Auswahl der wissenschaftlichen Literatur zum Thema „Multicilia“

Cilia are evolutionarily conserved microtubule-based structures that perform diverse biological functions. Cilia are assembled on basal bodies and anchored to the plasma membrane via distal appendages. In the male reproductive tract, multicilia in efferent ducts (EDs) move in a whip-like motion to prevent sperm agglutination. Previously, we demonstrated that the distal appendage protein CEP164 recruits Chibby1 (Cby1) to basal bodies to facilitate basal body docking and ciliogenesis. Mice lacking CEP164 in multiciliated cells (MCCs) (FoxJ1-Cre;CEP164fl/fl) show a significant loss of multicilia in the trachea, oviduct, and ependyma. In addition, we observed male sterility; however, the precise role of CEP164 in male fertility remained unknown. Here, we report that the seminiferous tubules and rete testis of FoxJ1-Cre;CEP164fl/fl mice exhibit substantial dilation, indicative of dysfunctional multicilia in the EDs. We found that multicilia were hardly detectable in the EDs of FoxJ1-Cre;CEP164fl/fl mice although FoxJ1-positive immature cells were present. Sperm aggregation and agglutination were commonly noticeable in the lumen of the seminiferous tubules and EDs of FoxJ1-Cre;CEP164fl/fl mice. In FoxJ1-Cre;CEP164fl/fl mice, the apical localization of Cby1 and the transition zone marker NPHP1 was severely diminished, suggesting basal body docking defects. TEM analysis of EDs further confirmed basal body accumulation in the cytoplasm of MCCs. Collectively, we conclude that male infertility in FoxJ1-Cre;CEP164fl/fl mice is caused by sperm agglutination and obstruction of EDs due to loss of multicilia. Our study, therefore, unravels an essential role of the distal appendage protein CEP164 in male fertility.

Cilia are microtubule-based hair-like organelles on the cell surface. Cilia have been implicated in various biological processes ranging from mechanosensation to fluid movement. Ciliary dysfunction leads to a plethora of human diseases, known as ciliopathies. Although non-motile primary cilia are ubiquitous, motile multicilia are found in restricted locations of the body, such as the respiratory tract, the oviduct, the efferent duct, and the brain ventricles. Multicilia beat in a whip-like motion to generate fluid flow over the apical surface of an epithelium. The concerted ciliary motion provides the driving force critical for clearing airway mucus and debris, transporting ova from the ovary to the uterus, maintaining sperm in suspension, and circulating cerebrospinal fluid in the brain. In the male reproductive tract, multiciliated cells (MCCs) were first described in the mid-1800s, but their importance in male fertility remained elusive until recently. MCCs exist in the efferent ducts, which are small, highly convoluted tubules that connect the testis to the epididymis and play an essential role in male fertility. In this review, we will introduce multiciliogenesis, discuss mouse models of male infertility with defective multicilia, and summarize our current knowledge on the biological function of multicilia in the male reproductive tract.

Recent molecular phylogenetic studies have led to the erection of the phylum Amoebozoa, uniting naked and testate lobose amoebae, the mycetozoan slime moulds and amitochondriate amoeboid protists (Archamoebae). Molecular data together with ultrastructural evidence have suggested a close relationship between Mycetozoa and Archamoebae, classified together in the Conosea, which was named after the cone of microtubules that, when present, is characteristic of their kinetids. However, the relationships of conoseans to other amoebozoans remain unclear. Here, we obtained the complete small-subunit (SSU) rRNA gene sequence (2746 bp) of the enigmatic, multiflagellated protist Multicilia marina, which has formerly been classified either in a distinct phylum, Multiflagellata, or among lobose amoebae. Our study clearly shows that Multicilia marina belongs to the Amoebozoa. Phylogenetic analyses including 60 amoebozoan SSU rRNA gene sequences revealed that Multicilia marina branches at the base of the Conosea, together with another flagellated amoebozoan, Phalansterium solitarium, as well as with Gephyramoeba sp., Filamoeba nolandi and two unidentified amoebae. This is the first report showing strong support for a clade containing all flagellated amoebozoans and we discuss the position of the root of the phylum Amoebozoa in the light of this result.

CAMSAP3, a protein that controls microtubule dynamics by binding to its minus-end, is localized at proximal regions of the axoneme in tracheal motile cilia. Its dysfunction results in a collapse of the central microtube pair, basal plate disorganization, and uncoordinated beating of multicilia.

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.

Centriole number is normally under tight control and is directly linked to ciliogenesis. In cells that use centrosomes as mitotic spindle poles, one pre-existing mother centriole is allowed to duplicate only one daughter centriole per cell cycle. In multiciliated cells, however, many centrioles are generated to serve as basal bodies of the cilia. Although deuterosomes were observed more than 40 years ago using electron microscopy and are believed to produce most of the basal bodies in a mother centriole-independent manner, the underlying molecular mechanisms have remained unknown until recently. From these findings arise more questions and a call for clarifications that will require multidisciplinary efforts.

The airway is covered by multicilia that beat in a metachronous manner toward the mouth to eliminate debris and infectious particles. Coordinated one-directional beating is an essential feature of multicilia in the airway to guarantee proper mucociliary clearance. Defects in ciliary motility lead to primary ciliary dyskinesia (PCD), with major symptoms including bronchitis and other chronic respiratory diseases. Recent work suggested that ciliary motility and planar polarity are required in the process of ciliary alignment that produces coordinated beating. However, the extent to which cilia motility is involved in this process in mammals has not yet been fully clarified. Here, to address the role of ciliary motility in the process of coordinated ciliary alignment, we analyzed Kintoun mice mutants ( Ktu−/−). Ktu−/− exhibited typical phenotypes of PCD with complete loss of ciliary motility in trachea and another ciliated tissue, the brain ependyma. Immunohistochemistry using antibodies against axonemal dynein confirmed the loss of multiple axonemal dynein components in mutant cilia. Observation of cilia orientation based on basal foot directions revealed that ciliary motility was not required in the alignment of airway cilia, whereas a strong requirement was observed in brain ependymal cells. Thus we conclude that the involvement of ciliary motility in the establishment of coordinated ciliary alignment varies among tissues.

La scoliose idiopathique (SI) est une déformation tridimensionnelle de la colonne vertébrale très répandue qui touche 3 à 4 % de la population en l'absence d'anomalies congénitales évidentes, et dont l'étiologie est mal comprise. Des études génétiques récentes chez le poisson zèbre ont montré que la perte de la fibre de Reissner (FR), un polymère protéique de sco-spondine sécrété par l'organe sous-commissural (SCO) et présent le long des cavités du système nerveux central, déclenche une scoliose chez les juvéniles (correspondant à "l'adolescence"). L'altération de la motilité des cils dans les embryons, nécessaire à la polymérisation du RF, entraîne également des défauts de courbure de l'axe, en corrélation avec une diminution de l'expression du gène du neuropeptide urp2 chez les mutants sco-spondine. Il n'est pas encore déterminé si ce scénario, où un défaut de circulation du LCR entraînant la perte du FR et une diminution de la signalisation de l'URP s'applique à d'autres mutants de poisson-zèbre qui n'altèrent pas directement la motilité des cils ou s'il peut être pertinent pour la SI humaine. Deux études ont également mis en évidence l'existence d'une signature neuro-inflammatoire, en aval de la perte du FR, qui contribue à la sévérité et à la pénétrance de la scoliose. Nous avons généré un mutant de poisson zèbre pour rpgrip1l, un gène codant pour une protéine de la zone de transition ciliaire, qui ne présente aucune anomalie embryonnaire et développe une scoliose avec une pénétrance presque complète chez les juvéniles. Le but de ma thèse était de déterminer la cascade d'événements menant à la scoliose chez les rpgrip1l-/-. Nous avons tiré profit du développement asynchrone de la scoliose chez rpgrip1l-/- pour montrer que les mutants droits présentent déjà des défauts ciliaires au niveau du tronc avec une augmentation du nombre de cellules immunitaires dans le cerveau et une augmentation du niveau d'expression d'urp1 et d'urp2. A l'apparition de la scoliose, les mutants rpgrip1l-/- perdent la FR et les touffes multi-ciliées proches du SCO. La réintroduction de RPGRIP1L dans les cellules ciliées motiles et les cellules progénitrices grâce à une transgénèse tissu-spécifique prévient l'apparition de la scoliose. En réduisant le niveau d'URP chez les rpgrip1l-/- par des croisements génétiques, nous avons démontré que l'altération de la signalisation URP ne contribue pas à la scoliose chez rpgrip1l-/-. De plus, un traitement anti-inflammatoire/anti-oxydant à long terme a réduit la sévérité et la pénétrance de la scoliose de 50%, suggérant que ces processus sont impliqués dans l'apparition et l'évolution de la scoliose chez rpgrip1l-/-. Enfin, grâce à une analyse protéomique et à des immuno-marquages, nous avons mis en évidence une astrogliose au sein du SCO et du ventricule rhombencéphalique qui se développe de manière asynchrone chez les mutants droits rpgrip1l-/- et qui persiste chez les mutants scoliotiques. Nous proposons que l'astrogliose au niveau du SCO associée au recrutement de cellules inflammatoires induisent localement la perte de touffes multi-ciliées, altérant la polymérisation du FR et conduisant finalement à la scoliose. Nous espérons que cette étude permettra de mieux comprendre les défauts moléculaires et cellulaires à l'origine de l'IS chez le poisson zèbre et qu'elle mettra en évidence l'astrogliose cérébrale en tant que défaut potentiel à l'origine de la SI chez l'homme
Idiopathic scoliosis (IS) is a highly prevalent three-dimensional spine deformity which affects 3-4% of the population in the absence of obvious congenital abnormalities, whose etiology is poorly understood. Recent genetic studies in zebrafish have shown that loss of Reissner fiber (RF), a sco-spondin protein polymer secreted by the subcomissural organ (SCO) and present along the cavities of the central nervous system, triggers scoliosis in juveniles (the "adolescent-like" stage). Impairment of cilia motility in embryos, necessary for RF polymerization, also leads to axis curvature defects, that correlates with decreased urp2 neuropeptide gene expression in sco-spondin mutants. It is yet not clear whether this scenario of CSF flow defect causing RF loss and decreased URP signaling holds true in other zebrafish mutants that do not impair directly cilia motility or may be relevant to human IS. Two studies also pointed to the existence of a neuro-inflammatory signature, downstream of RF loss, which contributes to scoliosis severity and penetrance. We generated a zebrafish mutant for rpgrip1l, a gene encoding a ciliary transition zone protein, which displays no embryonic anomaly and develops scoliosis with nearly full penetrance in juveniles. The goal of my thesis was to decipher the cascade of events leading to scoliosis in rpgrip1l-/-. We took advantage of the asynchronous scoliosis development in rpgrip1l-/- to show that the straight mutants already presented ciliary defects at trunk level with increase number of immune cells within the brain and urp1 and urp2 upregulation. At scoliosis onset, rpgrip1l-/- mutants had lost the RF and specific multi-ciliated tufts just lateral to the SCO. Reintroduction of RPGRIP1L into motile ciliated cells and progenitor cells thanks to tissue-specific transgenesis prevents scoliosis onset. By reducing URP level in rpgrip1l-/- via genetic crosses, we demonstrated that alteration of URP signaling does not contribute to the curvature phenotype of rpgrip1l-/-. Moreover, long-term anti-inflammatory/anti-oxidant treatment reduced the severity and penetrance of scoliosis by 50%, suggesting that inflammatory and/or oxidative processes are involved in rpgrip1l scoliosis onset and evolution. Finally, thanks to a proteomic analysis and immunostaining, we revealed an astrogliosis response within the SCO and rhombencephalic ventricle that develops asynchronously in straight rpgrip1l-/- and persists in scoliotic fish. We propose that the astrogliosis at SCO level associated with inflammatory cells recruitment induces locally the loss of multi-ciliated tufts, impairing RF polymerization and eventually leading to scoliosis. We hope these studies will provide new insights into the understanding of molecular and cellular defects leading to IS in zebrafish and highlight brain astrogliosis as a potential defect leading to human IS

Le processus de formation des cils mobiles multiples (multiciliogénèse) est composé de nombreuses étapes. Récemment, nous avons démontré que les microARNs de la famille miR-449 contrôlent plusieurs de ces étapes. Au cours de mon travail, je me suis concentré sur le rôle joué par miR-449 dans deux aspects du développement de l'épithélium multicilié. Dans les cellules multiciliées, un réseau dense d'actine sous-jacent l'aspect apicale de la membrane cellulaire (coiffe d'actine) est nécessaire pour l'ancrage des multiples corps basaux, et donc pour une ciliogenèse approprié. Dans le cadre de mon travail, j'ai participé à l' identification de la petite GTPase R-Ras comme une des véritables cibles de miR-449. J'ai démontré que la réorganisation de la coiffe d'actine et l'ensemble du processus de multiciliogénèse étaient compromis lorsque l'ARN messager de R-Ras se trouve protégé de la liaison avec miR-449. J'ai aussi contribué à identifier une nouvelle cible de miR-449, le gène Steel, qui code pour le ligand du récepteur transmembranaire à tyrosine-kinase KIT. La repression de Steel par miR449 est impliquée dans le processus par lequel les cellules multiciliées atteignent leur position finale dans l'épiderme embryonnaire de Xenopus. STEEL, qui agit probablement comme une molécule de guidage pour les cellules multiciliées qui expriment KIT, doit être réprimé par miR-449 dans ces mêmes cellules en cours de migration pour assurer leur deplacement directionnel approprié. En conclusion, mon travail a contribué à élucider le rôle complexe joué par le miARN miR-449 dans le processus de multiciliogénèse chez les vertébrés
The process of multiple motile cilia formation (multiciliogenesis) is composed of many different steps. Recently, we demonstrated that microRNAs of the miR-449 family control several of these steps. During my work, I focused on the role played by miR-449 in two aspects of the development of the mucociliary epithelium. In multiciliated cells, a dense actin network underlying the apical aspect of the cell membrane (actin cap) is required for the anchoring of the multiple basal bodies, and therefore for proper ciliogenesis. Small GTPases play important role in the formation of the actin cap. In the course of my work, I took part in the identification of transcripts coding the small GTPase R-Ras as bona fide targets of miR-449. I demonstrated that apical and subapical actin network reorganization and multiciliogenesis were impaired when R-Ras mRNA was protected from miR-449 binding. Moreover, the actin cap formation and multiciliogenesis were rescued when the translation of protected R-Ras transcripts was prevented. I also contributed to the finding that a new miR-449 target, the KIT receptor tyrosin kinase ligand STEEL, is involved in the process through which the multiciliated cells reach their final position within the developing frog epidermis. STEEL, which likely acts as a guidance molecule for the KIT-expressing multiciliated cells, needs to be repressed by miR-449 within the migrating cells to ensure their proper directional migration. Altogether, my work contributed to elucidate the complex role played by the miR-449 miRNA in the process of vertebrate multiciliogenesis

The subject of the diploma thesis is the design of a city block on brownfield areas in the Trnitá district of Brno. The aim was to make the most out of the land, minimize motor traffic and create objects with different functions (housing, administration, amenities, services) will be mixed. The main part of the whole area will be the Ponávka river, which will take advantage of the great potential of the watercourse, thus creating a quality public space for recreation. The principles of multiplicity and porosity have been used to design - from the maximum land abandonment, the mass was gradually taken away to create apartment houses, administrative areas, a common parcel with civic amenities and a walkable green roofs.

Chez les vertébrés, le battement coordonné des cils motiles présents par centaines à la surface apicale des cellules multiciliées (MCC) est requis pour propulser directionnellement les fluides biologiques à l'intérieur de certains organes (voies respiratoires, ventricules cérébraux, trompes utérines ou certaines structures embryonnaires). De nombreuses pathologies humaines sont associées à des défauts ciliaires ou à une perte des MCC (dyskinésies ciliaires, mucoviscidose, asthme,...). Dans ce contexte, mon travail de thèse a consisté à élucider les mécanismes complexes contrôlant la différenciation des MCC et donc la formation des cils motiles (multiciliogenèse). Par des approches de génomiques fonctionnelles à partir de deux modèles d'épithéliums multiciliés évolutivement éloignés (épithélium respiratoire humain et épiderme d'embryon de Xénope) nous avons identifié la famille des microARN (petits ARN non-codants régulateurs de l'expression génique) miR-449 comme majoritairement exprimée dans les MCC. Nous avons montré que miR-449 contrôle la multiciliogenèse i) en bloquant le cycle cellulaire, ii) en réprimant directement la voie de signalisation Notch et iii) en inhibant l'expression de la petite GTPase R-Ras. Enfin, nos travaux montrent que l'ensemble de ces mécanismes est conservé chez les vertébrés. En conclusion, miR-449 est un nouveau régulateur clé de la multiciliogenèse conservé au cours de l'évolution. Nos résultats pourraient ouvrir la voie à de nouvelles stratégies thérapeutiques utilisant des petits ARN régulateurs dans le traitement de certaines pathologies associées à des défauts ciliaires.

The strategy of information service’s marketing (case studies at pt. data consult,inc. and pt. agranet multicitra siberkom). Depok : University of Indonesia - Thesis - Postgraduate (S2) Department of Library Science, Faculty of Cultural Science. This research proposed to analize (1) The role of mission in implementing marketing strategy (2) The role of market analysis in implementing marketing strategy, and (3) The strategy of allocating resource which included strategy of 7 (seven) elements of marketing mix (products, price, place/distribution channel, promotion, process, people and phisical evidence) in implementing marketing strategy at PT. Data Consult, Inc and PT. Agranet Multicitra Siberkom / Agrakom especially for the product/services of Detikcom in period of 2001-2003. This research is a qualitative research using assessment data methods : in-dept interview either with direct conversation or e-mail, observation in the rules of reseacher-participant and bibliotheque. In doing this research, researcher enrich herself with open standards interview guide.The informan of this reseach are the decision makers, especially in marketing policy, consist of one staff from PT. Data Consult, Inc. and four staff from PT.Agranet Multicitra Siberkom Results of research at PT.Data Consult, Inc. showed (1) Company mission play in implementing marketing strategy, which is customized with current condition, (2) Market analysis play in any current applied marketing policy in the company, (3) The strategy of allocating resource which included strategy of 7 (seven) elements of marketing mix (products, price, place/distribution channel, promotion, process, people and phisical evidence) has been conducted in the process of implementation of marketing strategy at Data Consult. Perceiving the increased selling data of the whole products / services offered in period of 2001-2003, it may concluded if the implementation of marketing strategy at Data Consult in periode of 2001-2003 has been succesfully increased the amount of information products/services selling at Data Consult in periode of 2001-2003. Result of research at PT.Agranet Multicitra Siberkom showed that the basic strategy which is implemented by Agrakom in develop Detikcom is the principle of ‘how to make revenue is bigger than cost’. To achieve the goal, they developed marketing strategies which is suitable with online business characteristic. Basically, the mission and the result of analysis of the market play a big role in implementing marketing strategy of Detikcom. Performance evaluation of Detikcom in implementing marketing strategy above all seven elements of marketing mix (products, price, place/distribution channel, promotion, process, people and phisical evidence), able to be seen from the growth of sum of page view (opened pages) and sum of advertising which published at Detikcom in period of 2001-2003. Thus, it may concluded that the implementation of marketing strategy of Detikcom in periode 2001-2003 has been succesfully enhanced the sum of visitor and the ads selling in periode 2001-2003.