Beating Of Cilia Research Articles

The Motility of Axonemal Dynein is Regulated by the Tubulin Code

Microtubule diversity, arising from the utilization of various isoforms and posttranslational modifications of tubulin, affects many cellular processes, including the assembly and motility cilia and flagella. However, it is not known whether microtubule diversity directly regulates the activity of the axonemal dyneins, the motors that drive the beating of cilia. To test this possibility, we asked whether in vitro acetylation or deacetylation of α-tubulin's lysine 40 (K40), which is an important posttranslational modification of tubulin within the lumen of microtubules, can influence the activity of outer arm axonemal dynein in motility assays using purified proteins. Additionally, we asked whether proteolytic cleavage of α- and- β-tubulin's C-terminal tails (CTT), which are unstructured regions on the outside of the microtubule lattice and are the location of the detyrosination, polyglutamylation, and polyglycylation modifications as well as most of the genetic diversity between tubulins, regulate dynein's motility. By quantifying the motility with displacement-weighted velocity analysis, we found that both K40 acetylation CTT cleavage increase the motility of axonemal dynein. Our mathematical modeling further suggests that the increase in motility is due to an increase in the power stroke speed of dynein in both cases. These results show that axonemal dynein directly deciphers the tubulin code; specifically luminal acetylation of lysine 40 on α-tubulin increases the motility of dynein on the exterior of the microtubule, and CTTs, and their associated post-translational modifications, tend to decrease motility of dynein. This work has implications on allostery between the microtubule lumen and the external surface, the effect of post-translational modifications on molecular motors and microtubule associated proteins, and the regulation of the eukaryotic ciliary beat.

Ion channels and calcium signaling in motile cilia.

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.

Ciliobrevins as tools for studying dynein motor function.

Dyneins are a small class of molecular motors that bind to microtubules and walk toward their minus ends. They are essential for the transport and distribution of organelles, signaling complexes and cytoskeletal elements. In addition dyneins generate forces on microtubule arrays that power the beating of cilia and flagella, cell division, migration and growth cone motility. Classical approaches to the study of dynein function in axons involve the depletion of dynein, expression of mutant/truncated forms of the motor, or interference with accessory subunits. By necessity, these approaches require prolonged time periods for the expression or manipulation of cellular dynein levels. With the discovery of the ciliobrevins, a class of cell permeable small molecule inhibitors of dynein, it is now possible to acutely disrupt dynein both globally and locally. In this review, we briefly summarize recent work using ciliobrevins to inhibit dynein and discuss the insights ciliobrevins have provided about dynein function in various cell types with a focus on neurons. We temper this with a discussion of the need for studies that will elucidate the mechanism of action of ciliobrevin and as well as the need for experiments to further analyze the specificity of ciliobreviens for dynein. Although much remains to be learned about ciliobrevins, these small molecules are proving themselves to be valuable novel tools to assess the cellular functions of dynein.

Cilium height difference between strokes is more effective in driving fluid transport in mucociliary clearance: A numerical study.

Mucociliary clearance is the first line of defense in our airway. The purpose of this study is to identify and study key factors in the cilia motion that influence the transport ability of the mucociliary system. Using a rod-propel-fluid model, we examine the effects of cilia density, beating frequency, metachronal wavelength, and the extending height of the beating cilia. We first verify that asymmetry in the cilia motion is key to developing transport in the mucus flow. Next, two types of asymmetries between the effective and recovery strokes of the cilia motion are considered, the cilium beating velocity difference and the cilium height difference. We show that the cilium height difference is more efficient in driving the transport, and the more bend the cilium during the recovery stroke is, the more effective the transport would be. It is found that the transport capacity of the mucociliary system increases with cilia density and cilia beating frequency, but saturates above by a threshold value in both density and frequency. The metachronal wave that results from the phase lag among cilia does not contribute much to the mucus transport, which is consistent with the experimental observation of Sleigh (1989). We also test the effect of mucus viscosity, whose value is found to be inversely proportional to the transport ability. While multiple parts have to interplay and coordinate to allow for most effective mucociliary clearance, our findings from a simple model move us closer to understanding the effects of the cilia motion on the efficiency of this clearance system.

Analysis of unstable modes distinguishes mathematical models of flagellar motion

The mechanisms underlying the coordinated beating of cilia and flagella remain incompletely understood despite the fundamental importance of these organelles. The axoneme (the cytoskeletal structure of cilia and flagella) consists of microtubule doublets connected by passive and active elements. The motor protein dynein is known to drive active bending, but dynein activity must be regulated to generate oscillatory, propulsive waveforms. Mathematical models of flagellar motion generate quantitative predictions that can be analysed to test hypotheses concerning dynein regulation. One approach has been to seek periodic solutions to the linearized equations of motion. However, models may simultaneously exhibit both periodic and unstable modes. Here, we investigate the emergence and coexistence of unstable and periodic modes in three mathematical models of flagellar motion, each based on a different dynein regulation hypothesis: (i) sliding control; (ii) curvature control and (iii) control by interdoublet separation (the 'geometric clutch' (GC)). The unstable modes predicted by each model are used to critically evaluate the underlying hypothesis. In particular, models of flagella with 'sliding-controlled' dynein activity admit unstable modes with non-propulsive, retrograde (tip-to-base) propagation, sometimes at the same parameter values that lead to periodic, propulsive modes. In the presence of these retrograde unstable modes, stable or periodic modes have little influence. In contrast, unstable modes of the GC model exhibit switching at the base and propulsive base-to-tip propagation.

Physics of microswimmers—single particle motion and collective behavior: a review

Locomotion and transport of microorganisms in fluids is an essential aspect of life. Search for food, orientation toward light, spreading of off-spring, and the formation of colonies are only possible due to locomotion. Swimming at the microscale occurs at low Reynolds numbers, where fluid friction and viscosity dominates over inertia. Here, evolution achieved propulsion mechanisms, which overcome and even exploit drag. Prominent propulsion mechanisms are rotating helical flagella, exploited by many bacteria, and snake-like or whip-like motion of eukaryotic flagella, utilized by sperm and algae. For artificial microswimmers, alternative concepts to convert chemical energy or heat into directed motion can be employed, which are potentially more efficient. The dynamics of microswimmers comprises many facets, which are all required to achieve locomotion. In this article, we review the physics of locomotion of biological and synthetic microswimmers, and the collective behavior of their assemblies. Starting from individual microswimmers, we describe the various propulsion mechanism of biological and synthetic systems and address the hydrodynamic aspects of swimming. This comprises synchronization and the concerted beating of flagella and cilia. In addition, the swimming behavior next to surfaces is examined. Finally, collective and cooperate phenomena of various types of isotropic and anisotropic swimmers with and without hydrodynamic interactions are discussed.

Modulation of pumping rate by two species of marine bivalve molluscs in response to neurotransmitters: Comparison of in vitro and in vivo results

Most studies regarding the neuroanatomy and neurophysiology of molluscan ctenidia have focused on isolated ctenidial tissue preparations. This study investigated how bivalve molluscs modulate their feeding rates by examining the effects of a variety of neurotransmitters, including serotonin, dopamine, and the dopamine agonist apomorphine on both isolated ctenidial tissue and in intact members of two commercially important bivalve species: the blue mussel, Mytilus edulis; and the bay scallop Argopecten irradians. In particular, we examined the effect of changes in: 1) beat of the lateral cilia (in vitro), 2) distance between ctenidial filaments and/or plicae (in vivo), and 3) diameter of the siphonal openings (in vivo) on alteration of bulk water flow through the mantle cavity. Important differences were found between isolated tissue and whole animals, and between species. Drugs that stimulated ciliary beat in vitro did not increase water processing rate in vivo. None of the treatments increased water flow through the mantle cavity of intact animals. Results suggest that A. irradians was primarily modulating lateral ciliary activity, while M. edulis appeared to have a number of ways to control water processing activity, signifying that the two species may have different compensatory and regulatory mechanisms controlling feeding activity.

Analyzing Macromolecular Complexes in Situ Using Cellular Cryo‐Electron Microscopy

Cryo‐electron tomography (cryo‐ET) is a powerful technique for imaging biological structures in their native state and in an unperturbed cellular environment. We integrate high resolution imaging by either cryo‐ET and sub‐tomogram averaging or TYGRESS (Tomography‐Guided 3D Reconstruction of Subcellular Structures), with comparative genetics, biochemical methods and EM‐visible labeling to deconstruct the in situ 3D structure and functional organization of macromolecular complexes. We use cilia and flagella as model systems to advance techniques and approaches for high‐resolution imaging of complex cellular structures. Cilia and flagella are ubiquitous organelles with important biological roles in motility and sensation; defects in their assembly or function cause severe human diseases. The normal beating of cilia requires the precise coordination of thousands of dynein motors, but much remains to be learned about dynein's motility and regulation. We are in the process of generating a molecular blueprint of major ciliary complexes to gain a better understanding of the inner workings of these intriguing nano‐machines.

An investigation of non-Newtonian fluid flow due to metachronal beating of cilia in a tube

The aim of this study is to explain theoretically the role of ciliary motion on the transport of epididymal fluid through the ductus efferentes of the male reproductive track. For this purpose, a mathematical model has been developed for the flow of a non-Newtonian fluid in an axisymmetric tube due to metachronal wave of cilia motion for the more realistic consequences. Carreau viscous fluid model is considered to see the rheological effects on the pumping characteristics of the flow. Regular perturbation method has been employed to obtain the analytical expressions for the stream function, the velocity field and a relation between the pressure difference and the volume flow rate. It is found that the volume flow rate is influenced significantly by Weissenberg number We and the cilia length parameter ε. The computational results are presented graphically to see the effects of various physical parameters. Finally, the analysis is applied and compared with the observed value of the flow rate of spermatic fluid in the ductus efferentes of the male reproductive track. The volume flow rate is reported closed to the estimated value 6 × 10-3 ml/h in the human ductus efferentes when We = 0.5 and ε is near by 0.25.

Influence of magnetic field for metachoronical beating of cilia for nanofluid with Newtonian heating

Influence of magnetic field for metachoronical beating of cilia for nanofluid with Newtonian heating in an asymmetric channel is considered. The governing coupled equations are constructed under long wavelength and low Reynold's number approximation. The Newtonian heating is controlled by a dimensionless conjugate parameter, which varies between walls of channel. Numerical solutions are evaluated for nanoparticle fraction, heat transfer, stream function and pressure gradient. The important findings in this study are the variation of the conjugate parameter for Newtonian heating γ, Hartmann number M, thermophoresis parameter Nt and Brownian motion parameter Nb on pressure rise, nanoparticle fraction, heat transfer phenomena, pressure gradient and streamlines. The velocity field increases due to increase in M near the channel walls while velocity field decreases at the centre of the channel.

The planarian Schmidtea mediterranea as a model for studying motile cilia and multiciliated cells.

In the past few years, the freshwater planarian Schmidtea mediterranea has emerged as a powerful model system to study the assembly and function of cilia. S. mediterranea is a free-living flatworm that uses the beating of cilia on its ventral epidermis for locomotion. The ventral epidermis is composed of a single layer of multiciliated cells highly similar to the multiciliated cells that line the airway, the brain ventricles, and the oviducts in humans. The genome of S. mediterranea has been sequenced and efficient methods for targeting gene expression by RNA interference (RNAi) are available. Locomotion defects induced by perturbing the expression of ciliary genes can be often detected by simple visual screening, and more subtle defects can be detected by measuring locomotion speed. Cilia are present in large numbers and are directly accessible, which facilitates analyses by immunofluorescence and electron microscopy. Here we describe a set of methods for maintaining planarians in the lab. These include gene knockout by RNAi, cilia visualization by immunofluorescence, transmission electron microscopy, and live imaging.

The Effects of N-Acetyl Cysteine on Nasal Mucociliary Clearance in Healthy Volunteers: A Randomized, Double-Blind and Placebo-Controlled Study

Background: Mucociliary clearance is an important host defense function of the upper respiratory tract that requires the coordinated beating of cilia and results in the transport of mucus to the oropharynx. N-Acetyl Cysteine is a mucolytic drug currently used in pulmonary diseases. In the present study we sought to assess the effects of N-Acetyl Cysteine nasal mucociliary clearance in healthy volunteers. Methods: A total of 100 healthy individuals (55 male) with the mean age of 34.21 (± 12.63) years were included in the present randomized, double-blind and placebo-controlled study. Participants were assigned into two groups of case and control. The Mucociliary Clearance Time (MCT) was measured by saccharine test; measuring the time in minutes required for the subject to taste a saccharin particle placed on the inferior turbinate of the nasal. The variables studied were nasal MCT before and after taking the placebo or NAC, age, and sex. Data were analyzed using SPSS V.16. Results: Wilcoxon signed ranks test demonstrated considerable different of Saccharine Test Time (STT) between before and after taking the NAC (p=0.021), and no significant different of STT between before and after taking the placebo (p=0.723). Conclusion: N-Acetyl Cysteine exerts measurable effect on nasal mucociliary clearance in healthy volunteers; therefore may be beneficial in conditions associated with disruption of mucociliary clearance such as rhinitis and sinusitis. However, further studies are suggested to achieve more conclusive results.

A mathematical model for the flow of a Casson fluid due to metachronal beating of cilia in a tube.

A mathematical model is developed to study the transport mechanism of a Casson fluid flow inspired by the metachronal coordination between the beating cilia in a cylindrical tube. A two-dimensional system of nonlinear equations governing the flow problem is formulated by using axisymmetric cylindrical coordinates and then simplified by employing the long wavelength and low Reynolds number assumptions. Exact solutions are derived for the velocity components, the axial pressure gradient, and the stream function. However, the expressions for the pressure rise and the volume flow rate are evaluated numerically. The features of the flow characteristics such as pumping and trapping are illustrated and discussed with the help of graphs. It is observed that the volume flow rate is influenced significantly by the width of plug flow region H p as well as the cilia length parameter ε. The analysis is also applied and compared with the estimated value of the volume flow rate of epididymal fluid in the ductus efferentes of the human male reproductive tract.

Control of vertebrate core planar cell polarity protein localization and dynamics by Prickle 2.

Planar cell polarity (PCP) is a ubiquitous property of animal tissues and is essential for morphogenesis and homeostasis. In most cases, this fundamental property is governed by a deeply conserved set of 'core PCP' proteins, which includes the transmembrane proteins Van Gogh-like (Vangl) and Frizzled (Fzd), as well as the cytoplasmic effectors Prickle (Pk) and Dishevelled (Dvl). Asymmetric localization of these proteins is thought to be central to their function, and understanding the dynamics of these proteins is an important challenge in developmental biology. Among the processes that are organized by the core PCP proteins is the directional beating of cilia, such as those in the vertebrate node, airway and brain. Here, we exploit the live imaging capabilities of Xenopus to chart the progressive asymmetric localization of fluorescent reporters of Dvl1, Pk2 and Vangl1 in a planar polarized ciliated epithelium. Using this system, we also characterize the influence of Pk2 on the asymmetric dynamics of Vangl1 at the cell cortex, and we define regions of Pk2 that control its own localization and those impacting Vangl1. Finally, our data reveal a striking uncoupling of Vangl1 and Dvl1 asymmetry. This study advances our understanding of conserved PCP protein functions and also establishes a rapid, tractable platform to facilitate future in vivo studies of vertebrate PCP protein dynamics.

Heat transfer analysis of Rabinowitsch fluid flow due to metachronal wave of cilia

The present investigation concerns with the mechanical properties of a Rabinowitsch fluid model and the effects of thermal conductivity over it. Flow is considered to be occurring due to metachronal wave produced as a result of constant beating of cilia at the walls of a horizontal circular tube. The expressions for flow characteristics have been derived results are analyzed graphically and discussed briefly.

The planar cell polarity effector protein Wdpcp (Fritz) controls epithelial cell cortex dynamics via septins and actomyosin

Planar cell polarity (PCP) signaling controls polarized behaviors in diverse tissues, including the collective cell movements of gastrulation and the planar polarized beating of motile cilia. A major question in PCP signaling concerns the mechanisms linking this signaling cascade with more general cytoskeletal elements to drive polarized behavior. Previously, we reported that the PCP effector protein Wdpcp (formerly known as Fritz) interacts with septins and is critical for collective cell migration and cilia formation. Here, we report that Wdpcp is broadly involved in maintaining cortical tension in epithelial cells. In vivo 3D time-lapse imaging revealed that Wdpcp is necessary for basolateral plasma membrane stability in epithelial tissues, and we further show that Wdpcp controls cortical septin localization to maintain cortical rigidity in mucociliary epithelial cells. Finally, we show that Wdpcp acts via actomyosin to maintain balanced cortical tension in the epithelium. These data suggest that, in addition to its role in controlling plasma membrane dynamics in collective mesenchymal cell movements, Wdpcp is also essential for normal cell cortex stability during epithelial homeostasis.

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

Abstract 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.

Heat transfer study of an individual multiwalled carbon nanotube due to metachronal beating of cilia

In the present article, the consequences of cilia motion are reflected by the MWCNT. The problem is modeled in a symmetric channel with ciliated walls. Exact solutions of the governing flow problem are obtained for pressure gradient, temperature and velocities of the fluid. Streamlines for the velocity profile are plotted to discuss the trapping phenomenon.

Dry powder formulation of simvastatin

Objectives: This study focuses on the development of a dry powder inhaler (DPI) formulation of simvastatin (SV), and the effects of SV on the respiratory epithelium.Methods: Micronised SV samples were prepared by dry jet-milling. The long-term chemical stability and physicochemical properties of the formulations were characterised in terms of particles size, morphology, thermal and moisture responses. Furthermore, in vitro aerosol depositions were performed. The formulation was evaluated for cell viability and its effect on cilia beat activity, using ciliated nasal epithelial cells in vitro. The formulation transport across an established air interface Calu-3 bronchial epithelial cells and its ability to reduce mucus secretion was also investigated.Results: The particle size of the SV formulation and its aerosol performance were appropriate for inhalation therapy. Moreover, the formulation was found to be non-toxic to pulmonary epithelia cells and cilia beat activity up to a concentration of 10−6 M. Transport studies revealed that SV has the ability to penetrate into airway epithelial cells and is converted into its active SV hydroxy acid metabolite. Single dose of SV DPI also decreased mucus production after 4 days of dosing.Conclusion: This therapy could potentially be used for the local treatment of diseases like chronic obstructive pulmonary disease, cystic fibrosis, and bronchiectasis given its anti-inflammatory effects and ability to reduce mucus production.

Systematic discovery of novel ciliary genes through functional genomics in the zebrafish

Cilia are microtubule-based hair-like organelles that play many important roles in development and physiology, and are implicated in a rapidly expanding spectrum of human diseases, collectively termed ciliopathies. Primary ciliary dyskinesia (PCD), one of the most prevalent of ciliopathies, arises from abnormalities in the differentiation or motility of the motile cilia. Despite their biomedical importance, a methodical functional screen for ciliary genes has not been carried out in any vertebrate at the organismal level. We sought to systematically discover novel motile cilia genes by identifying the genes induced by Foxj1, a winged-helix transcription factor that has an evolutionarily conserved role as the master regulator of motile cilia biogenesis. Unexpectedly, we find that the majority of the Foxj1-induced genes have not been associated with cilia before. To characterize these novel putative ciliary genes, we subjected 50 randomly selected candidates to a systematic functional phenotypic screen in zebrafish embryos. Remarkably, we find that over 60% are required for ciliary differentiation or function, whereas 30% of the proteins encoded by these genes localize to motile cilia. We also show that these genes regulate the proper differentiation and beating of motile cilia. This collection of Foxj1-induced genes will be invaluable for furthering our understanding of ciliary biology, and in the identification of new mutations underlying ciliary disorders in humans.