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Control of the orientation of cilia by adenosinetriphosphate, calcium, and zinc in glycerol-extracted Paramecium caudatum.

The predominant orientation of cilia in glycerol-extracted Paramecium is toward the posterior of the specimen in a KCl solution. The cilia became reoriented toward the anterior shortly after transfer of the extracted cell to a mixture of ATP, calcium, and zinc. The degree of response was graded as a function of the concentration of each of the three essential factors. Minimum concentrations for the maximum response were 0.2 mM in ATP, 0.8 mM in calcium, and 0.0002 mM in zinc. The observations support the hypothesis that cation-induced ciliary reversal in live specimens is initiated by calcium ions which become displaced from an inferred cellular cation exchanger system.

Adenosine Triphosphate↗

Two heteromeric kinesin complexes in chemosensory neurons and sensory cilia of Caenorhabditis elegans.

Chemosensation in the nervous system of the nematode Caenorhabditis elegans depends on sensory cilia, whose assembly and maintenance requires the transport of components such as axonemal proteins and signal transduction machinery to their site of incorporation into ciliary structures. Members of the heteromeric kinesin family of microtubule motors are prime candidates for playing key roles in these transport events. Here we describe the molecular characterization and partial purification of two heteromeric kinesin complexes from C. elegans, heterotrimeric CeKinesin-II and dimeric CeOsm-3. Transgenic worms expressing green fluorescent protein driven by endogenous heteromeric kinesin promoters reveal that both CeKinesin-II and CeOsm-3 are expressed in amphid, inner labial, and phasmid chemosensory neurons. Additionally, immunolocalization experiments on fixed worms show an intense concentration of CeKinesin-II and CeOsm-3 polypeptides in the ciliated endings of these chemosensory neurons and a punctate localization pattern in the corresponding cell bodies and dendrites. These results, together with the phenotypes of known mutants in the pathway of sensory ciliary assembly, suggest that CeKinesin-II and CeOsm-3 drive the transport of ciliary components required for sequential steps in the assembly of chemosensory cilia.

Amino Acid Sequence↗

Bardet-Biedl syndrome genes are important in retrograde intracellular trafficking and Kupffer's vesicle cilia function.

Bardet-Biedl syndrome (BBS) is characterized by obesity, retinopathy, polydactyly, cognitive impairment, renal and cardiac anomalies as well as hypertension and diabetes. The nine known BBS genes do not appear to belong to the same functional category; yet mutation of these genes results in a nearly identical pleiotropic phenotype. Although the precise functions of the BBS proteins have yet to be determined, current data support a role in cilia function and intraflagellar transport. To gain insight into the biological processes controlled by BBS genes, we embarked on studies of six BBS orthologues from zebrafish. Knockdown of zebrafish bbs2, bbs4, bbs5, bbs6, bbs7 or bbs8 results in disruption of Kupffer's vesicle (KV), a ciliated organ thought to play a role in left-right patterning. KV defects are due to a progressive loss of cilia within the vesicle and result in subsequent alterations to organ laterality. We also note a specific defect altering retrograde melanosome transport. These studies are the first to comprehensively compare the diverse group of BBS genes in parallel and demonstrate a common role in intracellular trafficking, indicating that BBS proteins are involved in general organelle trafficking.

Animals↗

Function and structure of cilia in the fallopian tube of an infertile woman with Kartagener's syndrome.

In Kartagener's syndrome (KS), primary defects of the ciliary axoneme cause dyskinetic ciliary motion. Because ciliary motion is an important factor in normal ovum transport, ciliary dyskinesia may cause infertility. On the other hand, the existence of some ciliary activity, albeit abnormal, may account for fertility in some women with KS. In this case study, an infertile woman diagnosed with KS had normal results in all usual infertility tests. Biopsies of tubal mucosa were obtained at laparoscopy for ovum recovery during an in-vitro fertilization cycle. Ciliary activity, measured by laser light-scattering spectroscopy, was detected in all tubal specimens; however the majority of regions sampled showed no activity. In active regions, beat frequency ranged from 5 to 10 Hz, approximately 30% of normal. Electron microscopy showed similar morphological defects in both tubal and nasal mucosa. The number of cilia per cell was approximately 20% of normal. The major ultrastructural abnormality of cilia was an absence of the central microtubules. The only demonstrable explanation for this patient's infertility was primary ciliary dyskinesia associated with KS.

Adult↗

Homologues of radial spoke head proteins interact with Ca2+/calmodulin in Tetrahymena cilia.

Calmodulin (CaM) is an axonemal component. To examine the pathway of Ca(2+)/CaM signaling in cilia, using Ca(2+)/CaM-affinity column, we identified seven Ca(2+)/CaM-associated proteins from a crude dynein fraction and isolated 62 kDa (p62) and 66 kDa (p66) Ca(2+)/CaM-associated proteins in Tetrahymena cilia. The amino acid sequences deduced from the p62 and p66 cDNA sequences suggested that these proteins were similar to Chlamydomonas radial spoke proteins 4 and 6 (RSP4 and RSP6), components of the radial spoke head, and sea urchin sperm p63, which is a homologue of RSP4/6, and isolated as a key component that affect flagellar bending patterns. Although p62 and p66 do not have a conventional CaM-binding site, those have consecutive sequences which showed high normalized scores (>or= 5) from a CaM target database. These consecutive sequences were also found in RSP4, RSP6, and p63. These radial spoke heads proteins have a high similarity region composed of 15 amino acids between the five proteins. Immunoelectron microscopy using anti-CaM antibody showed that CaM was localized along the outer edge of the curved central pair microtubules in axoneme. Therefore, it is possible that the interaction between Ca(2+)/CaM and radial spoke head control axonemal curvature in the ciliary and flagellar waveform.

Amino Acid Sequence↗

Intraflagellar transport and cilia-dependent renal disease: the ciliary hypothesis of polycystic kidney disease.

Epithelial cells that line mammalian kidney nephrons have solitary nonmotile primary cilium projecting from their surface into the lumens of the ducts and tubules. Mutations that block the assembly of these cilia cause cystic kidney disease. The products of human autosomal dominant and recessive polycystic kidney disease genes and products of the nephronophthisis disease genes are at least partially localized to primary cilia. This suggests that the cilium serves as an organizing center for the early steps of the signal transduction pathway that is responsible for monitoring the integrity of the kidney nephron and controlling cell proliferation and differentiation.

Animals↗

Spermatozoa and cilia lacking axoneme in an infertile man.

A case of absence of axoneme both in spermatozoa and in the respiratory tract of the same individual is described. Spermatozoa are normal in shape and in head structure, but have tails containing only mitochondria, accessory fibers and fibrous sheath. Cilia in respiratory cells are present and normally long, but empty, flattened, devoid of microtubules and centrioles, resembling mucous cells microvilli, which are quite shorter. The role of the axoneme in the accessory fibers and cilia morphogenesis is discussed.

Adult↗

Cilium-attached and excised patch-clamp recordings of odourant-activated Ca-dependent K channels from chemosensory cilia of olfactory receptor neurons.

It has previously been proposed that a Ca2+-dependent K+ conductance is implicated in the inhibitory odourant response in rat and toad olfactory receptor neurons. Previous whole-cell and single-channel measurements on inside-out excised patches, in addition to immunochemical evidence, indicated the presence of Ca2+-dependent K+ channels in olfactory cilia, the transducing structures of these sensory cells. Ca2+-dependent K+ channels opened in 'on-cilium' membrane patches from C. caudiverbera upon odourant stimulation. Furthermore, after excision in the inside-out configuration, the channel could be opened by micromolar Ca2+, in a Ca2+-dependent fashion, but it was unresponsive to cyclic AMP. We estimated that the Ca2+ concentration in the proximity of a Ca2+-dependent K+ channel within the cilia reaches at least 100 microM during the odour response. The K+ channel displayed a higher selectivity for K+ than for Na+. Our results support a role for this Ca2+-dependent K+ channel in chemotransduction.

Animals↗

Calcium regulates the regeneration of cilia in Tetrahymena thermophila.

A study of calcium metabolism in Tetrahymena during the regeneration of cilia evidenced that the process is inhibited by nifedipine and trifluoperazine. This suggests that calcium ions play an important regulatory role in this process. This was confirmed by studies on calcium uptake and efflux which showed that there was a net increase in calcium uptake prior to the reinitiation of motility. The increase coincided with a period of sensitivity to the calcium antagonist TMB-8 and with an increase in the intracellular level of cGMP. The process was also inhibited by neomycin and stimulated by phorbol esters, which suggests that hydrolysis of phosphatidylinositol phosphates may take place as part of the calcium regulatory network during the regeneration of cilia.

Animals↗

Cilia in nematode sensory organs.

Electron microscopic studies revealed thle presenlce of true cilia in nerve processes connected with sensory organs of a nemnatodes. These structures are important in evaluating the relation between nematodes and the other aschelminths, from which they were separated partially on the basis of the supposed total absence of cilia.

Animals↗

Cilia regeneration in the sea urchin embryo: evidence for a pool of ciliary proteins.

Late gastrulae of paracentrotus lividus regenerated cilia after being deciliated in hypertonic sea water. Regeneration was not affected by actinomycin D or puromycin. Actinomycin D also did not affect ciliary protein synthesis during regeneration, although overall embryonic synthesis was depressed. Puromycin inhibited both total embryonic and ciliary protein synthesis. The data indicate that ciliary protein synthesis is controlled by a stable template and that the regenerating cilia are formed from a pool of ciliary proteins. It is suggested that the proteins of the mitotic apparatus and of the ciliamay be related.

Animals↗

Modified cilia in sensory organs of juvenile stages of a parasitic nematode.

Electron microscopical studies revealed the presence of dendritic nerve processes in sensory organs of the third- and fourth-stage juveniles of Haemonchus contortus, which contained structures resembling modified cilia. With few exceptions, the outer circle of fibers consisted of ten doublets, and in place of typical cilia-like central fibers were small microtubules or vesicles varying in number from zero to five.

Animals↗

Colchicine-inhibited cilia regeneration: explanation for lack of effect in tris buffer medium.

In Stentor coeruleus growth of new, daughter ciliates and experimentaly inducled regeneration of oral membranellar cilia are reversibly inhibited by low, nontoxic concentrations of colchicine. However, if the clulture medium containing colchicine (or Colcemid) is made up in tris(hydroxymethyl)aminomethane buffer, growth of ciliated daughters and regeneration of oral cilia proceed normally. The evidence suggests that the mechanism of this reversal of the effects of colchicine (or Colcemid) is due to a chemical reaction between tris(hydroxymethyl)-aminomethane (or its hydrochloride, or both) and colchicine (or Colcemid), wihich reduces the effective concentration of these mitotic spindle inhibitors reaching the stentors.

Animals↗

Effect of ciliostatic factors from Pseudomonas aeruginosa on rabbit respiratory cilia.

Heat-stable factors released by Pseudomonas aeruginosa in culture supernatants inhibit functional cilia of rabbit tracheal epithelium. Chloroform extraction removed heat-stable factors from stationary-phase culture supernatants. The extracts contained at least seven components separable by thin-layer chromatography (TLC). Cilioinhibitory components were identified as a phenazine derivative, pyo compounds (2-alkyl-4-hydroxyquinolines), and a rhamnolipid, also known as a hemolysin. Fluorescence and absorption spectra, relative migration on TLC, staining characteristics, and gas chromatography were the basis for identification. Inhibitory concentrations of each active component were established by quantitative measures of percent motility and beat frequency. Corresponding damage to ciliary ultrastructure was examined by electron microscopy. The pyo compounds produced ciliostasis at concentrations of 50 micrograms/ml, but without obvious ultrastructural lesions. The phenazine derivative also inhibited ciliary motility and caused some membrane disruption, although at substantially greater concentrations of 400 micrograms/ml. Limited exposure of tracheal explants to the rhamnolipid resulted in ciliostasis which was associated with altered ciliary membranes. More extensive exposure to rhamnolipid was associated with removal of dynein arms from axonemes. Pyocyanin at a concentration of 0.5 mg/ml did not inhibit ciliary beating under our conditions. The data suggest that the pyo compounds are the most effective per weight ciliostatic factors released by P. aeruginosa and rhamnolipid is the most destructive of cilia ultrastructure. By interfering with normal ciliary function, these ciliostatic factors may enable P. aeruginosa to more easily colonize the respiratory tract.

Animals↗

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

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

Animals↗

A temperature-sensitive mutation affecting cilia regeneration, nuclear development, and the cell cycle of Tetrahymena thermophila is rescued by cytoplasmic exchange.

A temperature-sensitive mutation was isolated that blocks cilia regeneration and arrests growth in Tetrahymena thermophila. Protein and RNA synthesis and ATP production appeared to be largely unaffected at the restrictive temperature, suggesting that the mutation is specific for cilia regeneration and growth. At the restrictive temperature, mutant cells arrested at a specific point in the cell cycle, after macronuclear S phase and shortly before micronuclear mitosis. Arrested cells did not undergo nuclear divisions, DNA replication, or cytokinesis, so the mutation appears to cause true cell cycle arrest. Surprisingly, the mutation does not appear to affect micronuclear mitosis directly but rather some event(s) prior to micronuclear mitosis that must be completed before cells can complete the cell cycle. The cell cycle arrest was transiently complemented by wild-type cytoplasm exchanged during conjugation with a wild-type cell. Each starved, wild-type cell apparently contained enough rescuing factor to support an average of six cell divisions. Thus, this mutation affects assembly and/or function of at least one but not all of the microtubule-based structures in T. thermophila.

Animals↗

Tolerance of the eye for implanted cilia.

Retained cilia in the eye following injury are rare. There are reports of long term tolerance to cilia in the anterior chamber but no information about the effect on the corneal endothelium. A 31-year-old man was seen with a 19-year history of a retained eyelash embedded in a corneal scar and extending across the anterior chamber to the chamber angle. The follicle was absent. There was no cellular inflammatory reaction, and the eye otherwise appeared normal on slit-lamp examination with normal vision. However, there was the appearance of an encrusted deposit on the surface of the lash and a markedly lowered endothelial cell count of 750 cells/mm2 compared with the fellow eye. The lash was not removed.

Adult↗