dysmotility in patients with reflux disease are warranted. PMID:24868489

  11. Actin motility: formin a SCAry tail.

    PubMed

    Alberts, Art; Way, Michael

    2011-01-11

    A new biochemical analysis has revealed that the Rickettsia bacterial protein Sca2--recently shown to be essential for virulence and actin-dependent motility--assembles actin filaments using a mechanism that functionally resembles the processive elongation tactics used by formins. PMID:21215933

  12. Semiautomated Motility Assay For Determining Toxicity

    NASA Technical Reports Server (NTRS)

    Noever, David A.; Cronise, Raymond

    1996-01-01

    Improved method of assessing toxicities of various substances based on observation of effects of those substances on motilities of manageably small number of cells of protozoan species Tetrahema pyriformis. Provides repeatable, standardized tests with minimal handling by technicians and with minimal exposure of technicians to chemicals. Rapid and economical alternative to Draize test.

  13. Targeting tumor cell motility to prevent metastasis

    PubMed Central

    Palmer, Trenis D.; Ashby, William J.; Lewis, John D.; Zijlstra, Andries

    2011-01-01

    Mortality and morbidity in patients with solid tumors invariably results from the disruption of normal biological function caused by disseminating tumor cells. Tumor cell migration is under intense investigation as the underlying cause of cancer metastasis. The need for tumor cell motility in the progression of metastasis has been established experimentally and is supported empirically by basic and clinical research implicating a large collection of migration-related genes. However, there are few clinical interventions designed to specifically target the motility of tumor cells and adjuvant therapy to specifically prevent cancer cell dissemination is severely limited. In an attempt to define motility targets suitable for treating metastasis, we have parsed the molecular determinants of tumor cell motility into five underlying principles including cell autonomous ability, soluble communication, cell-cell adhesion, cell-matrix adhesion, and integrating these determinants of migration on molecular scaffolds. The current challenge is to implement meaningful and sustainable inhibition of metastasis by developing clinically viable disruption of molecular targets that control these fundamental capabilities. PMID:21664937

  14. Primary cilia distribution and orientation during involution of the bovine mammary gland.

    PubMed

    Biet, J; Poole, C A; Stelwagen, K; Margerison, J K; Singh, K

    2016-05-01

    The regulation of mammary gland involution occurs through multiple levels including environmental factors, hormones, and local intramammary signals. Primary cilia (PC) are signaling organelles that sense biochemical and biophysical extracellular stimuli and are vital for cellular and tissue function. The aim of this study was to examine the distribution, incidence, and orientation of PC. Furthermore, we determined changes in expression levels of the signal transducer and activator of transcription (STAT)6 at the onset of bovine mammary gland involution. Mammary tissue was collected from pasture-fed, primiparous, nonpregnant Friesian dairy cows at mid lactation (n=5 per group) killed 6-h after milking (lactating controls) and during involution after 7 and 28 d of nonmilking (NM). Fluorescent immunohistochemistry and confocal microscopy of tissue sections showed that PC were present on luminal secretory epithelial cells (SEC), myoepithelial cells (MEC), and stromal fibroblast cells (SFC). Furthermore, in all 3 experimental groups, different PC positions or orientations relative to the cell surface were identified on SEC and MEC, which projected toward the lumen and were either straight, bent, or deflected against the apical cell surface, whereas PC in SFC were confined to the interalveolar space. However, by 28-d NM, fewer PC projected into the luminal space and most appeared deflected or projected toward the interalveolar space. Furthermore, by 28-d NM, with the increase in stromal connective tissue, more PC were detected within the interalveolar and interlobular stroma. At 28-d NM, we observed a decrease in luminal cilia relative to the total number of cilia. The number of ciliated cells in the total fraction (SEC, MEC, and SFC) was the same for all 3 groups, although in the luminal fraction (SEC and MEC), PC per nuclei increased by 28-d NM relative to lactation. At all 3 stages, we detected variations in shape and orientation of PC within the same alveolus, with

  15. Determining the Relative Contribution and Hierarchy of hha and qseBC in the Regulation of Flagellar Motility of Escherichia coli O157:H7

    PubMed Central

    Sharma, Vijay K.; Casey, Thomas A.

    2014-01-01

    In recent studies, we demonstrated that a deletion of hha caused increased secretion of locus of enterocyte encoded adherence proteins and reduced motility of enterohemorrhagic Escherichia coli (EHEC) O157:H7. In addition to the importance of hha in positive regulation of motility, a two-component quorum sensing pathway encoded by the qseBC genes has been shown to activate bacterial motility in response to mammalian stress hormones epinephrine and norepinephrine as well as bacterially produced autoinducer-3. In this study, we compared regulatory contribution and hierarchy of hha, a member of the Hha/YmoA family of nucleoid-associated proteins, to that of qseBC in the expression of EHEC O157:H7 motility. Since norepinephrine affects motility of EHEC O157:H7 through a qseBC-encoded two-component quorum sensing signaling, we also determined whether the hha-mediated regulation of motility is affected by norepinephrine and whether this effect is qseBC dependent. We used single (Δhha or ΔqseC) and double (Δhha ΔqseC) deletion mutants to show that hha exerts a greater positive regulatory effect in comparison to qseBC on the expression of motility by EHEC O157:H7. We also show that Hha is hierarchically superior in transcriptional regulation of motility than QseBC because transcription of qseC was significantly reduced in the hha deletion mutant compared to that in the parental and the hha-complemented mutant strains. These results suggest that hha regulates motility of EHEC O157:H7 directly as well as indirectly by controlling the transcription of qseBC. PMID:24465756

  16. Global genetic analysis in mice unveils central role for cilia in congenital heart disease

    PubMed Central

    Li, You; Klena, Nikolai T.; Gabriel, George C; Liu, Xiaoqin; Kim, Andrew J.; Lemke, Kristi; Chen, Yu; Chatterjee, Bishwanath; Devine, William; Damerla, Rama Rao; Chang, Chien-fu; Yagi, Hisato; San Agustin, Jovenal T.; Thahir, Mohamed; Anderton, Shane; Lawhead, Caroline; Vescovi, Anita; Pratt, Herbert; Morgan, Judy; Haynes, Leslie; Smith, Cynthia L.; Eppig, Janan T.; Reinholdt, Laura; Francis, Richard; Leatherbury, Linda; Ganapathiraju, Madhavi K.; Tobita, Kimimasa; Pazour, Gregory J.; Lo, Cecilia W.

    2015-01-01

    Congenital heart disease (CHD) is the most prevalent birth defect, affecting nearly 1% of live births1, but the incidence of CHD is up to ten fold higher in human fetuses2,3. A genetic contribution is strongly suggested by the association of CHD with chromosome abnormalities and high recurrence risk4. Here we report findings from a recessive forward genetic screen in fetal mice, showing the cilium and cilia transduced cell signaling play important roles in the pathogenesis of CHD. The cilium is an evolutionarily conserved organelle projecting from the cell surface with essential roles in diverse cellular processes. Using echocardiography, we ultrasound scanned 87,355 chemically mutagenized C57BL/6J fetal mice and recovered 218 CHD mouse models. Whole exome sequencing identified 91 recessive CHD mutations in 61 genes. This included 34 cilia-related genes, 16 genes involved in cilia transduced cell signaling, and 10 genes regulating vesicular trafficking, a pathway important for ciliogenesis and cell signaling. Surprisingly, many CHD genes encoded interacting proteins, suggesting an interactome protein network may provide a larger genomic context for CHD pathogenesis. These findings provide novel insights into the potential Mendelian genetic contribution to CHD in the fetal population, a segment of the human population not well studied. We note pathways identified show overlap with CHD candidate genes recovered in CHD patients5, suggesting they may have relevance to the more complex genetics of CHD overall. These CHD mouse models and >8,000 incidental mutations are sperm archived, creating a rich public resource for human disease modeling. PMID:25807483

  17. An animated model of reticulorumen motility.

    PubMed

    Gookin, Jody L; Foster, Derek M; Harvey, Alice M; McWhorter, Dan

    2009-01-01

    Understanding reticulorumen motility is important to the assessment of ruminant health and optimal production, and in the recognition, diagnosis, and treatment of disease. Accordingly, the teaching of reticulorumen motility is a staple of all veterinary curricula. This teaching has historically been based on written descriptions, line drawings, or pressure tracings obtained during contraction sequences. We developed an animated model of reticulorumen motility and hypothesized that veterinary students would prefer use of the model over traditional instructional methods. First-year veterinary students were randomly allocated to one of two online learning exercises: with the animated model (Group A) or with text and line drawings (Group B) depicting reticulorumen motility. Learning was assessed with a multiple-choice quiz and feedback on the learning alternatives was obtained by survey. Seventy-four students participated in the study, including 38/42 in Group A and 36/36 in Group B. Sixty-four out of 72 students (89%) responded that they would prefer use of the animated model if only one of the two learning methods was available. A majority of students agreed or strongly agreed that the animated model was easy to understand and improved their knowledge and appreciation of the importance of reticulorumen motility, and would recommend the model to other veterinary students. Interestingly, students in Group B achieved higher scores on examination than students in Group A. This could be speculatively attributed to the inclusion of an itemized list of contraction sequences in the text provided to Group B and failure of Group A students to read the text associated with the animations. PMID:20054084

  18. Maintenance of motility bias during cyanobacterial phototaxis.

    PubMed

    Chau, Rosanna Man Wah; Ursell, Tristan; Wang, Shuo; Huang, Kerwyn Casey; Bhaya, Devaki

    2015-04-01

    Signal transduction in bacteria is complex, ranging across scales from molecular signal detectors and effectors to cellular and community responses to stimuli. The unicellular, photosynthetic cyanobacterium Synechocystis sp. PCC6803 transduces a light stimulus into directional movement known as phototaxis. This response occurs via a biased random walk toward or away from a directional light source, which is sensed by intracellular photoreceptors and mediated by Type IV pili. It is unknown how quickly cells can respond to changes in the presence or directionality of light, or how photoreceptors affect single-cell motility behavior. In this study, we use time-lapse microscopy coupled with quantitative single-cell tracking to investigate the timescale of the cellular response to various light conditions and to characterize the contribution of the photoreceptor TaxD1 (PixJ1) to phototaxis. We first demonstrate that a community of cells exhibits both spatial and population heterogeneity in its phototactic response. We then show that individual cells respond within minutes to changes in light conditions, and that movement directionality is conferred only by the current light directionality, rather than by a long-term memory of previous conditions. Our measurements indicate that motility bias likely results from the polarization of pilus activity, yielding variable levels of movement in different directions. Experiments with a photoreceptor (taxD1) mutant suggest a supplementary role of TaxD1 in enhancing movement directionality, in addition to its previously identified role in promoting positive phototaxis. Motivated by the behavior of the taxD1 mutant, we demonstrate using a reaction-diffusion model that diffusion anisotropy is sufficient to produce the observed changes in the pattern of collective motility. Taken together, our results establish that single-cell tracking can be used to determine the factors that affect motility bias, which can then be coupled with

  19. Fibroblast Growth Factor Receptors (FGFRs) in Human Sperm: Expression, Functionality and Involvement in Motility Regulation

    PubMed Central

    Saucedo, Lucía; Buffa, Gabriela N.; Rosso, Marina; Guillardoy, Tomás; Góngora, Adrian; Munuce, María J.

    2015-01-01

    Fibroblast growth factors receptors (FGFRs) have been widely characterized in somatic cells, but there is scarce evidence of their expression and function in mammalian gametes. The objective of the present study was to evaluate the expression of FGFRs in human male germ cells, to determine sperm FGFR activation by the FGF2 ligand and their participation in the regulation of sperm motility. The expression of FGFR1, 2, 3 and 4 mRNAs and proteins in human testis and localization of these receptors in germ cells of the seminiferous epithelium was demonstrated. In ejaculated sperm, FGFRs were localized to the acrosomal region and flagellum. Sperm exposure to FGF2 caused an increase in flagellar FGFR phosphorylation and activation of extracellular signal-regulated kinase (ERK) and protein kinase B (PKB or Akt) signaling pathways. Incubation with FGF2 led to a significant increase in the percentage of total and progressive sperm motility, as well as in sperm kinematics. All responses were prevented by sperm preincubation with BGJ398, a specific inhibitor of FGFR tyrosine kinase activity. In addition to confirming the expression of FGFRs in germ cells of the human testis, our study describes for the first time the presence, localization and functionality of human sperm FGFRs, and provides evidence of the beneficial effect of FGF2 upon sperm motility. PMID:25970615

  20. Both contractile axial and lateral traction force dynamics drive amoeboid cell motility

    PubMed Central

    Bastounis, Effie; Meili, Ruedi; Álvarez-González, Begoña; Francois, Joshua; del Álamo, Juan C.; Lasheras, Juan C.

    2014-01-01

    Chemotaxing Dictyostelium discoideum cells adapt their morphology and migration speed in response to intrinsic and extrinsic cues. Using Fourier traction force microscopy, we measured the spatiotemporal evolution of shape and traction stresses and constructed traction tension kymographs to analyze cell motility as a function of the dynamics of the cell’s mechanically active traction adhesions. We show that wild-type cells migrate in a step-wise fashion, mainly forming stationary traction adhesions along their anterior–posterior axes and exerting strong contractile axial forces. We demonstrate that lateral forces are also important for motility, especially for migration on highly adhesive substrates. Analysis of two mutant strains lacking distinct actin cross-linkers (mhcA− and abp120− cells) on normal and highly adhesive substrates supports a key role for lateral contractions in amoeboid cell motility, whereas the differences in their traction adhesion dynamics suggest that these two strains use distinct mechanisms to achieve migration. Finally, we provide evidence that the above patterns of migration may be conserved in mammalian amoeboid cells. PMID:24637328

  1. Fibroblast Growth Factor Receptors (FGFRs) in Human Sperm: Expression, Functionality and Involvement in Motility Regulation.

    PubMed

    Saucedo, Lucía; Buffa, Gabriela N; Rosso, Marina; Guillardoy, Tomás; Góngora, Adrian; Munuce, María J; Vazquez-Levin, Mónica H; Marín-Briggiler, Clara

    2015-01-01

    Fibroblast growth factors receptors (FGFRs) have been widely characterized in somatic cells, but there is scarce evidence of their expression and function in mammalian gametes. The objective of the present study was to evaluate the expression of FGFRs in human male germ cells, to determine sperm FGFR activation by the FGF2 ligand and their participation in the regulation of sperm motility. The expression of FGFR1, 2, 3 and 4 mRNAs and proteins in human testis and localization of these receptors in germ cells of the seminiferous epithelium was demonstrated. In ejaculated sperm, FGFRs were localized to the acrosomal region and flagellum. Sperm exposure to FGF2 caused an increase in flagellar FGFR phosphorylation and activation of extracellular signal-regulated kinase (ERK) and protein kinase B (PKB or Akt) signaling pathways. Incubation with FGF2 led to a significant increase in the percentage of total and progressive sperm motility, as well as in sperm kinematics. All responses were prevented by sperm preincubation with BGJ398, a specific inhibitor of FGFR tyrosine kinase activity. In addition to confirming the expression of FGFRs in germ cells of the human testis, our study describes for the first time the presence, localization and functionality of human sperm FGFRs, and provides evidence of the beneficial effect of FGF2 upon sperm motility. PMID:25970615

  2. Influence of fluorescent tag on the motility properties of kinesin-1 in single-molecule assays.

    PubMed

    Norris, Stephen R; Núñez, Marcos F; Verhey, Kristen J

    2015-03-10

    Molecular motors such as kinesin and dynein use the energy derived from ATP hydrolysis to walk processively along microtubule tracks and transport various cargoes inside the cell. Recent advancements in fluorescent protein (FP) research enable motors to be fluorescently labeled such that single molecules can be visualized inside cells in multiple colors. The performance of these fluorescent tags can vary depending on their spectral properties and a natural tendency for oligomerization. Here we present a survey of different fluorescent tags fused to kinesin-1 and studied by single-molecule motility assays of mammalian cell lysates. We tested eight different FP tags and found that seven of them display sufficient fluorescence intensity and photostability to visualize motility events. Although none of the FP tags interfere with the enzymatic properties of the motor, four of the tags (EGFP, monomeric EGFP, tagRFPt, and mApple) cause aberrantly long motor run lengths. This behavior is unlikely to be due to electrostatic interactions and is probably caused by tag-dependent oligomerization events that appear to be facilitated by fusion to the dimeric kinesin-1. We also compared the single-molecule performance of various fluorescent SNAP and HALO ligands. We found that although both green and red SNAP ligands provide sufficient fluorescent signal, only the tetramethyl rhodamine (TMR) HALO ligand provides sufficient signal for detection in these assays. This study will serve as a valuable reference for choosing fluorescent labels for single-molecule motility assays. PMID:25762325

  3. Robust and stretchable indium gallium zinc oxide-based electronic textiles formed by cilia-assisted transfer printing.

    PubMed

    Yoon, Jongwon; Jeong, Yunkyung; Kim, Heeje; Yoo, Seonggwang; Jung, Hoon Sun; Kim, Yonghun; Hwang, Youngkyu; Hyun, Yujun; Hong, Woong-Ki; Lee, Byoung Hun; Choa, Sung-Hoon; Ko, Heung Cho

    2016-01-01

    Electronic textile (e-textile) allows for high-end wearable electronic devices that provide easy access for carrying, handling and using. However, the related technology does not seem to be mature because the woven fabric hampers not only the device fabrication process directly on the complex surface but also the transfer printing of ultrathin planar electronic devices. Here we report an indirect method that enables conformal wrapping of surface with arbitrary yet complex shapes. Artificial cilia are introduced in the periphery of electronic devices as adhesive elements. The cilia also play an important role in confining a small amount of glue and damping mechanical stress to maintain robust electronic performance under mechanical deformation. The example of electronic applications depicts the feasibility of cilia for 'stick-&-play' systems, which provide electronic functions by transfer printing on unconventional complex surfaces. PMID:27248982

  4. Robust and stretchable indium gallium zinc oxide-based electronic textiles formed by cilia-assisted transfer printing

    PubMed Central

    Yoon, Jongwon; Jeong, Yunkyung; Kim, Heeje; Yoo, Seonggwang; Jung, Hoon Sun; Kim, Yonghun; Hwang, Youngkyu; Hyun, Yujun; Hong, Woong-Ki; Lee, Byoung Hun; Choa, Sung-Hoon; Ko, Heung Cho

    2016-01-01

    Electronic textile (e-textile) allows for high-end wearable electronic devices that provide easy access for carrying, handling and using. However, the related technology does not seem to be mature because the woven fabric hampers not only the device fabrication process directly on the complex surface but also the transfer printing of ultrathin planar electronic devices. Here we report an indirect method that enables conformal wrapping of surface with arbitrary yet complex shapes. Artificial cilia are introduced in the periphery of electronic devices as adhesive elements. The cilia also play an important role in confining a small amount of glue and damping mechanical stress to maintain robust electronic performance under mechanical deformation. The example of electronic applications depicts the feasibility of cilia for ‘stick-&-play' systems, which provide electronic functions by transfer printing on unconventional complex surfaces. PMID:27248982

  5. Single-molecule imaging of Hedgehog pathway protein Smoothened in primary cilia reveals binding events regulated by Patched1

    PubMed Central

    Milenkovic, Ljiljana; Weiss, Lucien E.; Yoon, Joshua; Roth, Theodore L.; Su, YouRong S.; Sahl, Steffen J.; Scott, Matthew P.; Moerner, W. E.

    2015-01-01

    Accumulation of the signaling protein Smoothened (Smo) in the membrane of primary cilia is an essential step in Hedgehog (Hh) signal transduction, yet the molecular mechanisms of Smo movement and localization are poorly understood. Using ultrasensitive single-molecule tracking with high spatial/temporal precision (30 nm/10 ms), we discovered that binding events disrupt the primarily diffusive movement of Smo in cilia at an array of sites near the base. The affinity of Smo for these binding sites was modulated by the Hh pathway activation state. Activation, by either a ligand or genetic loss of the negatively acting Hh receptor Patched-1 (Ptch), reduced the affinity and frequency of Smo binding at the base. Our findings quantify activation-dependent changes in Smo dynamics in cilia and highlight a previously unknown step in Hh pathway activation. PMID:26100903

  6. Robust and stretchable indium gallium zinc oxide-based electronic textiles formed by cilia-assisted transfer printing

    NASA Astrophysics Data System (ADS)

    Yoon, Jongwon; Jeong, Yunkyung; Kim, Heeje; Yoo, Seonggwang; Jung, Hoon Sun; Kim, Yonghun; Hwang, Youngkyu; Hyun, Yujun; Hong, Woong-Ki; Lee, Byoung Hun; Choa, Sung-Hoon; Ko, Heung Cho

    2016-06-01

    Electronic textile (e-textile) allows for high-end wearable electronic devices that provide easy access for carrying, handling and using. However, the related technology does not seem to be mature because the woven fabric hampers not only the device fabrication process directly on the complex surface but also the transfer printing of ultrathin planar electronic devices. Here we report an indirect method that enables conformal wrapping of surface with arbitrary yet complex shapes. Artificial cilia are introduced in the periphery of electronic devices as adhesive elements. The cilia also play an important role in confining a small amount of glue and damping mechanical stress to maintain robust electronic performance under mechanical deformation. The example of electronic applications depicts the feasibility of cilia for `stick-&-play' systems, which provide electronic functions by transfer printing on unconventional complex surfaces.

  7. Colony Expansion of Socially Motile Myxococcus xanthus Cells Is Driven by Growth, Motility, and Exopolysaccharide Production

    PubMed Central

    Patra, Pintu; Kissoon, Kimberley; Cornejo, Isabel; Kaplan, Heidi B.; Igoshin, Oleg A.

    2016-01-01

    Myxococcus xanthus, a model organism for studies of multicellular behavior in bacteria, moves exclusively on solid surfaces using two distinct but coordinated motility mechanisms. One of these, social (S) motility is powered by the extension and retraction of type IV pili and requires the presence of exopolysaccharides (EPS) produced by neighboring cells. As a result, S motility requires close cell-to-cell proximity and isolated cells do not translocate. Previous studies measuring S motility by observing the colony expansion of cells deposited on agar have shown that the expansion rate increases with initial cell density, but the biophysical mechanisms involved remain largely unknown. To understand the dynamics of S motility-driven colony expansion, we developed a reaction-diffusion model describing the effects of cell density, EPS deposition and nutrient exposure on the expansion rate. Our results show that at steady state the population expands as a traveling wave with a speed determined by the interplay of cell motility and growth, a well-known characteristic of Fisher’s equation. The model explains the density-dependence of the colony expansion by demonstrating the presence of a lag phase–a transient period of very slow expansion with a duration dependent on the initial cell density. We propose that at a low initial density, more time is required for the cells to accumulate enough EPS to activate S-motility resulting in a longer lag period. Furthermore, our model makes the novel prediction that following the lag phase the population expands at a constant rate independent of the cell density. These predictions were confirmed by S motility experiments capturing long-term expansion dynamics. PMID:27362260

  8. Cytometry of mammalian sperm

    SciTech Connect

    Gledhill, B.L.

    1983-10-11

    Male germ cells respond dramatically to a variety of insults and are important reproductive dosimeters. Semen analyses are very useful in studies on the effects of drugs, chemicals, and environmental hazards on testicular function, male fertility and heritable germinal mutations. The accessibility of male cells makes them well suited for analytical cytology. We might automate the process of determining sperm morphology but should not do so solely for increased speed. Rather, richer tangible benefits will derive from cytometric evaluation through increased sensitivity, reduced subjectivity, standardization between investigators and laboratories, enhanced archival systems, and the benefits of easily exchanged standardized data. Inroads on the standardization of assays for motility and functional integrity are being made. Flow cytometric analysis of total DNA content of individual sperm is an insensitive means to detect exposure to reproductive toxins because of the small size and low frequency of the DNA content errors. Flow cytometry can be applied to determine the proportions of X- and Y-sperm in semen samples.

  9. The mammalian blastocyst.

    PubMed

    Frankenberg, Stephen R; de Barros, Flavia R O; Rossant, Janet; Renfree, Marilyn B

    2016-01-01

    The blastocyst is a mammalian invention that carries the embryo from cleavage to gastrulation. For such a simple structure, it exhibits remarkable diversity in its mode of formation, morphology, longevity, and intimacy with the uterine endometrium. This review explores this diversity in the light of the evolution of viviparity, comparing the three main groups of mammals: monotremes, marsupials, and eutherians. The principal drivers in blastocyst evolution were loss of yolk coupled with evolution of the placenta. An important outcome of blastocyst development is differentiation of two extraembryonic lineages (trophoblast and hypoblast) that contribute to the placenta. While in many species trophoblast segregation is often coupled with blastocyst formation, in marsupials and at least some Afrotherians, these events do not coincide. Thus, many questions regarding the conservation of molecular mechanisms controlling these events are of great interest but currently unresolved. For further resources related to this article, please visit the WIREs website. PMID:26799266

  10. Mammalian phospholipase C.

    PubMed

    Kadamur, Ganesh; Ross, Elliott M

    2013-01-01

    Phospholipase C (PLC) converts phosphatidylinositol 4,5-bisphosphate (PIP(2)) to inositol 1,4,5-trisphosphate (IP(3)) and diacylglycerol (DAG). DAG and IP(3) each control diverse cellular processes and are also substrates for synthesis of other important signaling molecules. PLC is thus central to many important interlocking regulatory networks. Mammals express six families of PLCs, each with both unique and overlapping controls over expression and subcellular distribution. Each PLC also responds acutely to its own spectrum of activators that includes heterotrimeric G protein subunits, protein tyrosine kinases, small G proteins, Ca(2+), and phospholipids. Mammalian PLCs are autoinhibited by a region in the catalytic TIM barrel domain that is the target of much of their acute regulation. In combination, the PLCs act as a signaling nexus that integrates numerous signaling inputs, critically governs PIP(2) levels, and regulates production of important second messengers to determine cell behavior over the millisecond to hour timescale. PMID:23140367

  11. Meckel-Gruber syndrome and the role of primary cilia in kidney, skeleton, and central nervous system development

    PubMed Central

    Barker, Amy R; Thomas, Rhys; Dawe, Helen R

    2014-01-01

    The ciliopathies are a group of related inherited diseases characterized by malformations in organ development. The diseases affect multiple organ systems, with kidney, skeleton, and brain malformations frequently observed. Research over the last decade has revealed that these diseases are due to defects in primary cilia, essential sensory organelles found on most cells in the human body. Here we discuss the genetic and cell biological basis of one of the most severe ciliopathies, Meckel-Gruber syndrome, and explain how primary cilia contribute to the development of the affected organ systems. PMID:24322779

  12. Evidence of 5-HT components in human sperm: implications for protein tyrosine phosphorylation and the physiology of motility

    PubMed Central

    Jiménez-Trejo, Francisco; Tapia-Rodríguez, Miguel; Cerbón, Marco; Kuhn, Donald M; Manjarrez-Gutiérrez, Gabriel; Mendoza-Rodríguez, C Adriana; Picazo, Ofir

    2016-01-01

    Serotonin (5-hydroxytryptamine; C10H12N2O (5-HT)) is produced in the CNS and in some cells of peripheral tissues. In the mammalian male reproductive system, both 5-HT and tryptophan hydroxylase (TPH) have been described in Leydig cells of the testis and in principal cells of the caput epididymis. In capacitated hamster sperm, it has been shown that 5-HT promotes the acrosomal reaction. The aim of this work was to explore the existence of components of the serotoninergic system and their relevance in human sperm physiology. We used both immunocytochemistry and western blot to detect serotoninergic markers such as 5-HT, TPH1, MAOA, 5-HT1B, 5-HT3, and 5HTT; HPLC for TPH enzymatic activity; Computer Assisted Semen Analysis assays to measure sperm motility parameters and pharmacological approaches to show the effect of 5-HT in sperm motility and tyrosine phosphorylation was assessed by western blot. We found the presence of serotoninergic markers (5-HT, TPH1, MAOA, 5-HT1B, 5-HT2A, 5-HT3, 5-HTT, and TPH enzymatic activity) in human sperm. In addition, we observed a significant increase in tyrosine phosphorylation and changes in sperm motility after 5-HT treatment. In conclusion, our data demonstrate the existence of components of a serotoninergic system in human sperm and support the notion for a functional role of 5-HT in mammalian sperm physiology, which can be modulated pharmacologically. PMID:23028123

  13. A novel biosensor to study cAMP dynamics in cilia and flagella

    PubMed Central

    Mukherjee, Shatanik; Jansen, Vera; Jikeli, Jan F; Hamzeh, Hussein; Alvarez, Luis; Dombrowski, Marco; Balbach, Melanie; Strünker, Timo; Seifert, Reinhard; Kaupp, U Benjamin; Wachten, Dagmar

    2016-01-01

    The cellular messenger cAMP regulates multiple cellular functions, including signaling in cilia and flagella. The cAMP dynamics in these subcellular compartments are ill-defined. We introduce a novel FRET-based cAMP biosensor with nanomolar sensitivity that is out of reach for other sensors. To measure cAMP dynamics in the sperm flagellum, we generated transgenic mice and reveal that the hitherto methods determining total cAMP levels do not reflect changes in free cAMP levels. Moreover, cAMP dynamics in the midpiece and principal piece of the flagellum are distinctively different. The sole cAMP source in the flagellum is the soluble adenylate cyclase (SACY). Although bicarbonate-dependent SACY activity requires Ca2+, basal SACY activity is suppressed by Ca2+. Finally, we also applied the sensor to primary cilia. Our new cAMP biosensor features unique characteristics that allow gaining new insights into cAMP signaling and unravel the molecular mechanisms underlying ciliary function in vitro and in vivo. DOI: http://dx.doi.org/10.7554/eLife.14052.001 PMID:27003291

  14. Co-expression of anoctamins in cilia of olfactory sensory neurons.

    PubMed

    Henkel, Bastian; Drose, Daniela R; Ackels, Tobias; Oberland, Sonja; Spehr, Marc; Neuhaus, Eva M

    2015-02-01

    Vertebrates can sense and identify a vast array of chemical cues. The molecular machinery involved in chemodetection and transduction is expressed within the cilia of olfactory sensory neurons. Currently, there is only limited information available on the distribution and density of individual signaling components within the ciliary compartment. Using super-resolution microscopy, we show here that cyclic-nucleotide-gated channels and calcium-activated chloride channels of the anoctamin family are localized to discrete microdomains in the ciliary membrane. In addition to ANO2, a second anoctamin, ANO6, also localizes to ciliary microdomains. This observation, together with the fact that ANO6 and ANO2 co-localize, indicates a role for ANO6 in olfactory signaling. We show that both ANO2 and ANO6 can form heteromultimers and that this heteromerization alters the recombinant channels' physiological properties. Thus, we provide evidence for interaction of ANO2 and ANO6 in olfactory cilia, with possible physiological relevance for olfactory signaling. PMID:25500808

  15. Primary cilia function regulates the length of the embryonic trunk axis and urogenital field in mice.

    PubMed

    Wainwright, Elanor N; Svingen, Terje; Ng, Ee Ting; Wicking, Carol; Koopman, Peter

    2014-11-15

    The issues of whether and how some organs coordinate their size and shape with the blueprint of the embryo axis, while others appear to regulate their morphogenesis autonomously, remain poorly understood. Mutations in Ift144, encoding a component of the trafficking machinery of primary cilia assembly, result in a range of embryo patterning defects, affecting the limbs, skeleton and neural system. Here, we show that embryos of the mouse mutant Ift144(twt) develop gonads that are larger than wild-type. Investigation of the early patterning of the urogenital ridge revealed that the anterior-posterior domain of the gonad/mesonephros was extended at 10.5 dpc, with no change in the length of the metanephros. In XY embryos, this extension resulted in an increase in testis cord number. Moreover, we observed a concomitant extension of the trunk axis in both sexes, with no change in the length of the tail domain or somite number. Our findings support a model in which: (1) primary cilia regulate embryonic trunk elongation; (2) the length of the trunk axis determines the size of the urogenital ridges; and (3) the gonad domain is partitioned into a number of testis cords that depends on the available space, rather than being divided a predetermined number of times to generate a specific number of cords. PMID:25224227

  16. Mechanical signals promote osteogenic fate through a primary cilia-mediated mechanism.

    PubMed

    Chen, Julia C; Hoey, David A; Chua, Mardonn; Bellon, Raymond; Jacobs, Christopher R

    2016-04-01

    It has long been suspected, but never directly shown, that bone formed to accommodate an increase in mechanical loading is related to the creation of osteoblasts from skeletal stem cells. Indeed, biophysical stimuli potently regulate osteogenic lineage commitmentin vitro In this study, we transplanted bone marrow cells expressing green fluorescent protein, to enable lineage tracing, and subjected mice to a biophysical stimulus, to elicit a bone-forming response. We detected cells derived from transplanted progenitors embedded within the bone matrix near active bone-forming surfaces in response to loading, demonstrating for the first time, that mechanical signals enhance the homing and attachment of bone marrow cells to bone surfaces and the commitment to an osteogenic lineage of these cellsin vivo Furthermore, we used an inducible Cre/Lox recombination system to delete kinesin family member 3A (Kif3a), a gene that is essential for primary cilia formation, at will in transplanted cells and their progeny, regardless of which tissue may have incorporated them. Disruption of the mechanosensing organelle, the primary cilium in a progenitor population, significantly decreased the amount of bone formed in response to mechanical stimulation. The collective results of our study directly demonstrate that, in a novel experimental stem cell mechanobiology model, mechanical signals enhance osteogenic lineage commitmentin vivoand that the primary cilium contributes to this process.-Chen, J. C., Hoey, D. A., Chua, M., Bellon, R., Jacobs, C. R. Mechanical signals promote osteogenic fate through a primary cilia-mediated mechanism. PMID:26675708

  17. Phosphorylation-dependent Akt-Inversin interaction at the basal body of primary cilia.

    PubMed

    Suizu, Futoshi; Hirata, Noriyuki; Kimura, Kohki; Edamura, Tatsuma; Tanaka, Tsutomu; Ishigaki, Satoko; Donia, Thoria; Noguchi, Hiroko; Iwanaga, Toshihiko; Noguchi, Masayuki

    2016-06-15

    A primary cilium is a microtubule-based sensory organelle that plays an important role in human development and disease. However, regulation of Akt in cilia and its role in ciliary development has not been demonstrated. Using yeast two-hybrid screening, we demonstrate that Inversin (INVS) interacts with Akt. Mutation in the INVS gene causes nephronophthisis type II (NPHP2), an autosomal recessive chronic tubulointerstitial nephropathy. Co-immunoprecipitation assays show that Akt interacts with INVS via the C-terminus. In vitro kinase assays demonstrate that Akt phosphorylates INVS at amino acids 864-866 that are required not only for Akt interaction, but also for INVS dimerization. Co-localization of INVS and phosphorylated form of Akt at the basal body is augmented by PDGF-AA Akt-null MEF cells as well as siRNA-mediated inhibition of Akt attenuated ciliary growth, which was reversed by Akt reintroduction. Mutant phosphodead- or NPHP2-related truncated INVS, which lack Akt phosphorylation sites, suppress cell growth and exhibit distorted lumen formation and misalignment of spindle axis during cell division. Further studies will be required for elucidating functional interactions of Akt-INVS at the primary cilia for identifying the molecular mechanisms underlying NPHP2. PMID:27220846

  18. RTTN Mutations Link Primary Cilia Function to Organization of the Human Cerebral Cortex

    PubMed Central

    Kheradmand Kia, Sima; Verbeek, Elly; Engelen, Erik; Schot, Rachel; Poot, Raymond A.; de Coo, Irenaeus F.M.; Lequin, Maarten H.; Poulton, Cathryn J.; Pourfarzad, Farzin; Grosveld, Frank G.; Brehm, António; de Wit, Marie Claire Y.; Oegema, Renske; Dobyns, William B.; Verheijen, Frans W.; Mancini, Grazia M.S.

    2012-01-01

    Polymicrogyria is a malformation of the developing cerebral cortex caused by abnormal organization and characterized by many small gyri and fusion of the outer molecular layer. We have identified autosomal-recessive mutations in RTTN, encoding Rotatin, in individuals with bilateral diffuse polymicrogyria from two separate families. Rotatin determines early embryonic axial rotation, as well as anteroposterior and dorsoventral patterning in the mouse. Human Rotatin has recently been identified as a centrosome-associated protein. The Drosophila melanogaster homolog of Rotatin, Ana3, is needed for structural integrity of centrioles and basal bodies and maintenance of sensory neurons. We show that Rotatin colocalizes with the basal bodies at the primary cilium. Cultured fibroblasts from affected individuals have structural abnormalities of the cilia and exhibit downregulation of BMP4, WNT5A, and WNT2B, which are key regulators of cortical patterning and are expressed at the cortical hem, the cortex-organizing center that gives rise to Cajal-Retzius (CR) neurons. Interestingly, we have shown that in mouse embryos, Rotatin colocalizes with CR neurons at the subpial marginal zone. Knockdown experiments in human fibroblasts and neural stem cells confirm a role for RTTN in cilia structure and function. RTTN mutations therefore link aberrant ciliary function to abnormal development and organization of the cortex in human individuals. PMID:22939636

  19. Cilia play a role in breaking left-right symmetry of the sea urchin embryo.

    PubMed

    Takemoto, Ayumi; Miyamoto, Tatsuo; Simono, Fumie; Kurogi, Nao; Shirae-Kurabayashi, Maki; Awazu, Akinori; Suzuki, Ken-Ichi T; Yamamoto, Takashi; Sakamoto, Naoaki

    2016-06-01

    Left-right asymmetry of bilaterian animals is established during early development. In mice, frogs and fishes, the ciliated left-right organizer plays an essential role in establishing bilateral asymmetry, and leftward flow of extracellular fluid generated by ciliary motion results in Nodal activity on the left side. However, H(+) /K(+) -ATPase activity is also involved in the determination of left-right asymmetry in a variety of animals, and it has been thought to be an ancestral mechanism in deuterostomes. In sea urchin, the determination of the left-right asymmetry based on H(+) /K(+) -ATPase activity was already clarified, but it remains to be uncovered whether ciliary motion is involved in the left-right asymmetry of the embryo. Here, we show evidence that ciliary motion is involved in the establishment of left-right asymmetry of sea urchin embryo. Furthermore, we show that the initial cilia generated on small micromeres during the early stage of embryogenesis may be involved in this process. These results suggest that the cilia-mediated mechanism for the determination of left-right asymmetry may be acquired at the base of the deuterostomes. PMID:27028068

  20. A novel biosensor to study cAMP dynamics in cilia and flagella.

    PubMed

    Mukherjee, Shatanik; Jansen, Vera; Jikeli, Jan F; Hamzeh, Hussein; Alvarez, Luis; Dombrowski, Marco; Balbach, Melanie; Strünker, Timo; Seifert, Reinhard; Kaupp, U Benjamin; Wachten, Dagmar

    2016-01-01

    The cellular messenger cAMP regulates multiple cellular functions, including signaling in cilia and flagella. The cAMP dynamics in these subcellular compartments are ill-defined. We introduce a novel FRET-based cAMP biosensor with nanomolar sensitivity that is out of reach for other sensors. To measure cAMP dynamics in the sperm flagellum, we generated transgenic mice and reveal that the hitherto methods determining total cAMP levels do not reflect changes in free cAMP levels. Moreover, cAMP dynamics in the midpiece and principal piece of the flagellum are distinctively different. The sole cAMP source in the flagellum is the soluble adenylate cyclase (SACY). Although bicarbonate-dependent SACY activity requires Ca(2+), basal SACY activity is suppressed by Ca(2+). Finally, we also applied the sensor to primary cilia. Our new cAMP biosensor features unique characteristics that allow gaining new insights into cAMP signaling and unravel the molecular mechanisms underlying ciliary function in vitro and in vivo. PMID:27003291

  1. Primary ciliary dyskinesia: evaluation using cilia beat frequency assessment via spectral analysis of digital microscopy images

    PubMed Central

    Kögler, João E.; Macchione, Mariangela; Shoemark, Amelia; Saldiva, Paulo H. N.; Rodrigues, Joaquim C.

    2011-01-01

    Ciliary beat frequency (CBF) measurements provide valuable information for diagnosing of primary ciliary dyskinesia (PCD). We developed a system for measuring CBF, used it in association with electron microscopy to diagnose PCD, and then analyzed characteristics of PCD patients.1 The CBF measurement system was based on power spectra measured through digital imaging. Twenty-four patients suspected of having PCD (age 1–19 yr) were selected from a group of 75 children and adolescents with pneumopathies of unknown causes. Ten healthy, nonsmoking volunteers (age ≥17 yr) served as a control group. Nasal brush samples were collected, and CBF and electron microscopy were performed. PCD was diagnosed in 12 patients: 5 had radial spoke defects, 3 showed absent central microtubule pairs with transposition, 2 had outer dynein arm defects, 1 had a shortened outer dynein arm, and 1 had a normal ultrastructure. Previous studies have reported that the most common cilia defects are in the dynein arm. As expected, the mean CBF was higher in the control group (P < 0.001) and patients with normal ultrastructure (P < 0.002), than in those diagnosed with cilia ultrastructural defects (i.e., PCD patients). An obstructive ventilatory pattern was observed in 70% of the PCD patients who underwent pulmonary function tests. All PCD patients presented bronchial wall thickening on chest computed tomography scans. The protocol and diagnostic techniques employed allowed us to diagnose PCD in 16% of patients in this study. PMID:21551013

  2. MOTILITY, AGGRESSION, AND THE BODILY I: AN INTERPRETATION OF WINNICOTT.

    PubMed

    Elkins, Jeremy

    2015-10-01

    Among the central ideas associated with the name of Winnicott, scant mention is made of motility. This is largely attributable to Winnicott himself, who never thematized motility and never wrote a paper specifically devoted to the topic. This paper suggests both that the idea of motility is nonetheless of central significance in Winnicott's thought, and that motility is of central importance in the development and constitution of the bodily I. In elaborating both these suggestions, the paper gives particular attention to the connections between motility, continuity, aggression, and creativity in Winnicott's work. PMID:26443951

  3. PKCα-Mediated Signals Regulate the Motile Responses of Cochlear Outer Hair Cells

    PubMed Central

    Park, Channy; Kalinec, Federico

    2015-01-01

    There is strong evidence that changes in the actin/spectrin-based cortical cytoskeleton of outer hair cells (OHCs) regulate their motile responses as well as cochlear amplification, the process that optimizes the sensitivity and frequency selectivity of the mammalian inner ear. Since a RhoA/protein kinase C (PKC)-mediated pathway is known to inhibit the actin-spectrin interaction in other cell models, we decided to investigate whether this signaling cascade could also participate in the regulation of OHC motility. We used high-speed video microscopy and confocal microscopy to explore the effects of pharmacological activation of PKCα, PKCβI, PKCβII, PKCδ, PKCε, and PKCζ with lysophosphatidic acid (LPA) and their inhibition with bisindolylmaleimide I, as well as inhibition of RhoA and Rho-associated protein kinase (ROCK) with C3 and Y-27632, respectively. Motile responses were induced in isolated guinea pig OHCs by stimulation with an 8 V/cm external alternating electrical field as 50 Hz bursts of square wave pulses (100 ms on/off). We found that LPA increased expression of PKCα and PKCζ only, with PKCα, but not PKCζ, phosphorylating the cytoskeletal protein adducin of both Ser-726 and Thr-445. Interestingly, however, inhibition of PKCα reduced adducin phosphorylation only at Ser-726. We also determined that LPA activation of a PKCα-mediated signaling pathway simultaneously enhanced OHC electromotile amplitude and cell shortening, and facilitated RhoA/ROCK/LIMK1-mediated cofilin phosphorylation. Altogether, our results suggest that PKCα-mediated signals, probably via adducin-mediated inhibition of actin-spectrin binding and cofilin-mediated depolymerization of actin filaments, play an essential role in the homeostatic regulation of OHC motility and cochlear amplification. PMID:25954875

  4. Ionic imbalance, in addition to molecular crowding, abates cytoskeletal dynamics and vesicle motility during hypertonic stress

    PubMed Central

    Nunes, Paula; Roth, Isabelle; Meda, Paolo; Féraille, Eric; Brown, Dennis; Hasler, Udo

    2015-01-01

    Cell volume homeostasis is vital for the maintenance of optimal protein density and cellular function. Numerous mammalian cell types are routinely exposed to acute hypertonic challenge and shrink. Molecular crowding modifies biochemical reaction rates and decreases macromolecule diffusion. Cell volume is restored rapidly by ion influx but at the expense of elevated intracellular sodium and chloride levels that persist long after challenge. Although recent studies have highlighted the role of molecular crowding on the effects of hypertonicity, the effects of ionic imbalance on cellular trafficking dynamics in living cells are largely unexplored. By tracking distinct fluorescently labeled endosome/vesicle populations by live-cell imaging, we show that vesicle motility is reduced dramatically in a variety of cell types at the onset of hypertonic challenge. Live-cell imaging of actin and tubulin revealed similar arrested microfilament motility upon challenge. Vesicle motility recovered long after cell volume, a process that required functional regulatory volume increase and was accelerated by a return of extracellular osmolality to isosmotic levels. This delay suggests that, although volume-induced molecular crowding contributes to trafficking defects, it alone cannot explain the observed effects. Using fluorescent indicators and FRET-based probes, we found that intracellular ATP abundance and mitochondrial potential were reduced by hypertonicity and recovered after longer periods of time. Similar to the effects of osmotic challenge, isovolumetric elevation of intracellular chloride concentration by ionophores transiently decreased ATP production by mitochondria and abated microfilament and vesicle motility. These data illustrate how perturbed ionic balance, in addition to molecular crowding, affects membrane trafficking. PMID:26045497

  5. Soft micromachines with programmable motility and morphology

    NASA Astrophysics Data System (ADS)

    Huang, Hen-Wei; Sakar, Mahmut Selman; Petruska, Andrew J.; Pané, Salvador; Nelson, Bradley J.

    2016-07-01

    Nature provides a wide range of inspiration for building mobile micromachines that can navigate through confined heterogenous environments and perform minimally invasive environmental and biomedical operations. For example, microstructures fabricated in the form of bacterial or eukaryotic flagella can act as artificial microswimmers. Due to limitations in their design and material properties, these simple micromachines lack multifunctionality, effective addressability and manoeuvrability in complex environments. Here we develop an origami-inspired rapid prototyping process for building self-folding, magnetically powered micromachines with complex body plans, reconfigurable shape and controllable motility. Selective reprogramming of the mechanical design and magnetic anisotropy of body parts dynamically modulates the swimming characteristics of the micromachines. We find that tail and body morphologies together determine swimming efficiency and, unlike for rigid swimmers, the choice of magnetic field can subtly change the motility of soft microswimmers.

  6. Symbiosis and the origin of eukaryotic motility

    NASA Technical Reports Server (NTRS)

    Margulis, L.; Hinkle, G.

    1991-01-01

    Ongoing work to test the hypothesis of the origin of eukaryotic cell organelles by microbial symbioses is discussed. Because of the widespread acceptance of the serial endosymbiotic theory (SET) of the origin of plastids and mitochondria, the idea of the symbiotic origin of the centrioles and axonemes for spirochete bacteria motility symbiosis was tested. Intracellular microtubular systems are purported to derive from symbiotic associations between ancestral eukaryotic cells and motile bacteria. Four lines of approach to this problem are being pursued: (1) cloning the gene of a tubulin-like protein discovered in Spirocheata bajacaliforniesis; (2) seeking axoneme proteins in spirochets by antibody cross-reaction; (3) attempting to cultivate larger, free-living spirochetes; and (4) studying in detail spirochetes (e.g., Cristispira) symbiotic with marine animals. Other aspects of the investigation are presented.

  7. Hydrodynamic Contributions to Amoeboid Cell Motility

    NASA Astrophysics Data System (ADS)

    Lewis, Owen; Guy, Robert

    2011-11-01

    Understanding the methods by which cells move is a fundamental problem in modern biology. Recent evidence has shown that the fluid dynamics of cytoplasm can play a vital role in cellular motility. The slime mold Physarum polycephalum provides an excellent model organism for the study of amoeboid motion. In this research, we use both analytic and computational models to investigate intracellular fluid flow in a simple model of Physarum. In both models, of we are specifically interested in stresses generated by cytoplasmic flow which act in the direction of cellular motility. In our numerical model, the Immersed Boundary Method is used to account for such stresses. We investigate the relationship between contraction waves, low waves and locomotive forces, and attempt characterize conditions necessary to generate directed motion.

  8. Hydrodynamic Contributions to Amoeboid Cell Motility

    NASA Astrophysics Data System (ADS)

    Lewis, Owen; Guy, Robert

    2012-11-01

    Understanding the methods by which cells move is a fundamental problem in modern biology. Recent evidence has shown that the fluid dynamics of cytoplasm can play a vital role in cellular motility. The slime mold Physarum polycephalum provides an excellent model organism for the study of amoeboid motion. In this research, we use a simply analytic model in conjuction with computational experiments to investigate intracellular fluid flow in a simple model of Physarum. Of particlar interest are stresses generated by cytoplasmic flow which may be used to aid in cellular motility. In our numerical model, the Immersed Boundary Method is used to account for such stresses. We investigate the relationship between contraction waves, flow waves, adhesion, and locomotive forces in an attempt to characterize conditions necessary to generate directed motion.

  9. Soft micromachines with programmable motility and morphology.

    PubMed

    Huang, Hen-Wei; Sakar, Mahmut Selman; Petruska, Andrew J; Pané, Salvador; Nelson, Bradley J

    2016-01-01

    Nature provides a wide range of inspiration for building mobile micromachines that can navigate through confined heterogenous environments and perform minimally invasive environmental and biomedical operations. For example, microstructures fabricated in the form of bacterial or eukaryotic flagella can act as artificial microswimmers. Due to limitations in their design and material properties, these simple micromachines lack multifunctionality, effective addressability and manoeuvrability in complex environments. Here we develop an origami-inspired rapid prototyping process for building self-folding, magnetically powered micromachines with complex body plans, reconfigurable shape and controllable motility. Selective reprogramming of the mechanical design and magnetic anisotropy of body parts dynamically modulates the swimming characteristics of the micromachines. We find that tail and body morphologies together determine swimming efficiency and, unlike for rigid swimmers, the choice of magnetic field can subtly change the motility of soft microswimmers. PMID:27447088

  10. Congo red uptake by motile Aeromonas species.

    PubMed

    Statner, B; George, W L

    1987-05-01

    Virulence of several species of enteropathogenic bacteria has been correlated with the ability of isolates to take up the dye Congo red. To determine whether Congo red uptake might be a useful marker for virulence of motile Aeromonas species, we examined 50 strains of diverse clinical origin on a medium containing 50 micrograms of Congo red per ml. All of the strains took up the dye to various degrees. For most strains, uptake was greatest at 37 degrees C and least at 22 degrees C. Production of acetyl methyl carbinol (Voges-Proskauer test) or lysine decarboxylase has been reported by some investigators to be a virulence marker for Aeromonas species. Congo red uptake did not correlate with either acetyl methyl carbinol or lysine decarboxylase production in our study. These data suggest that Congo red uptake may not be a useful marker for virulence of motile Aeromonas species. PMID:3584422

  11. Esophageal motility abnormalities in gastroesophageal reflux disease

    PubMed Central

    Martinucci, Irene; de Bortoli, Nicola; Giacchino, Maria; Bodini, Giorgia; Marabotto, Elisa; Marchi, Santino; Savarino, Vincenzo; Savarino, Edoardo

    2014-01-01

    Esophageal motility abnormalities are among the main factors implicated in the pathogenesis of gastroesophageal reflux disease. The recent introduction in clinical and research practice of novel esophageal testing has markedly improved our understanding of the mechanisms contributing to the development of gastroesophageal reflux disease, allowing a better management of patients with this disorder. In this context, the present article intends to provide an overview of the current literature about esophageal motility dysfunctions in patients with gastroesophageal reflux disease. Esophageal manometry, by recording intraluminal pressure, represents the gold standard to diagnose esophageal motility abnormalities. In particular, using novel techniques, such as high resolution manometry with or without concurrent intraluminal impedance monitoring, transient lower esophageal sphincter (LES) relaxations, hypotensive LES, ineffective esophageal peristalsis and bolus transit abnormalities have been better defined and strongly implicated in gastroesophageal reflux disease development. Overall, recent findings suggest that esophageal motility abnormalities are increasingly prevalent with increasing severity of reflux disease, from non-erosive reflux disease to erosive reflux disease and Barrett’s esophagus. Characterizing esophageal dysmotility among different subgroups of patients with reflux disease may represent a fundamental approach to properly diagnose these patients and, thus, to set up the best therapeutic management. Currently, surgery represents the only reliable way to restore the esophagogastric junction integrity and to reduce transient LES relaxations that are considered to be the predominant mechanism by which gastric contents can enter the esophagus. On that ground, more in depth future studies assessing the pathogenetic role of dysmotility in patients with reflux disease are warranted. PMID:24868489

  12. [Gastrointestinal motility and possibilities of influencing it].

    PubMed

    Duris, I; Payer, J; Huorka, M; Randus, V; Ondrejka, P

    1994-06-01

    The authors discuss factors which influence the motility of the smooth muscles in the pancreatobiliary region. They investigated some clinical and laboratory parameters after administration of the selective antagonist of calcium influx-Pineverium bromide-Dicetel. The drug influenced significantly in a positive way nausea, flatulence, pain and chronically elevated amylases. The authors mention a cycle of possible neurohumoral changes with which specific calcium channel antagonists could interfere. PMID:8073641

  13. Swimming Motility Reduces Deposition to Silica Surfaces

    SciTech Connect

    Lu, Nanxi; Massoudieh, Arash; Liang, Xiaomeng; Hu, Dehong; Kamai, Tamir; Ginn, Timothy R.; Zilles, Julie L.; Nguyen, Thanh H.

    2015-01-01

    The role of swimming motility on bacterial transport and fate in porous media was evaluated. We present microscopic evidence showing that strong swimming motility reduces attachment of Azotobacter vinelandii cells to silica surfaces. Applying global and cluster statistical analyses to microscopic videos taken under non-flow conditions, wild type, flagellated A. vinelandii strain DJ showed strong swimming ability with an average speed of 13.1 μm/s, DJ77 showed impaired swimming averaged at 8.7 μm/s, and both the non-flagellated JZ52 and chemically treated DJ cells were non-motile. Quantitative analyses of trajectories observed at different distances above the collector of a radial stagnation point flow cell (RSPF) revealed that both swimming and non-swimming cells moved with the flow when at a distance of at least 20 μm from the collector surface. Near the surface, DJ cells showed both horizontal and vertical movement diverging them from reaching surfaces, while chemically treated DJ cells moved with the flow to reach surfaces, suggesting that strong swimming reduced attachment. In agreement with the RSPF results, the deposition rates obtained for two-dimensional multiple-collector micromodels were also lowest for DJ, while DJ77 and JZ52 showed similar values. Strong swimming specifically reduced deposition on the upstream surfaces of the micromodel collectors.

  14. Bacteria motility at oil-water interfaces

    NASA Astrophysics Data System (ADS)

    Juarez, Gabriel; Smirga, Steven; Fernandez, Vicente; Stocker, Roman

    2012-11-01

    The swimming dynamics of bacteria are strongly influenced by interfaces: Motile bacteria often accumulate at rigid boundaries, such as liquid-solid interfaces, and at soft boundaries, such as liquid-air or liquid-liquid interfaces. Attachment of bacteria to these interfaces is crucial for the formation of biofilms (liquid-solid), pellicles (liquid-air), and oil-degrading communities (liquid-liquid). We investigated the motility of the oil-degrading bacteria Marinobacter aquaeolei in the presence of oil droplets. We created individual oil droplets using dedicated microfluidic devices and captured the swimming behavior of individual bacteria near the interface and their attachment dynamics to the droplets with high-speed and epifluorescent microscopy. We find that Marinobacter aquaeolei has a high affinity towards interfaces and their swimming dynamics at soft interfaces differ from both those in the bulk and at rigid boundaries. Characterizing the interaction and attachment of motile bacteria to liquid-liquid interfaces will promote a fundamental understanding to oil-microbe interactions in aquatic environments and potentially lead to improved oil bioremediation strategies.

  15. Motility of active fluid drops on surfaces

    NASA Astrophysics Data System (ADS)

    Khoromskaia, Diana; Alexander, Gareth P.

    2015-12-01

    Drops of active liquid crystal have recently shown the ability to self-propel, which was associated with topological defects in the orientation of active filaments [Sanchez et al., Nature 491, 431 (2013), 10.1038/nature11591]. Here, we study the onset and different aspects of motility of a three-dimensional drop of active fluid on a planar surface. We analyze theoretically how motility is affected by orientation profiles with defects of various types and locations, by the shape of the drop, and by surface friction at the substrate. In the scope of a thin drop approximation, we derive exact expressions for the flow in the drop that is generated by a given orientation profile. The flow has a natural decomposition into terms that depend entirely on the geometrical properties of the orientation profile, i.e., its bend and splay, and a term coupling the orientation to the shape of the drop. We find that asymmetric splay or bend generates a directed bulk flow and enables the drop to move, with maximal speeds achieved when the splay or bend is induced by a topological defect in the interior of the drop. In motile drops the direction and speed of self-propulsion is controlled by friction at the substrate.

  16. Effect of total laryngectomy on esophageal motility

    SciTech Connect

    Hanks, J.B.; Fisher, S.R.; Meyers, W.C.; Christian, K.C.; Postlethwait, R.W.; Jones, R.S.

    1981-01-01

    Total laryngectomy for cancer can result in dysphagia and altered esophageal motility. Manometric changes in the upper esophageal sphincter (UES), and in proximal and distal esophageal function have been reported. However, most studies have failed to take into account radiation therapy and appropriate controls. We selected ten male patients (54.3 +/- 1.9 yr) for longitudinal manometric evaluation prior to laryngectomy then at two weeks and again six months later. No patient received preoperative radiation therapy, had a previous history of esophageal surgery, or developed a postoperative wound infection or fistula. Seven of ten patients had positive nodes and received 6,000-6,600 rads postoperative radiation therapy. Preoperatively 4 of 10 patients complained of dysphagia which did not significantly change following surgery and radiation. Two of three patients who did not complain of dysphagia preoperatively and received radiation postoperatively developed dysphagia. No patient without dysphagia preoperatively who received no radiation therapy developed symptoms. Our studies show that laryngectomy causes alterations in the UES resting and peak pressures but not in the proximal or distal esophagus, or the lower esophageal sphincter. These data also imply radiation therapy may be associated with progressive alterations in motility and symptomatology. Further study regarding the effects of radiation on esophageal motility and function are urged.

  17. Hyaluronan stimulates pancreatic cancer cell motility

    PubMed Central

    Cheng, Xiao-Bo; Kohi, Shiro; Koga, Atsuhiro; Hirata, Keiji; Sato, Norihiro

    2016-01-01

    Hyaluronan (HA) accumulates in pancreatic ductal adenocarcinoma (PDAC), but functional significance of HA in the aggressive phenotype remains unknown. We used different models to investigate the effect of HA on PDAC cell motility by wound healing and transwell migration assay. Changes in cell motility were examined in 8 PDAC cell lines in response to inhibition of HA production by treatment with 4-methylumbelliferone (4-MU) and to promotion by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by co-culture with tumor-derived stromal fibroblasts. We also investigated changes in cell motility by adding exogenous HA. Additionally, mRNA expressions of hyaluronan synthases and hyaluronidases were examined using real time RT-PCR. Inhibition of HA by 4-MU significantly decreased the migration, whereas promotion of HA by TPA or co-culture with tumor-derived fibroblasts significantly increased the migration of PDAC cells. The changes in HA production by these treatments tended to be associated with changes in HAS3 mRNA expression. Furthermore, addition of exogenous HA, especially low-molecular-weight HA, significantly increased the migration of PDAC cells. These findings suggest that HA stimulates PDAC cell migration and thus represents an ideal therapeutic target to prevent invasion and metastasis. PMID:26684359

  18. Mammalian Wax Biosynthesis

    PubMed Central

    Cheng, Jeffrey B.; Russell, David W.

    2009-01-01

    Wax monoesters are synthesized by the esterification of fatty alcohols and fatty acids. A mammalian enzyme that catalyzes this reaction has not been isolated. We used expression cloning to identify cDNAs encoding a wax synthase in the mouse preputial gland. The wax synthase gene is located on the X chromosome and encodes a member of the acyltransferase family of enzymes that synthesize neutral lipids. Expression of wax synthase in cultured cells led to the formation of wax monoesters from straight chain saturated, unsaturated, and polyunsaturated fatty alcohols and acids. Polyisoprenols also were incorporated into wax monoesters by the enzyme. The wax synthase had little or no ability to synthesize cholesteryl esters, diacylglycerols, or triacylglycerols, whereas other acyltransferases, including the acyl-CoA:monoacylglycerol acyltransferase 1 and 2 enzymes and the acyl-CoA:diacylglycerol acyltransferase 1 and 2 enzymes, exhibited modest wax monoester synthesis activities. Confocal light microscopy indicated that the wax synthase was localized in membranes of the endoplasmic reticulum. Wax synthase mRNA was abundant in tissues rich in sebaceous glands such as the preputial gland and eyelid and was present at lower levels in other tissues. Coexpression of cDNAs specifying fatty acyl-CoA reductase 1 and wax synthase led to the synthesis of wax monoesters. The data suggest that wax monoester synthesis in mammals involves a two step biosynthetic pathway catalyzed by fatty acyl-CoA reductase and wax synthase enzymes. PMID:15220349

  19. Structure of mammalian metallothionein

    SciTech Connect

    Kaegi, J.H.R.; Vasak, M.; Lerch, K.; Gilg, D.E.O.; Hunziker, P.; Bernhard, W.R.; Good, M.

    1984-03-01

    All mammalian metallothioneins characterized contain a single polypeptide chain of 61 amino acid residues, among them 20 cysteines providing the ligands for seven metal-binding sites. Native metallothioneins are usually heterogeneous in metal composition, with Zn, Cd, and Cu occurring in varying proportions. However, forms containing only a single metal species, i.e., Zn, Cd, Ni, Co, Hg, Pb, Bi, have now been prepared by in vitro reconstitution from the metal-free apoprotein. By spectroscopic analysis of such derivatives it was established that all cysteine residues participate in metal binding, that each metal ion is bound to four thiolate ligands, and that the symmetry of each complex is close to that of a tetrahedron. To satisfy the requirements of the overall Me/sub 7/(Cys/sup -/)/sub 20/ stoichiometry, the complexes must be combined to form metal-thiolate cluster structures. The actual spatial organization of the clusters and the polypeptide chain remains to be established. An attractive possibility is the arrangement of the tetrahedral metal-thiolates in adamantane-like structures surrounded by properly folded segments of the chain providing the ligands. /sup 1/H-NMR data and infrared absorption measurements are consistent with a tightly folded structure rich in ..beta..-type conformation. 79 references, 11 figures, 4 tables.

  20. Mammalian Sirtuins and Energy Metabolism

    PubMed Central

    Li, Xiaoling; Kazgan, Nevzat

    2011-01-01

    Sirtuins are highly conserved NAD+-dependent protein deacetylases and/or ADP-ribosyltransferases that can extend the lifespan of several lower model organisms including yeast, worms and flies. The seven mammalian sirtuins, SIRT1 to SIRT7, have emerged as key metabolic sensors that directly link environmental signals to mammalian metabolic homeostasis and stress response. Recent studies have shed light on the critical roles of sirtuins in mammalian energy metabolism in response to nutrient signals. This review focuses on the involvement of two nuclear sirtuins, SIRT1 and SIRT6, and three mitochondrial sirtuins, SIRT3, SIRT4, and SIRT5, in regulation of diverse metabolic processes. PMID:21614150

  1. Physics of protein motility and motor proteins

    NASA Astrophysics Data System (ADS)

    Kolomeisky, Anatoly B.

    2013-09-01

    Motor proteins are enzymatic molecules that transform chemical energy into mechanical motion and work. They are critically important for supporting various cellular activities and functions. In the last 15 years significant progress in understanding the functioning of motor proteins has been achieved due to revolutionary breakthroughs in single-molecule experimental techniques and strong advances in theoretical modelling. However, microscopic mechanisms of protein motility are still not well explained, and the collective efforts of many scientists are needed in order to solve these complex problems. In this special section the reader will find the latest advances on the difficult road to mapping motor proteins dynamics in various systems. Recent experimental developments have allowed researchers to monitor and to influence the activity of single motor proteins with a high spatial and temporal resolution. It has stimulated significant theoretical efforts to understand the non-equilibrium nature of protein motility phenomena. The latest results from all these advances are presented and discussed in this special section. We would like to thank the scientists from all over the world who have reported their latest research results for this special section. We are also grateful to the staff and editors of Journal of Physics: Condensed Matter for their invaluable help in handling all the administrative and refereeing activities. The field of motor proteins and protein motility is fast moving, and we hope that this collection of articles will be a useful source of information in this highly interdisciplinary area. Physics of protein motility and motor proteins contents Physics of protein motility and motor proteinsAnatoly B Kolomeisky Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116 Yuan Zhang, Mirkó Palla, Andrew Sun and Jung-Chi Liao The load dependence of the physical properties of a molecular motor

  2. Generating conditional mutants to analyze ciliary functions: the use of Cre-lox technology to disrupt cilia in specific organs.

    PubMed

    O'Connor, Amber K; Kesterson, Robert A; Yoder, Bradley K

    2009-01-01

    The list of human disordered associated with cilia dysfunction, the ciliopathies, continues to highlight the importance of understanding the many roles of the long overlooked primary cilium. Much of the insights into the clinical importance of the cilium have come from analyses in model organisms, especially the mouse. However, the early embryonic lethality and severe developmental defects associated with cilia disruption has hindered progress in exploring cilia functions in late development or in adult tissues. This hurdle is being surmounted through the use of conditional alleles of genes encoding ciliary proteins and Cre deletor lines with inducible Cre activity or with lines expressing Cre in a cell-type-specific manner. Results from these approaches are providing important insights into the diverse array of cellular and tissue activities regulated by the cilium. Here we provide a recent account of the Cre/lox strategy. The generation and use of well-designed conditional alleles, as well as careful manipulation of embryonic stem cells are discussed. We also provide specific examples to illustrate the use of Cre/lox approaches to evaluate ciliary function in several tissues. With the recent characterization of multiple cilia proteomes along with efforts of several consortia to generate conditional alleles of all genes in the mouse, further use of conditional mutation approaches promise to yield many advances and surprises as we explore the functions of this increasingly complex organelle. PMID:20409823

  3. A high-resolution morphological and ultrastructural map of anterior sensory cilia and glia in Caenorhabditis elegans

    PubMed Central

    Doroquez, David B; Berciu, Cristina; Anderson, James R; Sengupta, Piali; Nicastro, Daniela

    2014-01-01

    Many primary sensory cilia exhibit unique architectures that are critical for transduction of specific sensory stimuli. Although basic ciliogenic mechanisms are well described, how complex ciliary structures are generated remains unclear. Seminal work performed several decades ago provided an initial but incomplete description of diverse sensory cilia morphologies in C. elegans. To begin to explore the mechanisms that generate these remarkably complex structures, we have taken advantage of advances in electron microscopy and tomography, and reconstructed three-dimensional structures of fifty of sixty sensory cilia in the C. elegans adult hermaphrodite at high resolution. We characterize novel axonemal microtubule organization patterns, clarify structural features at the ciliary base, describe new aspects of cilia–glia interactions, and identify structures suggesting novel mechanisms of ciliary protein trafficking. This complete ultrastructural description of diverse cilia in C. elegans provides the foundation for investigations into underlying ciliogenic pathways, as well as contributions of defined ciliary structures to specific neuronal functions. DOI: http://dx.doi.org/10.7554/eLife.01948.001 PMID:24668170

  4. Gli2 and Gli3 Localize to Cilia and Require the Intraflagellar Transport Protein Polaris for Processing and Function

    SciTech Connect

    Michaud III, Edward J; Haycraft, Courtney J; Aydin Son, Yesim; Zhang, Qihong; Yoder, Bradley

    2005-01-01

    Intraflagellar transport (IFT) proteins are essential for cilia assembly and have recently been associated with a number of developmental processes, such as left-right axis specification and limb and neural tube patterning. Genetic studies indicate that IFT proteins are required for Sonic hedgehog (Shh)signaling downstream of the Smoothened and Patched membrane proteins but upstream of the Glioma (Gli) transcription factors. However, the role that IFT proteins play in transduction of Shh signaling and the importance of cilia in this process remain unknown. Here we provide insights into the mechanism by which defects in an IFT protein, Tg737/Polaris, affect Shh signaling in the murine limb bud. Our data show that loss of Tg737 results in altered Gli3 processing that abrogates Gli3-mediated repression of Gli1 transcriptional activity. In contrast to the conclusions drawn from genetic analysis, the activity of Gli1 and truncated forms of Gli3 (Gli3R) are unaffected in Tg737 mutants at the molecular level, indicating that Tg737/Polaris is differentially involved in specific activities of the Gli proteins. Most important, a negative regulator of Shh signaling, Suppressor of fused, and the three full-length Gli transcription factors localize to the distal tip of cilia in addition to the nucleus. Thus, our data support a model where cilia have a direct role in Gli processing and Shh signal transduction.

  5. Somatic motility and hair bundle mechanics, are both necessary for cochlear amplification?

    PubMed Central

    Peng, Anthony W.; Ricci, Anthony J.

    2010-01-01

    Hearing organs have evolved to detect sounds across several orders of magnitude of both intensity and frequency. Detection limits are at the atomic level despite the energy associated with sound being limited thermodynamically. Several mechanisms have evolved to account for the remarkable frequency selectivity, dynamic range, and sensitivity of these various hearing organs, together termed the active process or cochlear amplifier. Similarities between hearing organs of disparate species provides insight into the factors driving the development of the cochlear amplifier. These properties include: a tonotopic map, the emergence of a two hair cell system, the separation of efferent and afferent innervations, the role of the tectorial membrane, and the shift from intrinsic tuning and amplification to a more end organ driven process. Two major contributors to the active process are hair bundle mechanics and outer hair cell electromotility, the former present in all hair cell organs tested, the latter only present in mammalian cochlear outer hair cells. Both of these processes have advantages and disadvantages, and how these processes interact to generate the active process in the mammalian system is highly disputed. A hypothesis is put forth suggesting that hair bundle mechanics provides amplification and filtering in most hair cells, while in mammalian cochlea, outer hair cell motility provides the amplification on a cycle by cycle basis driven by the hair bundle that provides frequency selectivity (in concert with the tectorial membrane) and compressive nonlinearity. Separating components of the active process may provide additional sites for regulation of this process. PMID:20430075

  6. The Wireless Motility Capsule: a One-Stop Shop for the Evaluation of GI Motility Disorders.

    PubMed

    Saad, Richard J

    2016-03-01

    The wireless motility and pH capsule (WMC) provides an office-based test to simultaneously assess both regional and whole gut transit. Ingestion of this non-digestible capsule capable of measuring temperature, pH, and the pressure of its immediate surroundings allows for the measurement of gastric, small bowel, and colonic transit times in an ambulatory setting. Approved by the US Food and Drug Administration for the evaluation of suspected conditions of delayed gastric emptying and the evaluation of colonic transit in chronic idiopathic constipation, WMC should be considered in suspected gastrointestinal motility disorders as it provides a single study capable of simultaneously assessing for regional, multiregional, or generalized motility disorders. Specific indications for testing with the WMC should include the evaluation of suspect cases of gastroparesis, small bowel dysmotility, and slow transit constipation, as well as symptom syndromes suggestive of a multiregional or generalized gastrointestinal transit delay. PMID:26908282

  7. Mammalian DNA Repair. Final Report

    SciTech Connect

    2003-01-24

    The Gordon Research Conference (GRC) on Mammalian DNA Repair was held at Harbortown Resort, Ventura Beach, CA. Emphasis was placed on current unpublished research and discussion of the future target areas in this field.

  8. First identification of proteins involved in motility of Mycoplasma gallisepticum.

    PubMed

    Indikova, Ivana; Vronka, Martin; Szostak, Michael P

    2014-01-01

    Mycoplasma gallisepticum, the most pathogenic mycoplasma in poultry, is able to glide over solid surfaces. Although this gliding motility was first observed in 1968, no specific protein has yet been shown to be involved in gliding. We examined M. gallisepticum strains and clonal variants for motility and found that the cytadherence proteins GapA and CrmA were required for gliding. Loss of GapA or CrmA resulted in the loss of motility and hemadsorption and led to drastic changes in the characteristic flask-shape of the cells. To identify further genes involved in motility, a transposon mutant library of M. gallisepticum was generated and screened for motility-deficient mutants, using a screening assay based on colony morphology. Motility-deficient mutants had transposon insertions in gapA and the neighbouring downstream gene crmA. In addition, insertions were seen in gene mgc2, immediately upstream of gapA, in two motility-deficient mutants. In contrast to the GapA/CrmA mutants, the mgc2 motility mutants still possessed the ability to hemadsorb. Complementation of these mutants with a mgc2-hexahistidine fusion gene restored the motile phenotype. This is the first report assigning specific M. gallisepticum proteins to involvement in gliding motility. PMID:25323771

  9. Passive versus active local microrheology in mammalian cells and amoebae

    NASA Astrophysics Data System (ADS)

    Riviere, C.; Gazeau, F.; Marion, S.; Bacri, J.-C.; Wilhelm, C.

    2004-12-01

    We compare in this paper the rotational magnetic microrheology detailed by Marion et al [18] and Wilhelm et al [19] to the passive tracking microrheology. The rotational microrheology has been designed to explore, using magnetic rotating probes, the local intracellular microenvironment of living cells in terms of viscoelasticity. Passive microrheology techniques is based on the analysis of spontaneous diffusive motions of Brownian probes. The dependence of mean square displacement (MSD) with the time then directly reflects the type of movement (sub-, hyper- or diffusive motions). Using the same intracellular probes, we performed two types of measurements (active and passive). Based on the fluctuation-dissipation theorem, one should obtain the same information from the both techniques in a thermally equilibrium system. Interestingly, our measurements differ, and the discordances directly inform on active biological processes, which add to thermally activated fluctuations in our out-of equilibrium systems. In both cell models used, mammalian Hela cells and amoebae Entamoeba Histolytica, a hyper-diffusive regime at a short time is observed, which highlights the presence of an active non-thermal driving force, acting on the probe. However, the nature of this active force in mammalian cells and amoebae is different, according to their different phenotypes. In mammalian cells active processes are governed by the transport, via molecular motors, on the microtubule network. In amoebae, which are highly motile cells free of microtubule network, the active processes are dominated by strong fluxes of cytoplasm driven by extension of pseudopodia, in random directions, leading to an amplitude of motion one order of magnitude higher than for mammalian cells. Figs 7, Refs 32.

  10. Mammalian Interphase Cdks

    PubMed Central

    2012-01-01

    Cyclin-dependent kinases (Cdks) drive cell cycle progression in all eukaryotes. Yeasts have a single major Cdk that mediates distinct cell cycle transitions via association with different cyclins. The closest homolog in mammals, Cdk1, drives mitosis. Mammals have additional Cdks—Cdk2, Cdk4, and Cdk6—that represent the major Cdks activated during interphase (iCdks). A large body of evidence has accrued that suggests that activation of iCdks dictates progression though interphase. In apparent contradiction, deficiency in each individual iCdk, respectively, in knockout mice proved to be compatible with live birth and in some instances fertility. Moreover, murine embryos could be derived with Cdk1 as the only functional Cdk. Thus, none of the iCdks is strictly essential for mammalian cell cycle progression, raising the possibility that Cdk1 is the dominant regulator in interphase. However, an absence of iCdks has been accompanied by major shifts in cyclin association to Cdk1, suggesting gain in function. After considerable tweaking, a chemical genetic approach has recently been able to examine the impact of acute inhibition of Cdk2 activity without marked distortion of cyclin/Cdk complex formation. The results suggest that, when expressed at its normal levels, Cdk2 performs essential roles in driving human cells into S phase and maintaining genomic stability. These new findings appear to have restored order to the cell cycle field, bringing it full circle to the view that iCdks indeed play important roles. They also underscore the caveat in knockdown and knockout approaches that protein underexpression can significantly perturb a protein interaction network. We discuss the implications of the new synthesis for future cell cycle studies and anti–Cdk-based therapy of cancer and other diseases. PMID:23634250

  11. Isotope Labeling in Mammalian Cells

    PubMed Central

    Dutta, Arpana; Saxena, Krishna; Klein-Seetharaman, Judith

    2011-01-01

    Isotope labeling of proteins represents an important and often required tool for the application of nuclear magnetic resonance (NMR) spectroscopy to investigate the structure and dynamics of proteins. Mammalian expression systems have conventionally been considered to be too weak and inefficient for protein expression. However, recent advances have significantly improved the expression levels of these systems. Here, we provide an overview of some of the recent developments in expression strategies for mammalian expression systems in view of NMR investigations. PMID:22167668

  12. Mutations in chemosensory cilia cause resistance to paraquat in nematode Caenorhabditis elegans.

    PubMed

    Fujii, Michihiko; Matsumoto, Yuki; Tanaka, Nanae; Miki, Kensuke; Suzuki, Toshikazu; Ishii, Naoaki; Ayusawa, Dai

    2004-05-01

    The relationship between oxidative stress and longevity is a matter of concern in various organisms. We isolated mutants resistant to paraquat from nematode Caenorhabditis elegans. One mutant named mev-4 was long-lived and showed cross-resistance to heat and Dyf phenotype (defective in dye filling). Genetic and sequence analysis revealed that mev-4 had a nonsense mutation on the che-11 gene, homologues of which are involved in formation of cilia and flagella in other organisms. The paraquat resistance was commonly observed in various Dyf mutants and did not depend on the daf-16 gene, whereas the extension of life span did depend on it. Expression of antioxidant enzyme genes seemed normal. These results suggest that chemosensory neurons are a target of oxidative stress and influence longevity dependent on the daf-16 signaling in C. elegans. PMID:14982934

  13. Swimming Motility Reduces Deposition to Silica Surfaces.

    PubMed

    Lu, Nanxi; Massoudieh, Arash; Liang, Xiaomeng; Hu, Dehong; Kamai, Tamir; Ginn, Timothy R; Zilles, Julie L; Nguyen, Thanh H

    2015-09-01

    The transport and fate of bacteria in porous media is influenced by physicochemical and biological properties. This study investigated the effect of swimming motility on the attachment of cells to silica surfaces through comprehensive analysis of cell deposition in model porous media. Distinct motilities were quantified for different strains using global and cluster-based statistical analyses of microscopic images taken under no-flow condition. The wild-type, flagellated strain DJ showed strong swimming as a result of the actively swimming subpopulation whose average speed was 25.6 μm/s; the impaired swimming of strain DJ77 was attributed to the lower average speed of 17.4 μm/s in its actively swimming subpopulation; and both the nonflagellated JZ52 and chemically treated DJ cells were nonmotile. The approach and deposition of these bacterial cells were analyzed in porous media setups, including single-collector radial stagnation point flow cells (RSPF) and two-dimensional multiple-collector micromodels under well-defined hydrodynamic conditions. In RSPF experiments, both swimming and nonmotile cells moved with the flow when at a distance ≥20 μm above the collector surface. Closer to the surface, DJ cells showed both horizontal and vertical movement, limiting their contact with the surface, while chemically treated DJ cells moved with the flow to reach the surface. These results explain how wild-type swimming reduces attachment. In agreement, the deposition in micromodels was also lowest for DJ compared with those for DJ77 and JZ52. Wild-type swimming specifically reduced deposition on the upstream surfaces of the micromodel collectors. Conducted under environmentally relevant hydrodynamic conditions, the results suggest that swimming motility is an important characteristic for bacterial deposition and transport in the environment. PMID:26436254

  14. Equilibrium shape of liquid lenses and correlations of beating cilia on the surface of frog embryos

    NASA Astrophysics Data System (ADS)

    Huisman, Fawn Mitsu

    This thesis reports the results of two unrelated projects: liquid lenses and cilia motion. A liquid lens is the equilibrium configuration of a non-wetting 3-fluid system, and the profile of the 3 interfaces can, in principle, be determined by solving the Young-Laplace differential equation for each interface if the surface tensions of all three interfaces are accurately known. Fluid interfacial tensions are not tabulated and are spectacularly sensitive to impurities, so in practice these quantities must be measured. We have developed a method of measuring these parameters by comparing the observed shape of liquid lenses to calculated equilibrium shapes. We have experimentally studied lenses of alkanes floating on water. These results were compared with various liquid lens systems. The profile of these lens systems was measured using ray tracing and Morie imaging, and found to be in excellent agreement with the calculated lens shapes for pure fluids. Upon the introduction of a surfactant, dodecyltrimethylammonium bromide(DTAB), we find that there is a minimum in the dihedral angle of the lens as a function of surfactant concentration, corresponding to the transition from partial-wetting to pseudo-partial wetting. The development and orientation of ciliated cells on Xenopus laevis larval skin was examined using high speed video microscopy. The intercellular orientation of wild-type, dominant negative mutants, and Vangl2MO morpholinos was studied by scoring the beating direction of ciliated cells and measuring the correlation with nearest neighbors. No significant difference between the mutant and wild type was found. Time lapse videos of developing cilia indicate that intracellular ordering is non-existent in early development, with ordering occurring by maturity. Further work needs to be done to determine what role fluid flow plays in that ordering.

  15. The unique paradigm of spirochete motility and chemotaxis

    PubMed Central

    Charon, Nyles W.; Cockburn, Andrew; Li, Chunhao; Liu, Jun; Miller, Kelly A.; Miller, Michael R.; Motaleb, Md.; Wolgemuth, Charles W.

    2013-01-01

    Spirochete motility is enigmatic: It differs from the motility of most other bacteria in that the entire bacterium is involved in translocation in the absence of external appendages. Using the Lyme disease spirochete Borrelia burgdorferi (Bb) as a model system, we explore the current research on spirochete motility and chemotaxis. Bb has periplasmic flagella (PFs) subterminally attached to each end of the protoplasmic cell cylinder, and surrounding the cell is an outer membrane. These internal helically shaped PFs allow the spirochete to swim by generating backward-moving waves by rotation. Exciting advances using cryoelectron microscopy tomography are presented with respect to in situ analysis of cell, PF, and motor structure. In addition, advances in the dynamics of motility, chemotaxis, gene regulation, and the role of motility and chemotaxis in the life cycle of Bb are summarized. The results indicate that the motility paradigms of flagellated bacteria do not apply to these unique bacteria. PMID:22994496

  16. Uncovering the Mystery of Gliding Motility in the Myxobacteria

    PubMed Central

    Nan, Beiyan; Zusman, David R.

    2012-01-01

    Bacterial gliding motility is the smooth movement of cells on solid surfaces unaided by flagella or pili. Many diverse groups of bacteria exhibit gliding, but the mechanism of gliding motility has remained a mystery since it was first observed more than a century ago. Recent studies on the motility of Myxococcus xanthus, a soil myxobacterium, suggest a likely mechanism for gliding in this organism. About forty M. xanthus genes were shown to be involved in gliding motility, and some of their protein products were labeled and localized within cells. These studies suggest that gliding motility in M. xanthus involves large multiprotein structural complexes, regulatory proteins, and cytoskeletal filaments. In this review, we summarize recent experiments that provide the basis for this emerging view of M. xanthus motility. We also discuss alternative models for gliding. PMID:21910630

  17. Filling an Emulsion Drop with Motile Bacteria

    NASA Astrophysics Data System (ADS)

    Vladescu, I. D.; Marsden, E. J.; Schwarz-Linek, J.; Martinez, V. A.; Arlt, J.; Morozov, A. N.; Marenduzzo, D.; Cates, M. E.; Poon, W. C. K.

    2014-12-01

    We have measured the spatial distribution of motile Escherichia coli inside spherical water droplets emulsified in oil. At low cell concentrations, the cell density peaks at the water-oil interface; at increasing concentration, the bulk of each droplet fills up uniformly while the surface peak remains. Simulations and theory show that the bulk density results from a "traffic" of cells leaving the surface layer, increasingly due to cell-cell scattering as the surface coverage rises above ˜10 %. Our findings show similarities with the physics of a rarefied gas in a spherical cavity with attractive walls.

  18. Gastrointestinal motility in space motion sickness

    NASA Technical Reports Server (NTRS)

    Thornton, William E.; Linder, Barry J.; Moore, Thomas P.; Pool, Sam L.

    1987-01-01

    Gastrointestinal symptoms in space motion sickness (SMS) are significantly different from those in ordinary motion sickness (MS). Recording and tabulation of sounds was the only technique that could be used as a measure of motility during spaceflight operations. There were 17 subjects, six unaffected by SMS, who made ambulatory recordings preflight and inflight. With one exception, all those affected had sharply reduced sounds, while those unaffected had increases or moderate reductions. The mechanism of vomiting in SMS appears to be secondary to this ileus, in contrast to vomiting in ordinary MS, where the emesis center is thought to be directly triggered by the vestibular system.

  19. Regulation of axonemal motility in demembranated equine sperm.

    PubMed

    Loux, Shavahn C; Macías-Garcia, Beatríz; González-Fernández, Lauro; Canesin, Heloisa DeSiqueira; Varner, Dickson D; Hinrichs, Katrin

    2014-12-01

    Equine in vitro fertilization is not yet successful because equine sperm do not effectively capacitate in vitro. Results of previous studies suggest that this may be due to failure of induction of hyperactivated motility in equine sperm under standard capacitating conditions. To evaluate factors directly affecting axonemal motility in equine sperm, we developed a demembranated sperm model and analyzed motility parameters in this model under different conditions using computer-assisted sperm analysis. Treatment of ejaculated equine sperm with 0.02% Triton X-100 for 30 sec maximized both permeabilization and total motility after reactivation. The presence of ATP was required for motility of demembranated sperm after reactivation, but cAMP was not. The calculated intracellular pH of intact equine sperm was 7.14 ± 0.07. Demembranated sperm showed maximal total motility at pH 7. Neither increasing pH nor increasing calcium levels, nor any interaction of the two, induced hyperactivated motility in demembranated equine sperm. Motility of demembranated sperm was maintained at free calcium concentrations as low as 27 pM, and calcium arrested sperm motility at much lower concentrations than those reported in other species. Calcium arrest of sperm motility was not accompanied by flagellar curvature, suggesting a failure of calcium to induce the tonic bend seen in other species and thought to support hyperactivated motility. This indicated an absence, or difference in calcium sensitivity, of the related asymmetric doublet-sliding proteins. These studies show a difference in response to calcium of the equine sperm axoneme to that reported in other species that may be related to the failure of equine sperm to penetrate oocytes in vitro under standard capacitating conditions. Further work is needed to determine the factors that stimulate hyperactivated motility at the axonemal level in equine sperm. PMID:25339104

  20. Chemokinetic motility responses of the cyanobacterium oscillatoria terebriformis

    NASA Technical Reports Server (NTRS)

    Richardson, Laurie L.; Castenholz, Richard W.

    1989-01-01

    Oscillatoria terebriformis, a gliding, filamentous, thermophilic cyanobacterium, exhibited an inhibition of gliding motility upon exposure to fructose. The observed response was transient, and the duration of nonmotility was directly proportional to the concentration of fructose. Upon resumption of motility, the rate of motility was also inversely proportional to the concentration of fructose. Sulfide caused a similar response. The effect of sulfide was specific and not due to either anoxia or negative redox potential. Exposure to glucose, acetate, lactate, or mat interstitial water did not elicit any motility response.

  1. Mechanics and polarity in cell motility

    NASA Astrophysics Data System (ADS)

    Ambrosi, D.; Zanzottera, A.

    2016-09-01

    The motility of a fish keratocyte on a flat substrate exhibits two distinct regimes: the non-migrating and the migrating one. In both configurations the shape is fixed in time and, when the cell is moving, the velocity is constant in magnitude and direction. Transition from a stable configuration to the other one can be produced by a mechanical or chemotactic perturbation. In order to point out the mechanical nature of such a bistable behaviour, we focus on the actin dynamics inside the cell using a minimal mathematical model. While the protein diffusion, recruitment and segregation govern the polarization process, we show that the free actin mass balance, driven by diffusion, and the polymerized actin retrograde flow, regulated by the active stress, are sufficient ingredients to account for the motile bistability. The length and velocity of the cell are predicted on the basis of the parameters of the substrate and of the cell itself. The key physical ingredient of the theory is the exchange among actin phases at the edges of the cell, that plays a central role both in kinematics and in dynamics.

  2. Regulation of macrophage motility by Irgm1

    PubMed Central

    Henry, Stanley C.; Traver, Maria; Daniell, Xiaojou; Indaram, Maanasa; Oliver, Tim; Taylor, Gregory A.

    2010-01-01

    IRG are a family of IFN-regulated proteins that are critical for resistance to infection. Mouse IRG proteins are divided into GMS and GKS subfamilies, based on a sequence within the G1 GTP-binding motif. The GMS proteins have a particularly profound impact on immunity, as typified by Irgm1, of which absence leads to a complete loss of resistance to a variety of intracellular bacteria and protozoa. The underlying molecular and cellular mechanisms are not clear. Here, we use time-lapse microscopy and cell-tracking analysis to demonstrate that Irgm1 is required for motility of IFN-γ-activated macrophages. The absence of Irgm1 led to decreased actin remodeling at the leading edge of migrating macrophages, as well as decreased Rac activation. Although Irgm1 did not localize to the leading edge of migrating macrophages, it was found to regulate the localization of a GKS IRG protein, Irgb6, which in turn, concentrated on the plasma membrane in the advancing lamellipodia, in close apposition to molecular components that regulate membrane remodeling, including Rac, paxillin, and actin. Thus, Irgm1 likely controls macrophage motility by regulating the positioning of specific GKS IRG proteins to the plasma membrane, which in turn, modulate cytoskeletal remodeling and membrane dynamics. PMID:19920210

  3. Gastrointestinal motility and functional gastrointestinal diseases.

    PubMed

    Kusano, Motoyasu; Hosaka, Hiroko; Kawada, Akiyo; Kuribayashi, Shiko; Shimoyama, Yasuyuki; Zai, Hiroaki; Kawamura, Osamu; Yamada, Masanobu

    2014-01-01

    Digestive tract motility patterns are closely related to the pathophysiology of functional gastrointestinal diseases (FGID), and these patterns differ markedly between the interdigestive period and the postprandial period. The characteristic motility pattern in the interdigestive period is so-called interdigestive migrating contraction (IMC). IMCs have a housekeeping role in the intestinal tract, and could also be related to FGID. IMCs arising from the stomach are called gastrointestinal IMCs (GI-IMC), while IMCs arising from the duodenum without associated gastric contractions are called intestinal IMCs (I-IMC). It is thought that I-IMCs are abnormal in FGID. Transport of food residue to the duodenum via gastric emptying is one of the most important postprandial functions of the stomach. In patients with functional dyspepsia (FD), abnormal gastric emptying is a possible mechanism of gastric dysfunction. Accordingly, delayed gastric emptying has attracted attention, with prokinetic agents and herbal medicines often being administered in Japan to accelerate gastric emptying in patients who have anorexia associated with dyspepsia. Recently, we found that addition of monosodium L-glutamate (MSG) to a high-calorie liquid diet rich in casein promoted gastric emptying in healthy men. Therefore, another potential method of improving delayed gastric emptying could be activation of chemosensors that stimulate the autonomic nervous system of the gastrointestinal tract, suggesting a role for MSG in the management of delayed gastric emptying in patients with FD. PMID:23886379

  4. The mechanics of motility in dissociated cytoplasm.

    PubMed Central

    Dembo, M

    1986-01-01

    We stimulate the dynamical behavior of dissociated cytoplasm using the Reactive Flow Model (Dembo, M., and F. Harlow, 1986, Biophys. J., 50:109-121). We find that for the most part the predicted dynamical behavior of the cytoplasm is governed by three nondimensional numbers. Several other nondimensional parameters, the initial conditions, and boundary conditions are found to have lesser effects. Of the three major nondimensional parameters, one (D#) controls the percentage of ectoplasm, the second (C#) controls the sharpness of the endoplasm-ectoplasm boundary, and the third (R#) controls the topological complexity of the endoplasm-ectoplasm distribution. If R# is very small, then the cytoplasm contracts into a single uniform mass, and there is no bulk streaming. If R# is very large, then the cytoplasmic mass breaks up into a number of clumps scattered throughout the available volume. Between these clumps the solution undergoes turbulent or chaotic patterns of streaming. Intermediate values of R# can be found such that the mass of cytoplasm remains connected and yet undergoes coherent modes of motility similar to flares (Taylor, D.L., J.S. Condeelis, P.L. Moore, and R.D. Allen, 1973, J. Cell Biol., 59:378-394) and rosettes (Kuroda, K., 1979, Cell Motility: Molecules and Organization, 347-362). Images FIGURE 1 FIGURE 1B FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 6 FIGURE 7 FIGURE 8 FIGURE 9 PMID:3801576

  5. Small intestine motility development in newborn mammals.

    PubMed

    Woliński, Jarosław; Słupecka-Ziemilska, Monika; Boryczka, Maria; Grzesiak, Paulina; Kwiatkowski, Jakub; Kotarba, Grzegorz

    2016-01-01

    Since the beginning of the 20th century, researchers have been working to improve the understanding of gastrointestinal motility. The first major discovery was the observation of a migrating myoelectric complex that turned out to be a universal occurrence among vertebrates. Further inquires resulted in a detailed description of its development during different stages of ontogeny. Some time before that, a cornerstone had been laid for a breakthrough that would come years later. That cornerstone came in the form of interstitial cells of Cajal whose true role could not be discerned until the discovery of a CD117 receptor - their main marker. With the ability to precisely mark interstitial cells of Cajal, a wave of subsequent new experiments and observations connected them to the occurrence of slow waves and allowed an understanding of the mechanism responsible for their generation. Some of these findings suggested that Cajal cells might have a role in the development of several motility disorders thus opening an avenue of research that requires the usage of both traditional and advanced diagnostic methods. PMID:27416626

  6. Bacterial Motility Reveals Unknown Molecular Organization.

    PubMed

    Duchesne, Ismaël; Rainville, Simon; Galstian, Tigran

    2015-11-17

    The water solubility of lyotropic liquid crystals (LCs) makes them very attractive to study the behavior of biological microorganisms in an environment where local symmetry is broken (as often encountered in nature). Several recent studies have shown a dramatic change in the behavior of flagellated bacteria when swimming in solutions of the lyotropic LC disodium cromoglycate (DSCG). In this study, the movements of Escherichia coli bacteria in DSCG-water solutions of different concentrations are observed to improve our understanding of this phenomenon. In addition, the viscosity of DSCG aqueous solutions is measured as a function of concentration at room temperature. We also experimentally identify a previously undescribed isotropic pretransition zone where bacteria start sticking to each other and to surfaces. Simple estimations show that the unbalanced osmotic pressure induced depletion force might be responsible for this sticking phenomenon. An estimate of the bacteria propulsive force and the DSCG aggregates length (versus concentration) are calculated from the measured viscosity of the medium. All these quantities are found to undergo a strong increase in the pretransition zone, starting at a threshold concentration of 6±1 wt % DSCG that is well below the known isotropic-LC transition (∼10 wt %). This study also shines light on the motility of flagellated bacteria in realistic environments, and it opens new avenues for interesting applications such as the use of motile microorganisms to probe the physical properties of their host or smart bandages that could guide bacteria out of wounds. PMID:26588572

  7. Immersed boundary-lattice Boltzmann method for simulation of muco-ciliary transport: effect of mucus depth at various amounts of cilia beat frequency

    NASA Astrophysics Data System (ADS)

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

    2015-12-01

    Numerical simulation based on immersed boundary-lattice Boltzmann method has been employed to study 2D muco-ciliary transport problem. The periciliary liquid (PCL) and mucus layers in this study are considered as the Newtonian and viscoelastic fluid respectively. An Oldroyd-B model is used as the constitutive equations of mucus layer. To simulate accurate effects of the cilia and PCL-mucus interface on the fluid, immersed boundary method is used. Numerical simulations have been performed to investigate the effects of mucus depth on the muco-ciliary clearance at various values of cilia beat frequencies. Our results show that, by increasing mucus depth, which results from air pollution and smoking, mean mucus velocity decreases. But it can be completely modified by increasing cilia beat frequency and the cilia beat frequency has great effect on the muco-ciliary clearance.

  8. EFCAB7 and IQCE regulate Hedgehog signaling by tethering the EVC-EVC2 complex to the base of primary cilia

    PubMed Central

    Pusapati, Ganesh V.; Hughes, Casey E; Dorn, Karolin V.; Zhang, Dapeng; Sugianto, Priscilla; Aravind, L.; Rohatgi, Rajat

    2014-01-01

    The Hedgehog (Hh) pathway depends on primary cilia in vertebrates, but the signaling machinery within cilia remains incompletely defined. We report the identification of a complex between two ciliary proteins, EFCAB7 and IQCE, which positively regulates the Hh pathway. The EFCAB7-IQCE module anchors the EVC-EVC2 complex in a signaling microdomain at the base of cilia. EVC and EVC2 genes are mutated in Ellis van Creveld and Weyers syndromes, characterized by impaired Hh signaling in skeletal, cardiac and orofacial tissues. EFCAB7 binds to a C-terminal disordered region in EVC2 that is deleted in Weyers patients. EFCAB7 depletion mimics the Weyers cellular phenotype— the mis-localization of EVC-EVC2 within cilia and impaired activation of the transcription factor GLI2. Evolutionary analysis suggests that emergence of these complexes might have been important for adaptation of an ancient organelle, the cilium, for an animal-specific signaling network. PMID:24582806

  9. Sirtuins: Guardians of Mammalian Healthspan

    PubMed Central

    Giblin, William; Skinner, Mary E.; Lombard, David B.

    2014-01-01

    The first link between sirtuins and longevity was made 15 years ago in yeast. These initial studies sparked efforts by many laboratories working in diverse model organisms to elucidate the relationships between sirtuins, lifespan, and age-associated dysfunction. Here we discuss the current understanding of how sirtuins relate to aging. We focus primarily on mammalian sirtuins SIRT1, SIRT3, and SIRT6, the three sirtuins for which the most relevant data are available. Strikingly, a large body of evidence now indicates that these and other mammalian sirtuins suppress a variety of age-related pathologies and promote healthspan. Moreover, increased expression of SIRT1 or SIRT6 extends mouse lifespan. Overall, these data point to important roles for sirtuins in promoting mammalian health, and perhaps in modulating the aging process. PMID:24877878

  10. Electroporation into Cultured Mammalian Embryos

    NASA Astrophysics Data System (ADS)

    Nomura, Tadashi; Takahashi, Masanori; Osumi, Noriko

    Over the last century, mammalian embryos have been used extensively as a common animal model to investigate fundamental questions in the field of developmental biology. More recently, the establishment of transgenic and gene-targeting systems in laboratory mice has enabled researchers to unveil the genetic mechanisms under lying complex developmental processes (Mak, 2007). However, our understanding of cell—cell interactions and their molecular basis in the early stages of mammalian embryogenesis is still very fragmentary. One of the major problems is the difficulty of precise manipulation and limited accessibility to mammalian embryos via uterus wall. Unfortunately, existing tissue and organotypic culture systems per se do not fully recapitulate three-dimensional, dynamic processes of organogenesis observed in vivo. Although transgenic animal technology and virus-mediated gene delivery are useful to manipulate gene expression, these techniques take much time and financial costs, which limit their use.

  11. Hook Adaptors Induce Unidirectional Processive Motility by Enhancing the Dynein-Dynactin Interaction.

    PubMed

    Olenick, Mara A; Tokito, Mariko; Boczkowska, Malgorzata; Dominguez, Roberto; Holzbaur, Erika L F

    2016-08-26

    Cytoplasmic dynein drives the majority of minus end-directed vesicular and organelle motility in the cell. However, it remains unclear how dynein is spatially and temporally regulated given the variety of cargo that must be properly localized to maintain cellular function. Recent work has suggested that adaptor proteins provide a mechanism for cargo-specific regulation of motors. Of particular interest, studies in fungal systems have implicated Hook proteins in the regulation of microtubule motors. Here we investigate the role of mammalian Hook proteins, Hook1 and Hook3, as potential motor adaptors. We used optogenetic approaches to specifically recruit Hook proteins to organelles and observed rapid transport of peroxisomes to the perinuclear region of the cell. This rapid and efficient translocation of peroxisomes to microtubule minus ends indicates that mammalian Hook proteins activate dynein rather than kinesin motors. Biochemical studies indicate that Hook proteins interact with both dynein and dynactin, stabilizing the formation of a supramolecular complex. Complex formation requires the N-terminal domain of Hook proteins, which resembles the calponin-homology domain of end-binding (EB) proteins but cannot bind directly to microtubules. Single-molecule motility assays using total internal reflection fluorescence microscopy indicate that both Hook1 and Hook3 effectively activate cytoplasmic dynein, inducing longer run lengths and higher velocities than the previously characterized dynein activator bicaudal D2 (BICD2). Together, these results suggest that dynein adaptors can differentially regulate dynein to allow for organelle-specific tuning of the motor for precise intracellular trafficking. PMID:27365401

  12. Protective effect of vitamin E on sperm motility and oxidative stress in valproic acid treated rats.

    PubMed

    Ourique, Giovana M; Saccol, Etiane M H; Pês, Tanise S; Glanzner, Werner G; Schiefelbein, Sun Hee; Woehl, Viviane M; Baldisserotto, Bernardo; Pavanato, Maria A; Gonçalves, Paulo B D; Barreto, Kátia P

    2016-09-01

    Long-term administration of valproic acid (VPA) is known to promote reproductive impairment mediated by increase in testicular oxidative stress. Vitamin E (VitE) is a lipophilic antioxidant known to be essential for mammalian spermatogenesis. However, the capacity of this vitamin to abrogate the VPA-mediated oxidative stress has not yet been assessed. In the current study, we evaluated the protective effect of VitE on functional abnormalities related to VPA-induced oxidative stress in the male reproductive system. VPA (400 mg kg(-1)) was administered by gavage and VitE (50 mg kg(-1)) intraperitoneally to male Wistar rats for 28 days. Analysis of spermatozoa from the cauda epididymides was performed. The testes and epididymides were collected for measurement of oxidative stress biomarkers. Treatment with VPA induced a decrease in sperm motility accompanied by an increase in oxidative damage to lipids and proteins, depletion of reduced glutathione and a decrease in total reactive antioxidant potential on testes and epididymides. Co-administration of VitE restored the antioxidant potential and prevented oxidative damage on testes and epididymides, restoring sperm motility. Thus, VitE protects the reproductive system from the VPA-induced damage, suggesting that it may be a useful compound to minimize the reproductive impairment in patients requiring long-term treatment with VPA. PMID:27424124

  13. DOCK2 is a Rac activator that regulates motility and polarity during neutrophil chemotaxis.

    PubMed

    Kunisaki, Yuya; Nishikimi, Akihiko; Tanaka, Yoshihiko; Takii, Ryosuke; Noda, Mayuko; Inayoshi, Ayumi; Watanabe, Ken-ichi; Sanematsu, Fumiyuki; Sasazuki, Takehiko; Sasaki, Takehiko; Fukui, Yoshinori

    2006-08-28

    Neutrophils are highly motile leukocytes, and they play important roles in the innate immune response to invading pathogens. Neutrophil chemotaxis requires Rac activation, yet the Rac activators functioning downstream of chemoattractant receptors remain to be determined. We show that DOCK2, which is a mammalian homologue of Caenorhabditis elegans CED-5 and Drosophila melanogaster Myoblast City, regulates motility and polarity during neutrophil chemotaxis. Although DOCK2-deficient neutrophils moved toward the chemoattractant source, they exhibited abnormal migratory behavior with a marked reduction in translocation speed. In DOCK2-deficient neutrophils, chemoattractant-induced activation of both Rac1 and Rac2 were severely impaired, resulting in the loss of polarized accumulation of F-actin and phosphatidylinositol 3,4,5-triphosphate (PIP3) at the leading edge. On the other hand, we found that DOCK2 associates with PIP3 and translocates to the leading edge of chemotaxing neutrophils in a phosphatidylinositol 3-kinase (PI3K)-dependent manner. These results indicate that during neutrophil chemotaxis DOCK2 regulates leading edge formation through PIP3-dependent membrane translocation and Rac activation. PMID:16943182

  14. Multiple roles of Pseudomonas aeruginosa TBCF10839 PilY1 in motility, transport and infection

    PubMed Central

    Bohn, Yu-Sing Tammy; Brandes, Gudrun; Rakhimova, Elza; Horatzek, Sonja; Salunkhe, Prabhakar; Munder, Antje; van Barneveld, Andrea; Jordan, Doris; Bredenbruch, Florian; Häußler, Susanne; Riedel, Kathrin; Eberl, Leo; Jensen, Peter Østrup; Bjarnsholt, Thomas; Moser, Claus; Hoiby, Niels; Tümmler, Burkhard; Wiehlmann, Lutz

    2008-01-01

    Polymorphonuclear neutrophils are the most important mammalian host defence cells against infections with Pseudomonas aeruginosa. Screening of a signature tagged mutagenesis library of the non-piliated P. aeruginosa strain TBCF10839 uncovered that transposon inactivation of its pilY1 gene rendered the bacterium more resistant against killing by neutrophils than the wild type and any other of the more than 3000 tested mutants. Inactivation of pilY1 led to the loss of twitching motility in twitching-proficient wild-type PA14 and PAO1 strains, predisposed to autolysis and impaired the secretion of quinolones and pyocyanin, but on the other hand promoted growth in stationary phase and bacterial survival in murine airway infection models. The PilY1 population consisted of a major full-length and a minor shorter PilY1* isoform. PilY1* was detectable in small extracellular quinolone-positive aggregates, but not in the pilus. P. aeruginosa PilY1 is not an adhesin on the pilus tip, but assists in pilus biogenesis, twitching motility, secretion of secondary metabolites and in the control of cell density in the bacterial population. PMID:19054330

  15. Gene expression in Pseudomonas aeruginosa swarming motility

    PubMed Central

    2010-01-01

    Background The bacterium Pseudomonas aeruginosa is capable of three types of motilities: swimming, twitching and swarming. The latter is characterized by a fast and coordinated group movement over a semi-solid surface resulting from intercellular interactions and morphological differentiation. A striking feature of swarming motility is the complex fractal-like patterns displayed by migrating bacteria while they move away from their inoculation point. This type of group behaviour is still poorly understood and its characterization provides important information on bacterial structured communities such as biofilms. Using GeneChip® Affymetrix microarrays, we obtained the transcriptomic profiles of both bacterial populations located at the tip of migrating tendrils and swarm center of swarming colonies and compared these profiles to that of a bacterial control population grown on the same media but solidified to not allow swarming motility. Results Microarray raw data were corrected for background noise with the RMA algorithm and quantile normalized. Differentially expressed genes between the three conditions were selected using a threshold of 1.5 log2-fold, which gave a total of 378 selected genes (6.3% of the predicted open reading frames of strain PA14). Major shifts in gene expression patterns are observed in each growth conditions, highlighting the presence of distinct bacterial subpopulations within a swarming colony (tendril tips vs. swarm center). Unexpectedly, microarrays expression data reveal that a minority of genes are up-regulated in tendril tip populations. Among them, we found energy metabolism, ribosomal protein and transport of small molecules related genes. On the other hand, many well-known virulence factors genes were globally repressed in tendril tip cells. Swarm center cells are distinct and appear to be under oxidative and copper stress responses. Conclusions Results reported in this study show that, as opposed to swarm center cells, tendril

  16. Breakup and then makeup: a predictive model of how cilia self-regulate hardness for posture control

    NASA Astrophysics Data System (ADS)

    Bandyopadhyay, Promode R.; Hansen, Joshua C.

    2013-06-01

    Functioning as sensors and propulsors, cilia are evolutionarily conserved organelles having a highly organized internal structure. How a paramecium's cilium produces off-propulsion-plane curvature during its return stroke for symmetry breaking and drag reduction is not known. We explain these cilium deformations by developing a torsional pendulum model of beat frequency dependence on viscosity and an olivo-cerebellar model of self-regulation of posture control. The phase dependence of cilia torsion is determined, and a bio-physical model of hardness control with predictive features is offered. Crossbridge links between the central microtubule pair harden the cilium during the power stroke; this stroke's end is a critical phase during which ATP molecules soften the crossbridge-microtubule attachment at the cilium inflection point where torsion is at its maximum. A precipitous reduction in hardness ensues, signaling the start of ATP hydrolysis that re-hardens the cilium. The cilium attractor basin could be used as reference for perturbation sensing.

  17. Quantification of motility of carabid beetles in farmland.

    PubMed

    Allema, A B; van der Werf, W; Groot, J C J; Hemerik, L; Gort, G; Rossing, W A H; van Lenteren, J C

    2015-04-01

    Quantification of the movement of insects at field and landscape levels helps us to understand their ecology and ecological functions. We conducted a meta-analysis on movement of carabid beetles (Coleoptera: Carabidae), to identify key factors affecting movement and population redistribution. We characterize the rate of redistribution using motility μ (L2 T-1), which is a measure for diffusion of a population in space and time that is consistent with ecological diffusion theory and which can be used for upscaling short-term data to longer time frames. Formulas are provided to calculate motility from literature data on movement distances. A field experiment was conducted to measure the redistribution of mass-released carabid, Pterostichus melanarius in a crop field, and derive motility by fitting a Fokker-Planck diffusion model using inverse modelling. Bias in estimates of motility from literature data is elucidated using the data from the field experiment as a case study. The meta-analysis showed that motility is 5.6 times as high in farmland as in woody habitat. Species associated with forested habitats had greater motility than species associated with open field habitats, both in arable land and woody habitat. The meta-analysis did not identify consistent differences in motility at the species level, or between clusters of larger and smaller beetles. The results presented here provide a basis for calculating time-varying distribution patterns of carabids in farmland and woody habitat. The formulas for calculating motility can be used for other taxa. PMID:25673121

  18. Laser radiation and motility patterns of human sperm

    SciTech Connect

    Lenzi, A.; Claroni, F.; Gandini, L.; Lombardo, F.; Barbieri, C.; Lino, A.; Dondero, F. )

    1989-01-01

    Human sperm were exposed in vitro to laser radiation. An increase in progressive sperm motility was associated with a faster rate of sperm ATP consumption. Computer-assisted analysis of sperm motility confirmed the positive effect of laser irradiation on velocity and linearity of sperm.

  19. Mechanics model for actin-based motility

    NASA Astrophysics Data System (ADS)

    Lin, Yuan

    2009-02-01

    We present here a mechanics model for the force generation by actin polymerization. The possible adhesions between the actin filaments and the load surface, as well as the nucleation and capping of filament tips, are included in this model on top of the well-known elastic Brownian ratchet formulation. A closed form solution is provided from which the force-velocity relationship, summarizing the mechanics of polymerization, can be drawn. Model predictions on the velocity of moving beads driven by actin polymerization are consistent with experiment observations. This model also seems capable of explaining the enhanced actin-based motility of Listeria monocytogenes and beads by the presence of Vasodilator-stimulated phosphoprotein, as observed in recent experiments.

  20. Metabolism and motility in prebiotic structures

    PubMed Central

    Hanczyc, Martin M.

    2011-01-01

    Easily accessible, primitive chemical structures produced by self-assembly of hydrophobic substances into oil droplets may result in self-moving agents able to sense their environment and move to avoid equilibrium. These structures would constitute very primitive examples of life on the Earth, even more primitive than simple bilayer vesicle structures. A few examples of simple chemical systems are presented that self-organize to produce oil droplets capable of movement, environment remodelling and primitive chemotaxis. These chemical agents are powered by an internal chemical reaction based on the hydrolysis of an oleic anhydride precursor or on the hydrolysis of hydrogen cyanide (HCN) polymer, a plausible prebiotic chemistry. Results are presented on both the behaviour of such droplets and the surface-active properties of HCN polymer products. Such motile agents would be capable of finding resources while escaping equilibrium and sustaining themselves through an internal metabolism, thus providing a working chemical model for a possible origin of life. PMID:21930579

  1. Method and system for enhancing microbial motility

    SciTech Connect

    Hazen, T.C.; Lopez-De-Victoria, G.

    1992-12-31

    A method and system for enhancing the motility of microorganisms by placing an effective amount of chlorinated hydrocarbons, preferably chlorinated alkenes, and most preferably trichloroethylene in spaced relation to the microbes so that the surprisingly strong, monomodal, chemotactic response of the chlorinated hydrocarbon on subsurface microbes can draw the microbes away from or towards and into a substance, as desired. In remediation of groundwater pollution, for example, TCE can be injected into the plume to increase the population of microbes at the plume whereby the plume can be more quickly degraded. A TCE-degrading microbe, such as Welchia alkenophilia, can be used to degrade the TCE following the degradation of the original pollutant.

  2. Evolutionary aspects of collective motility in pathogenic bacteria

    NASA Astrophysics Data System (ADS)

    Deforet, Maxime; Xavier, Joao

    Pseudomonas aeruginosa is a pathogenic bacteria that can use its single polar flagellum to swim through liquids. It can move collectively over semisolid surfaces, a behavior called swarming. It can also settle and form surface-attached communities called biofilms that protect them from antibiotics. The transition from single motility (swimming) to collective motility (swarming) is biologically relevant as it enables exploring environments that a single bacterium cannot explore on its own. It is also clinically relevant since swarming and biofilm formation are thought to be antagonistic. We investigate the mechanisms of bacterial collective motility using a multidisciplinary approach that combines mathematical modeling, quantitative experiments, and microbial genetics. We aim to identify how these mechanisms may evolve under the selective pressure of population expansion, and consequently reinforce or hinder collective motility. In particular, we clarify the role of growth rate and motility in invasive populations.

  3. A mechanism for cell motility by active polar gels

    PubMed Central

    Marth, W.; Praetorius, S.; Voigt, A.

    2015-01-01

    We analyse a generic motility model, with the motility mechanism arising by contractile stress due to the interaction of myosin and actin. A hydrodynamic active polar gel theory is used to model the cytoplasm of a cell and is combined with a Helfrich-type model to account for membrane properties. The overall model allows consideration of the motility without the necessity for local adhesion. Besides a detailed numerical approach together with convergence studies for the highly nonlinear free boundary problem, we also compare the induced flow field of the motile cell with that of classical squirmer models and identify the motile cell as a puller or pusher, depending on the strength of the myosin–actin interactions. PMID:25926698

  4. Thyroxin Is Useful to Improve Sperm Motility

    PubMed Central

    Mendeluk, Gabriela Ruth; Rosales, Mónica

    2016-01-01

    Background The aim of this study was to evaluate the non-genomic action of thyroxin on sperm kinetic and its probable use to improve sperm recovery after applying an en- richment method like “swim-up” in comparison with the available one, pentoxifylline. Materials and Methods This is an experimental study. A total of 50 patients were re- cruited, followed by infertility consultation. Conventional sperm assays were performed according to World Health Organization criteria-2010 (WHO-2010). A Computer Aided Semen Analysis System was employed to assess kinetic parameters and concentrations. Number of the motile sperm recovered after preparation technique was calculated. Results Addition of T4 (0.002 µg/ml) to semen samples increased hypermotility at 20 minutes (control: 14.18 ± 5.1% vs. 17.66 ± 8.88%, P<0.03, data expressed as mean ± SD) and remained unchanged after 40 minutes. Significant differences were found in the motile sperm recovered after swim-up (control: 8.93×106 ± 9.52× 06vs. 17.20×106 ± 21.16×106, P<0.03), achieving all of the tested samples a desirable threshold value for artificial insemination outcome, while adding pentoxifylline increased the number of recovered sperm after swim-up in 60% of the studied cases. No synergism between two treatments could be determined. Conclusion We propose a new physiological tool to artificially improve insemination. The discussion opens windows to investigate unknown pathways involved in sperm ca- pacitation and gives innovative arguments to better understand infertility mechanisms. PMID:27441054

  5. Hydrodynamics of helical-shaped bacterial motility

    NASA Astrophysics Data System (ADS)

    Wada, Hirofumi; Netz, Roland R.

    2009-08-01

    To reveal the underlying hydrodynamic mechanism for the directed propulsion of the bacterium Spiroplasma, we formulate a coarse-grained elastic polymer model with domains of alternating helicities along the contour. Using hydrodynamic simulations and analytic arguments, we show that the propagation of helical domain walls leads to the directed propulsion of the cell body opposite to the domain-wall traveling direction. Several key features of Spiroplasma motility are reproduced by our model. We in particular show that the helical pitch angle observed for Spiroplasma meliferum, ψ=35° , is optimized for maximal swimming speed and energy-conversion efficiency. Our analytic theory based on the slender-body hydrodynamic approximation agrees very well with our numerical data demonstrating how the chirality switch propagating along the helical cell body is converted to a translational thrust for the cell body itself. We in detail consider thermal effects on the propulsion efficiency in the form of orientational fluctuations and conformational fluctuations of the helix shape. The body length dependence of the cell motility is studied numerically and compared to our approximate analytic theory. For fixed pitch angle ψ=35° , the swimming speed is maximized at a ratio of cell-body length to domain length of about 2-3, which are typical values for real cells. We also propose simple analytic arguments for an enhancement of the swimming velocity with increasing solution viscosity by taking into account the effects of transient confinement of a helical cell body in a polymeric meshwork. Comparison with a generalized theory for the swimming speed of flagellated bacteria in polymeric meshworks shows that the presence of a finite-sized bacterial head gives rise to a maximal swimming speed at a finite solution viscosity, whereas in the absence of a head the swimming speed monotonically increases with increasing viscosity.

  6. Kinetics of Hedgehog-Dependent Full-Length Gli3 Accumulation in Primary Cilia and Subsequent Degradation ▿ † ‡

    PubMed Central

    Wen, Xiaohui; Lai, Cary K.; Evangelista, Marie; Hongo, Jo-Anne; de Sauvage, Frederic J.; Scales, Suzie J.

    2010-01-01

    Hedgehog (Hh) signaling in vertebrates depends on intraflagellar transport (IFT) within primary cilia. The Hh receptor Patched is found in cilia in the absence of Hh and is replaced by the signal transducer Smoothened within an hour of Hh stimulation. By generating antibodies capable of detecting endogenous pathway transcription factors Gli2 and Gli3, we monitored their kinetics of accumulation in cilia upon Hh stimulation. Localization occurs within minutes of Hh addition, making it the fastest reported readout of pathway activity, which permits more precise temporal and spatial localization of Hh signaling events. We show that the species of Gli3 that accumulates at cilium tips is full-length and likely not protein kinase A phosphorylated. We also confirmed that phosphorylation and βTrCP/Cul1 are required for endogenous Gli3 processing and that this is inhibited by Hh. Surprisingly, however, Hh-dependent inhibition of processing does not lead to accumulation of full-length Gli3, but instead renders it labile, leading to its proteasomal degradation via the SPOP/Cul3 complex. In fact, full-length Gli3 disappears with faster kinetics than the Gli3 repressor, the latter not requiring SPOP/Cul3 or βTrCP/Cul1. This may contribute to the increased Gli3 activator/repressor ratios found in IFT mutants. PMID:20154143

  7. A mechanism for vertebrate Hedgehog signaling: recruitment to cilia and dissociation of SuFu–Gli protein complexes

    PubMed Central

    Tukachinsky, Hanna; Lopez, Lyle V.

    2010-01-01

    In vertebrates, Hedgehog (Hh) signaling initiated in primary cilia activates the membrane protein Smoothened (Smo) and leads to activation of Gli proteins, the transcriptional effectors of the pathway. In the absence of signaling, Gli proteins are inhibited by the cytoplasmic protein Suppressor of Fused (SuFu). It is unclear how Hh activates Gli and whether it directly regulates SuFu. We find that Hh stimulation quickly recruits endogenous SuFu–Gli complexes to cilia, suggesting a model in which Smo activates Gli by relieving inhibition by SuFu. In support of this model, we find that Hh causes rapid dissociation of the SuFu–Gli complex, thus allowing Gli to enter the nucleus and activate transcription. Activation of protein kinase A (PKA), an inhibitor of Hh signaling, blocks ciliary localization of SuFu–Gli complexes, which in turn prevents their dissociation by signaling. Our results support a simple mechanism in which Hh signals at vertebrate cilia cause dissociation of inactive SuFu–Gli complexes, a process inhibited by PKA. PMID:20956384

  8. Biological and Chemical Removal of Primary Cilia Affects Mechanical Activation of Chondrogenesis Markers in Chondroprogenitors and Hypertrophic Chondrocytes

    PubMed Central

    Deren, Matthew E.; Yang, Xu; Guan, Yingjie; Chen, Qian

    2016-01-01

    Chondroprogenitors and hypertrophic chondrocytes, which are the first and last stages of the chondrocyte differentiation process, respectively, are sensitive to mechanical signals. We hypothesize that the mechanical sensitivity of these cells depends on the cell surface primary cilia. To test this hypothesis, we removed the primary cilia by biological means with transfection with intraflagellar transport protein 88 (IFT88) siRNA or by chemical means with chloral hydrate treatment. Transfection of IFT88 siRNA significantly reduced the percentage of ciliated cells in both chondroprogenitor ATDC5 cells as well as primary hypertrophic chondrocytes. Cyclic loading (1 Hz, 10% matrix deformation) of ATDC5 cells in three-dimensional (3D) culture stimulates the mRNA levels of chondrogenesis marker Type II collagen (Col II), hypertrophic chondrocyte marker Type X collagen (Col X), and a molecular regulator of chondrogenesis and chondrocyte hypertrophy bone morphogenetic protein 2 (BMP-2). The reduction of ciliated chondroprogenitors abolishes mechanical stimulation of Col II, Col X, and BMP-2. In contrast, cyclic loading stimulates Col X mRNA levels in hypertrophic chondrocytes, but not those of Col II and BMP-2. Both biological and chemical reduction of ciliated hypertrophic chondrocytes reduced but failed to abolish mechanical stimulation of Col X mRNA levels. Thus, primary cilia play a major role in mechanical stimulation of chondrogenesis and chondrocyte hypertrophy in chondroprogenitor cells and at least a partial role in hypertrophic chondrocytes. PMID:26861287

  9. Biochemical studies of olfaction: binding specificity of odorants to a cilia preparation from rainbow trout olfactory rosettes.

    PubMed

    Rhein, L D; Cagan, R H

    1983-08-01

    Cilia isolated from the olfactory epithelium (olfactory rosettes) of rainbow trout (Salmo gairdneri) bind amino acids, which are odor stimuli to this species. We demonstrate that L-threonine, L-serine, and L-alanine bind to a common site, TSA, in the cilia preparation. All possible mixtures of two of the amino acids as competitors, with the third as the 3H-labeled ligand, were studied. The effect of two combined (unlabeled) competitors was always substantially less than additive compared with their actions singly. Along with additional inhibition studies using mixtures of inhibitors, the data show that the three odorants must interact with at least one common binding site, TSA. Binding of L-[3H]lysine to site L was unaffected by addition of L-threonine, L-serine, or L-alanine, establishing its independence from site TSA. L-Arginine inhibited binding of L-[3H]lysine, showing that both of these basic amino acids interact with site L. The data establish the presence, in trout olfactory cilia, of at least two separate and noninteracting populations of odorant binding sites, TSA and L. PMID:6409997

  10. Detection of planar polarity proteins in mammalian cochlea.

    PubMed

    Montcouquiol, Mireille; Jones, Jennifer M; Sans, Nathalie

    2008-01-01

    The "core genes" were identified as a group of genes believed to function as a conserved signaling cassette for the specification of planar polarity in Drosophila Melanogaster, and includes frizzled (fz), van gogh (vang) or strabismus (stbm), prickle (Pk), dishevelled (dsh), flamingo (fmi), and diego. The mutation of each of these genes not only causes the disruption of planar polarity within the wing or the eye of the animal, but also affects the localization of all the other protein members of the core group. These properties emphasize the importance of the interrelations between the proteins of this group. All of these core genes have homologs in vertebrates. Studies in Danio Rerio (zebrafish) and Xenopus laevis (frog) have uncovered other roles for some of these molecules in gastrulation and neurulation, during which the shape of a given tissue will undergo major transformation through cell movements. A disruption in these processes can lead to severe neural tube defects in diverse organisms, including humans. In fact, a large body of evidence suggests that planar polarity proteins are not involved in one specific cascade but in many different ones and many different mechanisms such as, but not limited to, hair or cilia orientation, asymmetric division, cellular movements, or neuronal migration. In mice cochleae, mutations in planar polarity genes lead to defects in the orientation of the stereociliary bundles at the apex of each hair cell. This phenotype established the cochlea as one of the clearest examples of planar polarity in mammals. Although significant progress has been made toward understanding the molecular basis required for the development of planar polarity in invertebrates, similar advances in vertebrates are more recent and rely mainly on the identification of a group of mammalian mutants that affect hair cell stereociliary bundle orientation. These include mutation of vangl2, scrb1, celsr1, PTK-7, dvl1-2, and more recently fz3 and fz6. In this

  11. Bacterial signaling and motility: Sure bets

    SciTech Connect

    Zhulin, Igor B

    2008-01-01

    The IX International Conference on Bacterial Locomotion and Signal Transduction (BLAST IX) was held from 14 to 19 January 2007 in Laughlin, NV, a town in the Mojave Desert on the Nevada-Arizona border near old Route 66 and along the banks of the Colorado River. This area is a home to rattlesnakes, sagebrush, abandoned gold mines, and compulsive gamblers. What better venue could scientists possibly dream of for a professional meeting? So there they were, about 190 scientists gathered in the Aquarius Casino Resort, the largest hotel and casino in Laughlin, discussing the latest advances in the field. Aside from a brief excursion to an abandoned gold mine and a dinner cruise on the Colorado River, the scientists focused on nothing but their data and hypotheses, in spirited arguments and rebuttals, and outlined their visions and future plans in a friendly and open environment. The BLAST IX program was dense, with nearly 50 talks and over 90 posters. For that reason, this meeting report will not attempt to be comprehensive; instead it will first provide general background information on the central topics of the meeting and then highlight only a few talks that were of special interest to us and hopefully to the wider scientific community. We will also attempt to articulate some of the future directions or perspectives to the best of our abilities. The best known and understood bacterial motility mechanism is swimming powered by flagella. The rotation of bacterial flagella drives this form of bacterial movement in an aqueous environment. A bacterial flagellum consists of a helical filament attached to the cell body through a complex structure known as the hook-basal body, which drives flagellar rotation. The essential components of the basal body are the MotA-MotB motor-stator proteins bound to the cytoplasmic membrane. These stator proteins interact with proteins that comprise the supramembrane and cytoplasmic rings, which are components of the motor imbedded in the

  12. DNA repair in mammalian embryos.

    PubMed

    Jaroudi, Souraya; SenGupta, Sioban

    2007-01-01

    Mammalian cells have developed complex mechanisms to identify DNA damage and activate the required response to maintain genome integrity. Those mechanisms include DNA damage detection, DNA repair, cell cycle arrest and apoptosis which operate together to protect the conceptus from DNA damage originating either in parental gametes or in the embryo's somatic cells. DNA repair in the newly fertilized preimplantation embryo is believed to rely entirely on the oocyte's machinery (mRNAs and proteins deposited and stored prior to ovulation). DNA repair genes have been shown to be expressed in the early stages of mammalian development. The survival of the embryo necessitates that the oocyte be sufficiently equipped with maternal stored products and that embryonic gene expression commences at the correct time. A Medline based literature search was performed using the keywords 'DNA repair' and 'embryo development' or 'gametogenesis' (publication dates between 1995 and 2006). Mammalian studies which investigated gene expression were selected. Further articles were acquired from the citations in the articles obtained from the preliminary Medline search. This paper reviews mammalian DNA repair from gametogenesis to preimplantation embryos to late gestational stages. PMID:17141556

  13. Slow motility in hair cells of the frog amphibian papilla: Ca2+-dependent shape changes.

    PubMed

    Farahbakhsh, Nasser A; Narins, Peter M

    2006-02-01

    We investigated the process of slow motility in non-mammalian auditory hair cells by recording the time course of shape change in hair cells of the frog amphibian papilla. The tall hair cells in the rostral segment of this organ, reported to be the sole recipients of efferent innervation, were found to shorten in response to an increase in the concentration of the intracellular free calcium. These shortenings are composed of two partially-overlapping phases: an initial rapid iso-volumetric contraction, followed by a slower length decrease accompanied with swelling. It is possible to unmask the iso-volumetric contraction by delaying the cell swelling with the help of K+ or Cl- channel inhibitors, quinine or furosemide. Furthermore, it appears that the longitudinal contraction in these cells is Ca2+-calmodulin-dependent: in the presence of W-7, a calmodulin inhibitor, only a slow, swelling phase could be observed. These findings suggest that amphibian rostral AP hair cells resemble their mammalian counterparts in expressing both a Ca2+-calmodulin-dependent contractile structure and an "osmotic" mechanism capable of mediating length change in response to extracellular stimuli. Such a mechanism might be utilized by the efferent neurotransmitters for adaptive modulation of mechano-electrical transduction, sensitivity enhancement, frequency selectivity, and protection against over-stimulation. PMID:16426781

  14. Flash Photolysis of Caged Compounds in the Cilia of Olfactory Sensory Neurons

    PubMed Central

    Boccaccio, Anna; Sagheddu, Claudia; Menini, Anna

    2011-01-01

    Photolysis of caged compounds allows the production of rapid and localized increases in the concentration of various physiologically active compounds1. Caged compounds are molecules made physiologically inactive by a chemical cage that can be broken by a flash of ultraviolet light. Here, we show how to obtain patch-clamp recordings combined with photolysis of caged compounds for the study of olfactory transduction in dissociated mouse olfactory sensory neurons. The process of olfactory transduction (Figure 1) takes place in the cilia of olfactory sensory neurons, where odorant binding to receptors leads to the increase of cAMP that opens cyclic nucleotide-gated (CNG) channels2. Ca entry through CNG channels activates Ca-activated Cl channels. We show how to dissociate neurons from the mouse olfactory epithelium3 and how to activate CNG channels or Ca-activated Cl channels by photolysis of caged cAMP4 or caged Ca5. We use a flash lamp6,7 to apply ultraviolet flashes to the ciliary region to uncage cAMP or Ca while patch-clamp recordings are taken to measure the current in the whole-cell voltage-clamp configuration8-11. PMID:22064384

  15. Primary cilia act as mechanosensors during bone healing around an implant.

    PubMed

    Leucht, P; Monica, S D; Temiyasathit, S; Lenton, K; Manu, A; Longaker, M T; Jacobs, C R; Spilker, R L; Guo, H; Brunski, J B; Helms, J A

    2013-03-01

    The primary cilium is an organelle that senses cues in a cell's local environment. Some of these cues constitute molecular signals; here, we investigate the extent to which primary cilia can also sense mechanical stimuli. We used a conditional approach to delete Kif3a in pre-osteoblasts and then employed a motion device that generated a spatial distribution of strain around an intra-osseous implant positioned in the mouse tibia. We correlated interfacial strain fields with cell behaviors ranging from proliferation through all stages of osteogenic differentiation. We found that peri-implant cells in the Col1Cre;Kif3a(fl/fl) mice were unable to proliferate in response to a mechanical stimulus, failed to deposit and then orient collagen fibers to the strain fields caused by implant displacement, and failed to differentiate into bone-forming osteoblasts. Collectively, these data demonstrate that the lack of a functioning primary cilium blunts the normal response of a cell to a defined mechanical stimulus. The ability to manipulate the genetic background of peri-implant cells within the context of a whole, living tissue provides a rare opportunity to explore mechanotransduction from a multi-scale perspective. PMID:22784673

  16. Emergence of polar order and cooperativity in hydrodynamically coupled model cilia

    PubMed Central

    Bruot, Nicolas; Cicuta, Pietro

    2013-01-01

    As a model of ciliary beat, we use two-state oscillators that have a defined direction of oscillation and have strong synchronization properties. By allowing the direction of oscillation to vary according to the interaction with the fluid, with a timescale longer than the timescale of synchronization, we show in simulations that several oscillators can align in a direction set by the geometrical configuration of the system. In this system, the alignment depends on the state of synchronization of the system, and is therefore linked to the beat pattern of the model cilia. By testing various configurations from two to 64 oscillators, we deduce empirically that, when the synchronization state of neighbouring oscillators is in phase, the angles of the oscillators align in a configuration of high hydrodynamic coupling. In arrays of oscillators that break the planar symmetry, a global direction of alignment emerges reflecting this polarity. In symmetric configurations, where several directions are geometrically equivalent, the array still displays strong internal cooperative behaviour. It also appears that the shape of the array is more important than the lattice type and orientation in determining the preferred direction. PMID:23883957

  17. Mutations in DYNC2LI1 disrupt cilia function and cause short rib polydactyly syndrome

    PubMed Central

    Taylor, S. Paige; Dantas, Tiago J.; Duran, Ivan; Wu, Sulin; Lachman, Ralph S.; Nelson, Stanley F.; Cohn, Daniel H.; Vallee, Richard B.; Krakow, Deborah

    2015-01-01

    The short rib polydactyly syndromes (SRPS) are a heterogeneous group of autosomal recessive, perinatal-lethal skeletal disorders characterized primarily by short, horizontal ribs, short limbs, and poly-dactyly. Mutations in several genes affecting intraflagellar transport (IFT) cause SRPS but they do not account for all cases. Here we identify additional SRPS genes and further unravel the functional basis for IFT. We perform whole exome sequencing and identify mutations in a new disease-producing gene, cytoplasmic dynein-2 light intermediate chain 1, DYNC2LI1, segregating with disease in three families. Using primary fibroblasts, we show that DYNC2LI1 is essential for dynein-2 complex stability and that mutations in DYNC2LI1 result in variable-length, including hyperelongated, cilia, Hedgehog pathway impairment, and ciliary IFT accumulations. The findings in this study expand our understanding of SRPS locus heterogeneity and demonstrate the importance of DYNC2LI1 in dynein-2 complex stability, cilium function, Hedgehog regulation, and skeletogenesis. PMID:26077881

  18. Remodeling Cildb, a popular database for cilia and links for ciliopathies

    PubMed Central

    2014-01-01

    Background New generation technologies in cell and molecular biology generate large amounts of data hard to exploit for individual proteins. This is particularly true for ciliary and centrosomal research. Cildb is a multi–species knowledgebase gathering high throughput studies, which allows advanced searches to identify proteins involved in centrosome, basal body or cilia biogenesis, composition and function. Combined to localization of genetic diseases on human chromosomes given by OMIM links, candidate ciliopathy proteins can be compiled through Cildb searches. Methods Othology between recent versions of the whole proteomes was computed using Inparanoid and ciliary high throughput studies were remapped on these recent versions. Results Due to constant evolution of the ciliary and centrosomal field, Cildb has been recently upgraded twice, with new species whole proteomes and new ciliary studies, and the latter version displays a novel BioMart interface, much more intuitive than the previous ones. Conclusions This already popular database is designed now for easier use and is up to date in regard to high throughput ciliary studies. PMID:25422781

  19. Ionic control of the reversal response of cilia in Paramecium caudatum. A calcium hypothesis.

    PubMed

    Naitoh, Y

    1968-01-01

    The duration of ciliary reversal of Paramecium caudatum in response to changes in external ionic factors was determined with various ionic compositions of both equilibration and stimulation media. The reversal response was found to occur when calcium ions bound by an inferred cellular cation exchange system were liberated in exchange for externally applied cations other than calcium. Factors which affect the duration of the response were (a) initial amount of calcium bound by the cation exchange system, (b) final amount of calcium bound by the system after equilibration with the stimulation medium, and (c) concentration of calcium ions in the stimulation medium. An empirical equation is presented which relates the duration of the response to these three factors. On the basis of these and previously published data, the following hypothesis is proposed for the mechanism underlying ciliary reversal in response to cationic stimulation: Ca(++) liberated from the cellular cation exchange system activates a contractile system which is energized by ATP. Contraction of this component results in the reversal of effective beat direction of cilia by a mechanism not yet understood. The duration of reversal in live paramecia is related to the time course of bound calcium release. PMID:4966766

  20. Photolysis of caged calcium in cilia induces ciliary reversal in Paramecium caudatum.

    PubMed

    Iwadate, Yoshiaki

    2003-04-01

    Intracellular Ca(2+) concentration controls both the pattern and frequency of ciliary and flagellar beating in eukaryotes. In Paramecium, it is widely accepted that the reversal of the direction of ciliary beating (ciliary reversal) is induced by an increase in intra-ciliary Ca(2+) levels. Despite this, the Ca(2+)-sensitive region of the cilium that initiates ciliary reversal has not been clearly identified. We injected caged calcium into living P. caudatum cells and applied ultraviolet (UV) light to portions of the injected cells to raise artificially the intracellular Ca(2+) level ([Ca(2+)](i)). UV application to the upper ciliary region above the basal body induced ciliary reversal in injected cells. Furthermore, UV application to the tips of cilia induced weak ciliary reversal. Larger areas of photolysis in the cilium gave rise to greater angles of ciliary reversal. These results strongly suggest that the Ca(2+)-sensitive region for ciliary reversal is distributed all over the cilium, above the basal body. PMID:12604576

  1. [Correction of the position of the cilia in facial paralysis: Technical note].

    PubMed

    Caillot, A; Labbé, D

    2015-06-01

    Facial paralysis is a incapacitating pathology that we treat with lengthening temporalis myoplasty for reanimation of the smile. To treat lagophthalmia, we use the extension of the levator of the upper eyelid according Tessier and the asymmetric external blepharorraphy. These techniques can optionally be combined with other techniques, as needed. However, many patients are embarrassed by the appearance of the lashes of the upper eyelid homolateral side facial paralysis. The cilia are lowered and horizontalised, creating a functional disorder by partial "amputation" of the visual field and aesthetic inconvenience. We describe a surgical technique to correct the malposition of the lashes. This technique can be carried out independently or in the lengthening of the temporal myoplasty or another surgical procedure on the eye. In case of extension of the levator of the upper eyelid, the technique we propose requires no additional incision. This is a simple technique and increases very little surgical time. It is fast, little or no morbid, reproducible and provides a significant improvement in the aesthetic and functional patient. This simple technique allows to provide both aesthetic and functional refinement for patients with facial paralysis sequelae. PMID:25708730

  2. Primary cilia act as mechanosensors during bone healing around an implant

    PubMed Central

    Leucht, P.; Monica, S.D.; Temiyasathit, S.; Lenton, K.; Manu, A.; Longaker, M.T.; Jacobs, C.R.; Spilker, R.L.; Guo, H.; Brunski, J.B.; Helms, J.A.

    2012-01-01

    The primary cilium is an organelle that senses cues in a cell’s local environment. Some of these cues constitute molecular signals; here, we investigate the extent to which primary cilia can also sense mechanical stimuli. We used a conditional approach to delete Kif3a in pre-osteoblasts and then employed a motion device that generated a spatial distribution of strain around an intra-osseous implant positioned in the mouse tibia. We correlated interfacial strain fields with cell behaviors ranging from proliferation through all stages of osteogenic differentiation. We found that peri-implant cells in the Col1Cre;Kif3afl/fl mice were unable to proliferate in response to a mechanical stimulus, failed to deposit and then orient collagen fibers to the strain fields caused by implant displacement, and failed to differentiate into bone-forming osteoblasts. Collectively, these data demonstrate that the lack of a functioning primary cilium blunts the normal response of a cell to a defined mechanical stimulus. The ability to manipulate the genetic background of peri-implant cells within the context of a whole, living tissue provides a rare opportunity to explore mechanotransduction from a multi-scale perspective. PMID:22784673

  3. Self-organization in cytoskeletal mixtures: from synthetic cilia to flowing networks

    NASA Astrophysics Data System (ADS)

    Sanchez, Tim

    2013-03-01

    Inspired by biological functions such as ciliary beating and cytoplasmic streaming, we have developed a highly tunable and robust model system from biological components that self-organizes to produce a broad range of far-from-equilibrium materials with remarkable emergent properties. Using only simple components - microtubules, kinesin motor clusters, and a depletion agent that bundles MTs - we reproduced several essential biological functions, including cilia-like beating, the emergence of metachronal waves in bundle arrays, and internally generated flows in active cytoskeletal gels. The occurrence of these biomimetic functions as self-organized processes provides unique insight into the mechanisms that drive these processes in biology. Beyond these biomimetic behaviors, we have also used the same components to engineer novel active materials which have no biological analogues: active streaming 2D nematics, and finally self-propelled emulsion droplets. These observations exemplify how assemblages of animate microscopic objects exhibit highly sought-after collective and biomimetic properties, challenging us to develop a theoretical framework that would allow for a systematic engineering of their far-from-equilibrium material properties.

  4. PEX6 is Expressed in Photoreceptor Cilia and Mutated in Deafblindness with Enamel Dysplasia and Microcephaly.

    PubMed

    Zaki, Maha S; Heller, Raoul; Thoenes, Michaela; Nürnberg, Gudrun; Stern-Schneider, Gabi; Nürnberg, Peter; Karnati, Srikanth; Swan, Daniel; Fateen, Ekram; Nagel-Wolfrum, Kerstin; Mostafa, Mostafa I; Thiele, Holger; Wolfrum, Uwe; Baumgart-Vogt, Eveline; Bolz, Hanno J

    2016-02-01

    Deafblindness is part of several genetic disorders. We investigated a consanguineous Egyptian family with two siblings affected by congenital hearing loss and retinal degeneration, initially diagnosed as Usher syndrome type 1. At teenage, severe enamel dysplasia, developmental delay, and microcephaly became apparent. Genome-wide homozygosity mapping and whole-exome sequencing detected a homozygous missense mutation, c.1238G>T (p.Gly413Val), affecting a highly conserved residue of peroxisomal biogenesis factor 6, PEX6. Biochemical profiling of the siblings revealed abnormal and borderline plasma phytanic acid concentration, and cerebral imaging revealed white matter disease in both. We show that Pex6 localizes to the apical extensions of secretory ameloblasts and differentiated odontoblasts at early stages of dentin synthesis in mice, and to cilia of retinal photoreceptor cells. We propose PEX6, and possibly other peroxisomal genes, as candidate for the rare cooccurrence of deafblindness and enamel dysplasia. Our study for the first time links peroxisome biogenesis disorders to retinal ciliopathies. PMID:26593283

  5. Dynamic self-assembly of motile bacteria in liquid crystals

    PubMed Central

    Mushenheim, Peter C.; Trivedi, Rishi R.; Tuson, Hannah H.

    2014-01-01

    This paper reports an investigation of dynamical behaviors of motile rod-shaped bacteria within anisotropic viscoelastic environments defined by lyotropic liquid crystals (LCs). In contrast to passive microparticles (including non-motile bacteria) that associate irreversibly in LCs via elasticity-mediated forces, we report that motile Proteus mirabilis bacteria form dynamic and reversible multi-cellular assemblies when dispersed in a lyotropic LC. By measuring the velocity of the bacteria through the LC (8.8 +/− 0.2 μm/s) and by characterizing the ordering of the LC about the rod-shaped bacteria (tangential anchoring), we conclude that the reversibility of the inter-bacterial interaction emerges from the interplay of forces generated by the flagella of the bacteria and the elasticity of the LC, both of which are comparable in magnitude (tens of pN) for motile Proteus mirabilis cells. We also measured the dissociation process, which occurs in a direction determined by the LC, to bias the size distribution of multi-cellular bacterial complexes in a population of motile Proteus mirabilis relative to a population of non-motile cells. Overall, these observations and others reported in this paper provide insight into the fundamental dynamical behaviors of bacteria in complex anisotropic environments and suggest that motile bacteria in LCs are an exciting model system for exploration of principles for the design of active materials. PMID:24652584

  6. Quorum sensing positively regulates flagellar motility in pathogenic Vibrio harveyi.

    PubMed

    Yang, Qian; Defoirdt, Tom

    2015-04-01

    Vibrios belonging to the Harveyi clade are among the major pathogens of aquatic organisms. Quorum sensing (QS) is essential for virulence of V. harveyi towards different hosts. However, most virulence factors reported to be controlled by QS to date are negatively regulated by QS, therefore suggesting that their impact on virulence is limited. In this study, we report that QS positively regulates flagellar motility. We found that autoinducer synthase mutants showed significantly lower swimming motility than the wild type, and the swimming motility could be restored by adding synthetic signal molecules. Further, motility of a luxO mutant with inactive QS (LuxO D47E) was significantly lower than that of the wild type and of a luxO mutant with constitutively maximal QS activity (LuxO D47A). Furthermore, we found that the expression of flagellar genes (both early, middle and late genes) was significantly lower in the luxO mutant with inactive QS when compared with wild type and the luxO mutant with maximal QS activity. Motility assays and gene expression also revealed the involvement of the quorum-sensing master regulator LuxR in the QS regulation of motility. Finally, the motility inhibitor phenamil significantly decreased the virulence of V. harveyi towards gnotobiotic brine shrimp larvae. PMID:24528485

  7. Primary cilium - antenna-like structure on the surface of most mammalian cell types

    NASA Astrophysics Data System (ADS)

    Dvorak, J.; Sitorova, V.; Hadzi Nikolov, D.; Mokry, J.; Richter, I.; Kasaova, L.; Filip, S.; Ryska, A.; Petera, J.

    2011-12-01

    The primary cilium is a sensory solitary non-motile microtubule-based organelle protruding in the quiescent phase of the cell cycle from the surface of the majority of human cells, including embryonic cells, stem cells and stromal cells of malignant tumors. The presence of a primary cilium on the surface of a cell is transient, limited to the quiescent G1(G0) phase and the beginning of the S phase of the cell cycle. The primary cilium is formed from the mother centriole. Primary cilia are key coordinators of signaling pathways during development and tissue homeostasis and, when deffective, they are a major cause of human diseases and developmental disorders, now commonly referred to as ciliopathies. Most cancer cells do not possess a primary cilium. The loss of the primary cilium is a regular feature of neoplastic transformation in the majority of solid tumors. The primary cilium could serve as a tumor suppressor organelle. The aim of this paper was to provide a review of the current knowledge of the primary cilium.

  8. Flagella-independent surface motility in Salmonella enterica serovar Typhimurium

    PubMed Central

    Park, Sun-Yang; Pontes, Mauricio H.; Groisman, Eduardo A.

    2015-01-01

    Flagella are multiprotein complexes necessary for swimming and swarming motility. In Salmonella enterica serovar Typhimurium, flagella-mediated motility is repressed by the PhoP/PhoQ regulatory system. We now report that Salmonella can move on 0.3% agarose media in a flagella-independent manner when experiencing the PhoP/PhoQ-inducing signal low Mg2+. This motility requires the PhoP-activated mgtA, mgtC, and pagM genes, which specify a Mg2+ transporter, an inhibitor of Salmonella’s own F1Fo ATPase, and a small protein of unknown function, respectively. The MgtA and MgtC proteins are necessary for pagM expression because pagM mRNA levels were lower in mgtA and mgtC mutants than in wild-type Salmonella, and also because pagM expression from a heterologous promoter rescued motility in mgtA and mgtC mutants. PagM promotes group motility by a surface protein(s), as a pagM-expressing strain conferred motility upon a pagM null mutant, and proteinase K treatment eliminated motility. The pagM gene is rarely found outside subspecies I of S. enterica and often present in nonfunctional allelic forms in organisms lacking the identified motility. Deletion of the pagM gene reduced bacterial replication on 0.3% agarose low Mg2+ media but not in low Mg2+ liquid media. Our findings define a form of motility that allows Salmonella to scavenge nutrients and to escape toxic compounds in low Mg2+ semisolid environments. PMID:25624475

  9. Flagella-independent surface motility in Salmonella enterica serovar Typhimurium.

    PubMed

    Park, Sun-Yang; Pontes, Mauricio H; Groisman, Eduardo A

    2015-02-10

    Flagella are multiprotein complexes necessary for swimming and swarming motility. In Salmonella enterica serovar Typhimurium, flagella-mediated motility is repressed by the PhoP/PhoQ regulatory system. We now report that Salmonella can move on 0.3% agarose media in a flagella-independent manner when experiencing the PhoP/PhoQ-inducing signal low Mg(2+). This motility requires the PhoP-activated mgtA, mgtC, and pagM genes, which specify a Mg(2+) transporter, an inhibitor of Salmonella's own F1Fo ATPase, and a small protein of unknown function, respectively. The MgtA and MgtC proteins are necessary for pagM expression because pagM mRNA levels were lower in mgtA and mgtC mutants than in wild-type Salmonella, and also because pagM expression from a heterologous promoter rescued motility in mgtA and mgtC mutants. PagM promotes group motility by a surface protein(s), as a pagM-expressing strain conferred motility upon a pagM null mutant, and proteinase K treatment eliminated motility. The pagM gene is rarely found outside subspecies I of S. enterica and often present in nonfunctional allelic forms in organisms lacking the identified motility. Deletion of the pagM gene reduced bacterial replication on 0.3% agarose low Mg(2+) media but not in low Mg(2+) liquid media. Our findings define a form of motility that allows Salmonella to scavenge nutrients and to escape toxic compounds in low Mg(2+) semisolid environments. PMID:25624475

  10. Pattern-formation mechanisms in motility mutants of Myxococcus xanthus

    PubMed Central

    Starruß, Jörn; Peruani, Fernando; Jakovljevic, Vladimir; Søgaard-Andersen, Lotte; Deutsch, Andreas; Bär, Markus

    2012-01-01

    Formation of spatial patterns of cells is a recurring theme in biology and often depends on regulated cell motility. Motility of the rod-shaped cells of the bacterium Myxococcus xanthus depends on two motility machineries, type IV pili (giving rise to S-motility) and the gliding motility apparatus (giving rise to A-motility). Cell motility is regulated by occasional reversals. Moving M. xanthus cells can organize into spreading colonies or spore-filled fruiting bodies, depending on their nutritional status. To ultimately understand these two pattern-formation processes and the contributions by the two motility machineries, as well as the cell reversal machinery, we analyse spatial self-organization in three M. xanthus strains: (i) a mutant that moves unidirectionally without reversing by the A-motility system only, (ii) a unidirectional mutant that is also equipped with the S-motility system, and (iii) the wild-type that, in addition to the two motility systems, occasionally reverses its direction of movement. The mutant moving by means of the A-engine illustrates that collective motion in the form of large moving clusters can arise in gliding bacteria owing to steric interactions of the rod-shaped cells, without the need of invoking any biochemical signal regulation. The two-engine strain mutant reveals that the same phenomenon emerges when both motility systems are present, and as long as cells exhibit unidirectional motion only. From the study of these two strains, we conclude that unidirectional cell motion induces the formation of large moving clusters at low and intermediate densities, while it results in vortex formation at very high densities. These findings are consistent with what is known from self-propelled rod models, which strongly suggests that the combined effect of self-propulsion and volume exclusion interactions is the pattern-formation mechanism leading to the observed phenomena. On the other hand, we learn that when cells occasionally reverse

  11. Motility, Force Generation, and Energy Consumption of Unicellular Parasites.

    PubMed

    Hochstetter, Axel; Pfohl, Thomas

    2016-07-01

    Motility is a key factor for pathogenicity of unicellular parasites, enabling them to infiltrate and evade host cells, and perform several of their life-cycle events. State-of-the-art methods of motility analysis rely on a combination of optical tweezers with high-resolution microscopy and microfluidics. With this technology, propulsion forces, energies, and power generation can be determined so as to shed light on the motion mechanisms, chemotactic behavior, and specific survival strategies of unicellular parasites. With these new tools in hand, we can elucidate the mechanisms of motility and force generation of unicellular parasites, and identify ways to manipulate and eventually inhibit them. PMID:27157805

  12. Effects of trifluoromethyl ketones on the motility of Proteus vulgaris.

    PubMed

    Wolfart, Krisztina; Molnar, Annamaria; Kawase, Masami; Motohashi, Noboru; Molnar, Joseph

    2004-09-01

    In the present study, we showed the inhibition of motility by trifluoromethyl ketone (TF) derivatives (1-8) in Proteus vulgaris (P. vulgaris) cultures. Among them, 1-(2-benzoxazoyl)-3,3,3-trifluoro-2-propanone (1) showed a much stronger inhibitory effect on the motility of P. vulgaris than other TF compounds at 10% MIC. Our results suggest the possibility of an inhibitory action of TF compounds on the proton motive forces by affecting the action of biological motor and proton efflux in the membranes, resulting in a reduction of the ratio of running and the increased number of tumbling and non-motile cells. PMID:15340240

  13. Mechanisms of mammalian iron homeostasis

    PubMed Central

    Pantopoulos, Kostas; Porwal, Suheel Kumar; Tartakoff, Alan; Devireddy, L.

    2012-01-01

    Iron is vital for almost all organisms because of its ability to donate and accept electrons with relative ease. It serves as a cofactor for many proteins and enzymes necessary for oxygen and energy metabolism, as well as for several other essential processes. Mammalian cells utilize multiple mechanisms to acquire iron. Disruption of iron homeostasis is associated with various human diseases: iron deficiency resulting from defects in acquisition or distribution of the metal causes anemia; whereas iron surfeit resulting from excessive iron absorption or defective utilization causes abnormal tissue iron deposition, leading to oxidative damage. Mammals utilize distinct mechanisms to regulate iron homeostasis at the systemic and cellular levels. These involve the hormone hepcidin and iron regulatory proteins, which collectively ensure iron balance. This review outlines recent advances in iron regulatory pathways, as well as in mechanisms underlying intracellular iron trafficking, an important but less-studied area of mammalian iron homeostasis. PMID:22703180

  14. An overview of mammalian pluripotency.

    PubMed

    Wu, Jun; Yamauchi, Takayoshi; Izpisua Belmonte, Juan Carlos

    2016-05-15

    Mammalian pluripotency is the ability to give rise to all somatic cells as well as the germ cells of an adult mammal. It is a unique feature of embryonic epiblast cells, existing only transiently, as cells pass through early developmental stages. By contrast, pluripotency can be captured and stabilized indefinitely in cell culture and can also be reactivated in differentiated cells via nuclear reprogramming. Pluripotent stem cells (PSCs) are the in vitro carriers of pluripotency and they can inhabit discrete pluripotent states depending on the stage at which they were derived and their culture conditions. Here, and in the accompanying poster, we provide a summary of mammalian pluripotency both in vivo and in vitro, and highlight recent and future applications of PSCs for basic and translational research. PMID:27190034

  15. Phosphorylation of a Myosin Motor by TgCDPK3 Facilitates Rapid Initiation of Motility during Toxoplasma gondii egress

    PubMed Central

    Gaji, Rajshekhar Y.; Johnson, Derrick E.; Treeck, Moritz; Wang, Mu; Hudmon, Andy; Arrizabalaga, Gustavo

    2015-01-01

    Members of the family of calcium dependent protein kinases (CDPK’s) are abundant in certain pathogenic parasites and absent in mammalian cells making them strong drug target candidates. In the obligate intracellular parasite Toxoplasma gondii TgCDPK3 is important for calcium dependent egress from the host cell. Nonetheless, the specific substrate through which TgCDPK3 exerts its function during egress remains unknown. To close this knowledge gap we applied the proximity-based protein interaction trap BioID and identified 13 proteins that are either near neighbors or direct interactors of TgCDPK3. Among these was Myosin A (TgMyoA), the unconventional motor protein greatly responsible for driving the gliding motility of this parasite, and whose phosphorylation at serine 21 by an unknown kinase was previously shown to be important for motility and egress. Through a non-biased peptide array approach we determined that TgCDPK3 can specifically phosphorylate serines 21 and 743 of TgMyoA in vitro. Complementation of the TgmyoA null mutant, which exhibits a delay in egress, with TgMyoA in which either S21 or S743 is mutated to alanine failed to rescue the egress defect. Similarly, phosphomimetic mutations in the motor protein overcome the need for TgCDPK3. Moreover, extracellular Tgcdpk3 mutant parasites have motility defects that are complemented by expression of S21+S743 phosphomimetic of TgMyoA. Thus, our studies establish that phosphorylation of TgMyoA by TgCDPK3 is responsible for initiation of motility and parasite egress from the host-cell and provides mechanistic insight into how this unique kinase regulates the lytic cycle of Toxoplasma gondii. PMID:26544049

  16. Olfactory sensitivity in mammalian species.

    PubMed

    Wackermannová, M; Pinc, L; Jebavý, L

    2016-07-18

    Olfaction enables most mammalian species to detect and discriminate vast numbers of chemical structures called odorants and pheromones. The perception of such chemical compounds is mediated via two major olfactory systems, the main olfactory system and the vomeronasal system, as well as minor systems, such as the septal organ and the Grueneberg ganglion. Distinct differences exist not only among species but also among individuals in terms of their olfactory sensitivity; however, little is known about the mechanisms that determine these differences. In research on the olfactory sensitivity of mammals, scientists thus depend in most cases on behavioral testing. In this article, we reviewed scientific studies performed on various mammalian species using different methodologies and target chemical substances. Human and non-human primates as well as rodents and dogs are the most frequently studied species. Olfactory threshold studies on other species do not exist with the exception of domestic pigs. Olfactory testing performed on seals, elephants, and bats focused more on discriminative abilities than on sensitivity. An overview of olfactory sensitivity studies as well as olfactory detection ability in most studied mammalian species is presented here, focusing on comparable olfactory detection thresholds. The basics of olfactory perception and olfactory sensitivity factors are also described. PMID:27070753

  17. Eye Motility Alterations in Retinitis Pigmentosa

    PubMed Central

    Galeoto, Giovanni; Fratipietro, Manuela

    2015-01-01

    Purpose. We evaluated a sample of individuals with retinitis pigmentosa (RP) with the aim of assessing the presence or absence of ocular motility (OM) disorders. Materials and Methods. We included 23 out of the 25 individuals from the sample (9 females and 14 males) with an average visual acuity of 6/10. Results. The cover test about the vertical deviation in near distance showed an r/l in 3.45% and an l/r in 6.9%. The assessment of OM showed that 39.1% of the sample had a severe hyperfunction of the IO of the right eye and a severe hyperfunction (34.5%) of the SO of the left eye; 21.8% had a moderate hypofunction of right SO with a moderate percentage of hypofunction of 17.5% for the SO of the left eye; 30.5%, however, showed a serious hypofunction of the SR of both eyes; 21.7% of the sample showed a hyperfunction in both eyes of the IR. Conclusion. This alteration, however, is not attributable to either a high refractive defect (medium-low myopia: −1 diopter ±3 SD) or to a severely impaired binocular vision (visual acuity, motor fusion, and stereopsis are normal or within a range of values commonly accepted). Therefore, the disorders of OM lead to a genetic origin. PMID:26124957

  18. Colloidal motility and patterning by physical chemotaxis

    NASA Astrophysics Data System (ADS)

    Palacci, Jeremie; Abecassis, Benjamin; Cottin-Bizonne, Cecile; Ybert, Christophe; Bocquet, Lyderic

    2009-11-01

    We developped a microfluidic setup to show the motility of colloids or biomolecules under a controlled salt gradient thanks to the diffusiophoresis phenomenon [1,2]. We can therefore mimic chemotaxis on simple physical basis with thrilling analogies with the biological chemotaxis of E. Coli bacteria: salt dependance of the velocity [3] and log-sensing behavior [4]. In addition with a temporally tunable gradient we show we can generate an effective osmotic potential to trap colloids or DNA. These experimental observations are supported by numerical simulations and an asymptotic ratchet model. Finally, we use these traps to generate various patterns and because concentration gradients are ubiquitous in nature, we question for the role of such a mecanism in morphogenesis [5] or positioning perspectives in cells [6]. [4pt] [1] B. Abecassis, C. Cottin-Bizonne, C. Ybert, A. Ajdari, and L. Bocquet, Nat. Mat., 7(10):785--789, 2008. [2] Anderson, Ann. Rev. Fluid Mech, 21, 1989. [3] Y. L. Qi and J. Adler, PNAS, 86(21):8358--8362, 1989. [4] Y. V. Kalinin, L. L. Jiang, Y. H. Tu, and M. M. Wu, Biophys. J., 96(6):2439--2448, 2009. [4] J. B. Moseley, A. Mayeux, A. Paoletti, and P. Nurse, Nat., 459(7248):857--U8, 2009. [6] L. Wolpert, Dev., 107:3--12, 1989

  19. Ciliopathy-associated gene Cc2d2a promotes assembly of subdistal appendages on the mother centriole during cilia biogenesis

    PubMed Central

    Veleri, Shobi; Manjunath, Souparnika H.; Fariss, Robert N.; May-Simera, Helen; Brooks, Matthew; Foskett, Trevor A.; Gao, Chun; Longo, Teresa A.; Liu, Pinghu; Nagashima, Kunio; Rachel, Rivka A.; Li, Tiansen; Dong, Lijin; Swaroop, Anand

    2014-01-01

    The primary cilium originates from the mother centriole and participates in critical functions during organogenesis. Defects in cilia biogenesis or function lead to pleiotropic phenotypes. Mutations in centrosome-cilia gene CC2D2A result in Meckel and Joubert syndromes. Here we generate a Cc2d2a-/- mouse that recapitulates features of Meckel syndrome including embryonic lethality and multi-organ defects. Cilia are absent in Cc2d2a-/- embryonic node and other somatic tissues; disruption of cilia-dependent Shh signaling appears to underlie exencephaly in mutant embryos. The Cc2d2a-/- mouse embryonic fibroblasts (MEFs) lack cilia though mother centriole and pericentriolar proteins are detected. Odf2, associated with subdistal appendages, is absent and ninein is reduced in mutant MEFs. In Cc2d2a-/- MEFs, subdistal appendages are lacking or abnormal by transmission-EM. Consistent with this, CC2D2A localizes to subdistal appendages by immuno-EM in wild type cells. We conclude that CC2D2A is essential for the assembly of subdistal appendages, which anchor cytoplasmic microtubules and prime the mother centriole for axoneme biogenesis. PMID:24947469

  20. Ghrelin as a target for gastrointestinal motility disorders.

    PubMed

    Greenwood-Van Meerveld, Beverley; Kriegsman, Michael; Nelson, Richard

    2011-11-01

    The therapeutic potential of ghrelin and synthetic ghrelin receptor (GRLN-R) agonists for the treatment of gastrointestinal (GI) motility disorders is based on their ability to stimulate coordinated patterns of propulsive GI motility. This review focuses on the latest findings that support the therapeutic potential of GRLN-R agonists for the treatment of GI motility disorders. The review highlights the preclinical and clinical prokinetic effects of ghrelin and a series of novel ghrelin mimetics to exert prokinetic effects on the GI tract. We build upon a series of excellent reviews to critically discuss the evidence that supports the potential of GRLN-R agonists to normalize GI motility in patients with GI hypomotility disorders such as gastroparesis, post-operative ileus (POI), idiopathic chronic constipation and functional bowel disorders. PMID:21453735

  1. Mucin Promotes Rapid Surface Motility in Pseudomonas aeruginosa

    PubMed Central

    Yeung, Amy T. Y.; Parayno, Alicia; Hancock, Robert E. W.

    2012-01-01

    ABSTRACT An important environmental factor that determines the mode of motility adopted by Pseudomonas aeruginosa is the viscosity of the medium, often provided by adjusting agar concentrations in vitro. However, the viscous gel-like property of the mucus layer that overlays epithelial surfaces is largely due to the glycoprotein mucin. P. aeruginosa is known to swim within 0.3% (wt/vol) agar and swarm on the surface at 0.5% (wt/vol) agar with amino acids as a weak nitrogen source. When physiological concentrations or as little as 0.05% (wt/vol) mucin was added to the swimming agar, in addition to swimming, P. aeruginosa was observed to undergo highly accelerated motility on the surface of the agar. The surface motility colonies in the presence of mucin appeared to be circular, with a bright green center surrounded by a thicker white edge. While intact flagella were required for the surface motility in the presence of mucin, type IV pili and rhamnolipid production were not. Replacement of mucin with other wetting agents indicated that the lubricant properties of mucin might contribute to the surface motility. Based on studies with mutants, the quorum-sensing systems (las and rhl) and the orphan autoinducer receptor QscR played important roles in this form of surface motility. Transcriptional analysis of cells taken from the motility zone revealed the upregulation of genes involved in virulence and resistance. Based on these results, we suggest that mucin may be promoting a new or highly modified form of surface motility, which we propose should be termed “surfing.” PMID:22550036

  2. Dexmedetomidine and Fentanyl Exhibit Temperature Dependent Effects on Human Respiratory Cilia

    PubMed Central

    Welchering, Nils; Ochoa, Sebastian; Tian, Xin; Francis, Richard; Zahid, Maliha; Muñoz, Ricardo; Lo, Cecilia W.

    2015-01-01

    Background: Dexmedetomidine (dex) is commonly used in intensive care due to its effective sedation and analgesia with few adverse effects and minimal respiratory depression. However, we recently observed that exposing mouse epithelial respiratory cells to dex decreased ciliary beat frequency (CBF), suggesting dex may pose pulmonary risk. Objective: The purpose of this study is to determine the effects of dex at clinically relevant doses on CBF in human respiratory epithelia. Methods: Human nasal epithelial cilia were obtained from the inferior nasal turbinate with a rhinoprobe and placed in culture medium at 15°C and 37°C. At 5 and 30 min, video-microscopy was used to assess CBF, either without (control) or with different concentrations (1, 5, and 10 nM) of dex, fentanyl (fen), and dex + fen combination. Results: At 15°C, CBF was lower in the dex group compared to controls at 5 and 30 min. At 37°C, there was a significant increase in CBF with dex at 5 and 30 min, except for dex at 5 nM after 5 min, which showed a significant decrease. At 15°C the combination of dex + fen showed a positive interaction, causing less ciliary inhibition as expected. In contrast, no interaction between drugs was seen between dex and fen at 37°C. Conclusion: At low temperatures, dex reduces CBF in human respiratory epithelia, whereas dex increases CBF at physiologic temperature in vitro. Whether these effects translate into clinical consequences during hypothermia, as with cardiopulmonary bypass surgery will require further studies. PMID:25717467

  3. Mucociliary and long-term particle clearance in airways of patients with immotile cilia

    PubMed Central

    Möller, Winfried; Häußinger, Karl; Ziegler-Heitbrock, Löms; Heyder, Joachim

    2006-01-01

    Spherical monodisperse ferromagnetic iron oxide particles of 1.9 μm geometric and 4.2 μm aerodynamic diameter were inhaled by seven patients with primary ciliary dyskinesia (PCD) using the shallow bolus technique, and compared to 13 healthy non-smokers (NS) from a previous study. The bolus penetration front depth was limiting to the phase1 dead space volume. In PCD patients deposition was 58+/-8 % after 8 s breath holding time. Particle retention was measured by the magnetopneumographic method over a period of nine months. Particle clearance from the airways showed a fast and a slow phase. In PCD patients airway clearance was retarded and prolonged, 42+/-12 % followed the fast phase with a mean half time of 16.8+/-8.6 hours. The remaining fraction was cleared slowly with a half time of 121+/-25 days. In healthy NS 49+/-9 % of particles were cleared in the fast phase with a mean half time of 3.0+/-1.6 hours, characteristic of an intact mucociliary clearance. There was no difference in the slow clearance phase between PCD patients and healthy NS. Despite non-functioning cilia the effectiveness of airway clearance in PCD patients is comparable to healthy NS, with a prolonged kinetics of one week, which may primarily reflect the effectiveness of cough clearance. This prolonged airway clearance allows longer residence times of bacteria and viruses in the airways and may be one reason for increased frequency of infections in PCD patients. PMID:16423294

  4. Microfabricated ratchet structures for concentrating and patterning motile bacterial cells

    NASA Astrophysics Data System (ADS)

    Yub Kim, Sang; Lee, Eun Se; Lee, Ho Jae; Lee, Se Yeon; Kuk Lee, Sung; Kim, Taesung

    2010-09-01

    We present a novel microfabricated concentrator for Escherichia coli that can be a stand-alone and self-contained microfluidic device because it utilizes the motility of cells. First of all, we characterize the motility of E. coli cells and various ratcheting structures that can guide cells to move in a desired direction in straight and circular channels. Then, we combine these ratcheting microstructures with the intrinsic tendency of cells to swim on the right side in microchannels to enhance the concentration rates up to 180 fold until the concentrators are fully filled with cells. Furthermore, we demonstrate that cells can be positioned and concentrated with a constant spacing distance on a surface, allowing spatial patterning of motile cells. These results can be applied to biosorption or biosensor devices that are powered by motile cells because they can be highly concentrated without any external mechanical and electrical energy sources. Hence, we believe that the concentrator design holds considerable potential to be applied for concentrating and patterning other motile microbes and providing a versatile structure for motility study of bacterial cells.

  5. Upper gastrointestinal motility: prenatal development and problems in infancy.

    PubMed

    Singendonk, Maartje M J; Rommel, Nathalie; Omari, Taher I; Benninga, Marc A; van Wijk, Michiel P

    2014-09-01

    Deglutition, or swallowing, refers to the process of propulsion of a food bolus from the mouth into the stomach and involves the highly coordinated interplay of swallowing and breathing. At 34 weeks gestational age most neonates are capable of successful oral feeding if born at this time; however, the maturation of respiration is still in progress at this stage. Infants can experience congenital and developmental pharyngeal and/or gastrointestinal motility disorders, which might manifest clinically as gastro-oesophageal reflux (GER) symptoms, feeding difficulties and/or refusal, choking episodes and airway changes secondary to micro or overt aspiration. These problems might lead to impaired nutritional intake and failure to thrive. These gastrointestinal motility disorders are mostly classified according to the phase of swallowing in which they occur, that is, the oral preparatory, oral, pharyngeal and oesophageal phases. GER is a common phenomenon in infancy and is referred to as GERD when it causes troublesome complications. GER is predominantly caused by transient relaxation of the lower oesophageal sphincter. In oesophageal atresia, oesophageal motility disorders develop in almost all patients after surgery; however, a congenital origin of disordered motility has also been proposed. This Review highlights the prenatal development of upper gastrointestinal motility and describes the most common motility disorders that occur in early infancy. PMID:24890279

  6. Comparison of motility stimulants for cryopreserved human semen.

    PubMed

    Hammitt, D G; Bedia, E; Rogers, P R; Syrop, C H; Donovan, J F; Williamson, R A

    1989-09-01

    Caffeine, pentoxifylline, 2-deoxyadenosine, cyclic adenosine monophosphate (cAMP), relaxin, adenosine, kallikrein, and calcium were compared for their ability to stimulate motility of cryopreserved sperm. Caffeine, pentoxifylline, and 2-deoxyadenosine significantly increased the percentage of motile sperm at 15, 30, 45, and 60 minutes after administration. Sperm velocity was significantly increased by caffeine at 0, 15, 30, and 45 minutes, and by pentoxifylline at 0, 45, and 60 minutes. Consistent stimulation was not observed for other chemicals. Caffeine, pentoxifylline, and 2-deoxyadenosine were then examined for their ability to provide motility stimulation after removal with washing. With the exception of caffeine, percent motility and velocity for stimulated and untreated sperm were similar after washing. A significant reduction in motility was observed at 48 hours after washing for caffeine. The percentage of hamster oocytes penetrated at 24 hours after washing was significantly reduced for caffeine, 2-deoxyadenosine, and pentoxifylline combined with 2-deoxyadenosine. Pentoxifylline-treated sperm showed no reduction in fertilizing capacity. These results indicate that, of the chemicals examined, pentoxifylline is superior for motility stimulation of cryopreserved sperm. PMID:2550282

  7. Mass sperm motility is associated with fertility in sheep.

    PubMed

    David, Ingrid; Kohnke, Philippa; Lagriffoul, Gilles; Praud, Olivier; Plouarboué, Franck; Degond, Pierre; Druart, Xavier

    2015-10-01

    The study was to focus on the relationship between wave motion (mass sperm motility, measured by a mass sperm motility score, manually assessed by artificial insemination (AI) center operators) and fertility in male sheep. A dataset of 711,562 artificial inseminations performed in seven breeds by five French AI centers during the 2001-2005 time period was used for the analysis. Factors influencing the outcome of the insemination, which is a binary response observed at lambing of either success (1) or failure (0), were studied using a joint model within each breed and AI center (eight separate analyses). The joint model is a multivariate model where all information related to the female, the male and the insemination process were included to improve the estimation of the factor effects. Results were consistent for all analyses. The male factors affecting AI results were the age of the ram and the mass motility. After correction for the other factors of variation, the lambing rate increased quasi linearly from three to more than ten points with the mass sperm motility score depending on the breed and the AI center. The consistency of the relationship for all breeds indicated that mass sperm motility is predictive of the fertility resulting when sperm are used from a specific ejaculate. Nonetheless, predictability could be improved if an objective measurement of mass sperm motility were available as a substitute for the subjective scoring currently in use in AI centers. PMID:26364125

  8. On the motility of military microrobots

    SciTech Connect

    Solem, J.C.

    1991-07-01

    I show that at the physical limits of technology, crude robots on the size scale of 10--100 {mu}m may be possible. An interesting aspect of such miniscule vehicles is the means by which they might move about. I address this question with a number of detailed calculations for microrobots traveling by air, land, and sea. The Reynolds number for airborne robots is close to unity -- the viscous forces dominate the inertial forces. I show that there is no sense to using a lifting airfoil, a microrobotic helicopter could fly by simply gripping the viscous air around it. Swimming robots encounter a higher Reynolds number and I explore a variety of propulsion mechanisms. The best propulsion appears to be a fan propeller using blades of rather unusual design. Surprisingly, the corkscrew-flagellum propulsion of the motile form of Escherichia coli is a good deal less efficient than this fan propulsion. Nature is known for her parsimonious use of energy: perhaps she uses the flagellum because it is easy to fabricate from protein. Hopping seems to be the most effective mode of transport for earth-bound robots. It is stealthy, predator-evading, and energy-efficient and provides mobility over many types of terrain. I calculate optimum hopping strategies as a function of weight and atmospheric viscosity. It would be interesting to see how these equations apply to insects. Finally, I show various ways adhesion and electric fields can be used for walking on walls. The research is avant-garde, but may be useful when micromechanical technology reaches the projected level of competence. 5 figs.

  9. Shear stress blunts tubuloglomerular feedback partially mediated by primary cilia and nitric oxide at the macula densa.

    PubMed

    Wang, Lei; Shen, Chunyu; Liu, Haifeng; Wang, Shaohui; Chen, Xinshan; Roman, Richard J; Juncos, Luis A; Lu, Yan; Wei, Jin; Zhang, Jie; Yip, Kay-Pong; Liu, Ruisheng

    2015-10-01

    The present study tested whether primary cilia on macula densa serve as a flow sensor to enhance nitric oxide synthase 1 (NOS1) activity and inhibit tubuloglomerular feedback (TGF). Isolated perfused macula densa was loaded with calcein red and 4,5-diaminofluorescein diacetate to monitor cell volume and nitric oxide (NO) generation. An increase in tubular flow rate from 0 to 40 nl/min enhanced NO production by 40.0 ± 1.2%. The flow-induced NO generation was blocked by an inhibitor of NOS1 but not by inhibition of the Na/K/2Cl cotransporter or the removal of electrolytes from the perfusate. NO generation increased from 174.8 ± 21 to 276.1 ± 24 units/min in cultured MMDD1 cells when shear stress was increased from 0.5 to 5.0 dynes/cm(2). The shear stress-induced NO generation was abolished in MMDD1 cells in which the cilia were disrupted using a siRNA to ift88. Increasing the NaCl concentration of the tubular perfusate from 10 to 80 mM NaCl in the isolated perfused juxtaglomerular preparation reduced the diameter of the afferent arteriole by 3.8 ± 0.1 μm. This response was significantly blunted to 2.5 ± 0.2 μm when dextran was added to the perfusate to increase the viscosity and shear stress. Inhibition of NOS1 blocked the effect of dextran on TGF response. In vitro, the effects of raising perfusate viscosity with dextran on tubular hydraulic pressure were minimized by reducing the outflow resistance to avoid stretching of tubular cells. These results suggest that shear stress stimulates primary cilia on the macula densa to enhance NO generation and inhibit TGF responsiveness. PMID:26269519

  10. Whole exome re-sequencing implicates CCDC38 and cilia structure and function in resistance to smoking related airflow obstruction.

    PubMed

    Wain, Louise V; Sayers, Ian; Soler Artigas, María; Portelli, Michael A; Zeggini, Eleftheria; Obeidat, Ma'en; Sin, Don D; Bossé, Yohan; Nickle, David; Brandsma, Corry-Anke; Malarstig, Anders; Vangjeli, Ciara; Jelinsky, Scott A; John, Sally; Kilty, Iain; McKeever, Tricia; Shrine, Nick R G; Cook, James P; Patel, Shrina; Spector, Tim D; Hollox, Edward J; Hall, Ian P; Tobin, Martin D

    2014-05-01

    Chronic obstructive pulmonary disease (COPD) is a leading cause of global morbidity and mortality and, whilst smoking remains the single most important risk factor, COPD risk is heritable. Of 26 independent genomic regions showing association with lung function in genome-wide association studies, eleven have been reported to show association with airflow obstruction. Although the main risk factor for COPD is smoking, some individuals are observed to have a high forced expired volume in 1 second (FEV1) despite many years of heavy smoking. We hypothesised that these "resistant smokers" may harbour variants which protect against lung function decline caused by smoking and provide insight into the genetic determinants of lung health. We undertook whole exome re-sequencing of 100 heavy smokers who had healthy lung function given their age, sex, height and smoking history and applied three complementary approaches to explore the genetic architecture of smoking resistance. Firstly, we identified novel functional variants in the "resistant smokers" and looked for enrichment of these novel variants within biological pathways. Secondly, we undertook association testing of all exonic variants individually with two independent control sets. Thirdly, we undertook gene-based association testing of all exonic variants. Our strongest signal of association with smoking resistance for a non-synonymous SNP was for rs10859974 (P = 2.34 × 10(-4)) in CCDC38, a gene which has previously been reported to show association with FEV1/FVC, and we demonstrate moderate expression of CCDC38 in bronchial epithelial cells. We identified an enrichment of novel putatively functional variants in genes related to cilia structure and function in resistant smokers. Ciliary function abnormalities are known to be associated with both smoking and reduced mucociliary clearance in patients with COPD. We suggest that genetic influences on the development or function of cilia in the bronchial epithelium may

  11. Inv acts as a molecular anchor for Nphp3 and Nek8 in the proximal segment of primary cilia.

    PubMed

    Shiba, Dai; Manning, Danielle K; Koga, Hisashi; Beier, David R; Yokoyama, Takahiko

    2010-02-01

    A primary cilium is an antenna-like structure extending from the surface of most vertebrate cells. It is structurally divided along its vertical axis into sub-compartments that include the ciliary tip, the shaft, the ciliary necklace segment, the transitional zone and the basal body. We recently discovered that the shaft of the primary cilia has a distinct molecular compartment, termed the "Inv compartment", which is characterized by the accumulation of Inv at the base of primary cilia. Inv was discovered as a causative gene in inv mutant mice. It was later found to be responsible for the infantile type of nephronophthisis (NPHP2). Nephronophthisis (NPHP) is an autosomal recessive kidney disease. Nine causative genes have been identified, with all examined products thought to function in cilia, basal body and/or centrioles. However, their exact intra-ciliary localization and relationship have not been clear. Here, we report that products of Nphp3 and Nek8 (the mouse orthologs of the causative genes for NPHP3 and NPHP9, respectively) localize to the Inv compartment. We also show that Inv is essential for the compartmental localization of Nphp3 and Nek8, whereas localization of Inv does not require Nphp3 or Nek8. Nphp1 and Nphp4 also localize at the proximal region of the cilium, but not in Inv compartment. Our results indicate that Inv acts as an anchor for Nphp3 and Nek8 in the Inv compartment, and suggest that Inv compartment is a candidate site for intra-ciliary interaction of Inv, Nphp3 and Nek8. PMID:20169535

  12. Mammalian Sperm Fertility Related Proteins

    PubMed Central

    Ashrafzadeh, Ali; Karsani, Saiful Anuar; Nathan, Sheila

    2013-01-01

    Infertility is an important aspect of human and animal reproduction and still presents with much etiological ambiguity. As fifty percent of infertility is related to the male partner, molecular investigations on sperm and seminal plasma can lead to new knowledge on male infertility. Several comparisons between fertile and infertile human and other species sperm proteome have shown the existence of potential fertility markers. These proteins have been categorized into energy related, structural and other functional proteins which play a major role in sperm motility, capacitation and sperm-oocyte binding. The data from these studies show the impact of sperm proteome studies on identifying different valuable markers for fertility screening. In this article, we review recent development in unraveling sperm fertility related proteins. PMID:24151436

  13. Reduction of meckelin leads to general loss of cilia, ciliary microtubule misalignment and distorted cell surface organization

    PubMed Central

    2014-01-01

    Background Meckelin (MKS3), a conserved protein linked to Meckel Syndrome, assists in the migration of centrioles to the cell surface for ciliogenesis. We explored for additional functions of MKS3p using RNA interference (RNAi) and expression of FLAG epitope tagged protein in the ciliated protozoan Paramecium tetraurelia. This cell has a highly organized cell surface with thousands of cilia and basal bodies that are grouped into one or two basal body units delineated by ridges. The highly systematized nature of the P. tetraurelia cell surface provides a research model of MKS and other ciliopathies where changes in ciliary structure, subcellular organization and overall arrangement of the cell surface can be easily observed. We used cells reduced in IFT88 for comparison, as the involvement of this gene’s product with cilia maintenance and growth is well understood. Results FLAG-MKS3p was found above the plane of the distal basal body in the transition zone. Approximately 95% of those basal bodies observed had staining for FLAG-MKS3. The RNAi phenotype for MKS3 depleted cells included global shortening and loss of cilia. Basal body structure appeared unaffected. On the dorsal surface, the basal bodies and their associated rootlets appeared rotated out of alignment from the normal anterior-posterior rows. Likewise, cortical units were abnormal in shape and out of alignment from normal rows. A GST pull down using the MKS3 coiled-coil domain suggests previously unidentified interacting partners. Conclusions Reduction of MKS3p shows that this protein affects development and maintenance of cilia over the entire cell surface. Reduction of MKS3p is most visible on the dorsal surface. The anterior basal body is attached to and moves along the striated rootlet of the posterior basal body in preparation for duplication. We propose that with reduced MKS3p, this attachment and guidance of the basal body is lost. The basal body veers off course, causing basal body rows to be

  14. Cyclic GMP Balance Is Critical for Malaria Parasite Transmission from the Mosquito to the Mammalian Host

    PubMed Central

    Lakshmanan, Viswanathan; Fishbaugher, Matthew E.; Morrison, Bob; Baldwin, Michael; Macarulay, Michael; Vaughan, Ashley M.; Mikolajczak, Sebastian A.

    2015-01-01

    ABSTRACT Transmission of malaria occurs during Anopheles mosquito vector blood meals, when Plasmodium sporozoites that have invaded the mosquito salivary glands are delivered to the mammalian host. Sporozoites display a unique form of motility that is essential for their movement across cellular host barriers and invasion of hepatocytes. While the molecular machinery powering motility and invasion is increasingly well defined, the signaling events that control these essential parasite activities have not been clearly delineated. Here, we identify a phosphodiesterase (PDEγ) in Plasmodium, a regulator of signaling through cyclic nucleotide second messengers. Reverse transcriptase PCR (RT-PCR) analysis and epitope tagging of endogenous PDEγ detected its expression in blood stages and sporozoites of Plasmodium yoelii. Deletion of PDEγ (pdeγ−) rendered sporozoites nonmotile, and they failed to invade the mosquito salivary glands. Consequently, PDEγ deletion completely blocked parasite transmission by mosquito bite. Strikingly, pdeγ− sporozoites showed dramatically elevated levels of cyclic GMP (cGMP), indicating that a perturbation in cyclic nucleotide balance is involved in the observed phenotypic defects. Transcriptome sequencing (RNA-Seq) analysis of pdeγ− sporozoites revealed reduced transcript abundance of genes that encode key components of the motility and invasion apparatus. Our data reveal a crucial role for PDEγ in maintaining the cyclic nucleotide balance in the malaria parasite sporozoite stage, which in turn is essential for parasite transmission from mosquito to mammal. PMID:25784701

  15. Evolutionary paths to mammalian cochleae.

    PubMed

    Manley, Geoffrey A

    2012-12-01

    Evolution of the cochlea and high-frequency hearing (>20 kHz; ultrasonic to humans) in mammals has been a subject of research for many years. Recent advances in paleontological techniques, especially the use of micro-CT scans, now provide important new insights that are here reviewed. True mammals arose more than 200 million years (Ma) ago. Of these, three lineages survived into recent geological times. These animals uniquely developed three middle ear ossicles, but these ossicles were not initially freely suspended as in modern mammals. The earliest mammalian cochleae were only about 2 mm long and contained a lagena macula. In the multituberculate and monotreme mammalian lineages, the cochlea remained relatively short and did not coil, even in modern representatives. In the lineage leading to modern therians (placental and marsupial mammals), cochlear coiling did develop, but only after a period of at least 60 Ma. Even Late Jurassic mammals show only a 270 ° cochlear coil and a cochlear canal length of merely 3 mm. Comparisons of modern organisms, mammalian ancestors, and the state of the middle ear strongly suggest that high-frequency hearing (>20 kHz) was not realized until the early Cretaceous (~125 Ma). At that time, therian mam