A 210 kDa protein is located in a membrane-microtubule linker at the distal end of mature and nascent basal bodies

1999 ◽  
Vol 112 (11) ◽  
pp. 1633-1644 ◽  
Author(s):  
K.F. Lechtreck ◽  
A. Teltenkotter ◽  
A. Grunow
Keyword(s):  
Basal Body ◽  
Body Position ◽  
Immunogold Electron Microscopy ◽  
Basal Bodies ◽  
Western Blots ◽  
Whole Cells ◽  
Extracellular Matrix Components ◽  
Green Flagellate ◽  
Protein Rich ◽  
Novel Protein

A monoclonal antibody raised against purified flagellar basal apparatuses from the green flagellate Spermatozopsis similis reacted with a protein of 210 kDa (p210) in western blots. The protein was partially cloned by immunoscreening of a cDNA library. The sequence encoded a novel protein rich in alanine (25%) and proline (20%), which contained regions similar to proteins of comparable amino acid composition such as extracellular matrix components or the membrane-cytoskeletal linker synapsin. Using a polyclonal antibody (anti-p210) raised against the C-terminal part of p210, it was shown that the protein was highly enriched in the basal apparatuses. Immunogold electron microscopy of isolated cytoskeletons or whole cells revealed that p210 was located in the flagellar transition region. The protein was part of the Y-shaped fibrous linkers between the doublet microtubules and the flagellar membrane, as indicated by statistical analysis of post-labeled sections using anti-centrin and anti-tubulin as controls. In premitotic cells p210 was located in a fibrous layer at the distal end of nascent basal bodies, which was perforated by the outgrowing axoneme. During deflagellation the protein remained at the basal body but we observed changes in its distribution, indicating that p210 partially moved to the tip of the basal body. p210 can be used as a marker to determine basal body position, orientation (parallel or antiparallel) and number in S. similis by indirect immunofluorescence. We suppose that p210 is involved in linking basal bodies to the plasma membrane, which is an important step during ciliogenesis.

scholarly journals The UNI1 and UNI2 Genes Function in the Transition of Triplet to Doublet Microtubules between the Centriole and Cilium in Chlamydomonas

2009 ◽  
Vol 20 (1) ◽  
pp. 368-378 ◽  
Author(s):  
Brian P. Piasecki ◽  
Carolyn D. Silflow
Keyword(s):  
Basal Body ◽  
Double Mutant ◽  
Genetic System ◽  
Eukaryotic Cells ◽  
Immunogold Electron Microscopy ◽  
Basal Bodies ◽  
Wild Type ◽  
Unicellular Alga ◽  
Mutant Cells

One fundamental role of the centriole in eukaryotic cells is to nucleate the growth of cilia. The unicellular alga Chlamydomonas reinhardtii provides a simple genetic system to study the role of the centriole in ciliogenesis. Wild-type cells are biflagellate, but “uni” mutations result in failure of some centrioles (basal bodies) to assemble cilia (flagella). Serial transverse sections through basal bodies in uni1 and uni2 single and double mutant cells revealed a previously undescribed defect in the transition of triplet microtubules to doublet microtubules, a defect correlated with failure to assemble flagella. Phosphorylation of the Uni2 protein is reduced in uni1 mutant cells. Immunogold electron microscopy showed that the Uni2 protein localizes at the distal end of the basal body where microtubule transition occurs. These results provide the first mechanistic insights into the function of UNI1 and UNI2 genes in the pathway mediating assembly of doublet microtubules in the axoneme from triplet microtubules in the basal body template.

scholarly journals Basal body reorientation mediated by a Ca2+-modulated contractile protein.

1987 ◽  
Vol 105 (2) ◽  
pp. 903-912 ◽  
Author(s):  
G I McFadden ◽  
D Schulze ◽  
B Surek ◽  
J L Salisbury ◽  
M Melkonian
Keyword(s):  
Green Algae ◽  
Basal Body ◽  
Flagellar Apparatus ◽  
Immunoblot Analysis ◽  
Contractile Protein ◽  
Immunogold Electron Microscopy ◽  
Basal Bodies ◽  
Photophobic Response ◽  
Parallel Configuration

A rapid, Ca2+-dependent change in the angle between basal bodies (up to 180 degrees) is associated with light-induced reversal of swimming direction (the "photophobic" response) in a number of flagellated green algae. In isolated, detergent-extracted, reactivated flagellar apparatus complexes of Spermatozopsis similis, axonemal beat form conversion to the symmetrical/undulating flagellar pattern and basal body reorientation (from the antiparallel to the parallel configuration) are simultaneously induced at greater than or equal to 10(-7) M Ca2+. Basal body reorientation, however, is independent of flagellar beating since it is induced at greater than or equal to 10(-7) M Ca2+ when flagellar beating is inhibited (i.e., in the presence of 1 microM orthovanadate in reactivation solutions; in the absence of ATP or dithiothreitol in isolation and reactivation solutions), or when axonemes are mechanically removed from flagellar apparatuses. Although frequent axonemal beat form reversals were induced by varying the Ca2+ concentration, antiparallel basal body configuration could not be restored in isolated flagellar apparatuses. Observations of the photophobic response in vivo indicate that even though the flagella resume the asymmetric, breaststroke beat form 1-2 s after photostimulation, antiparallel basal body configuration is not restored until a few minutes later. Using an antibody generated against the 20-kD Ca2+-modulated contractile protein of striated flagellar roots of Tetraselmis striata (Salisbury, J. L., A. Baron, B. Surek, and M. Melkonian, 1984, J. Cell Biol., 99:962-970), we have found the distal connecting fiber of Spermatozopsis similis to be immunoreactive by indirect immunofluorescence and immunogold electron microscopy. Electrophoretic and immunoblot analysis indicates that the antigen of S. similis flagellar apparatuses consists, like the Tetraselmis protein, of two acidic isoforms of 20 kD. We conclude that the distal basal body connecting fiber is a contractile organelle and reorients basal bodies during the photophobic response in certain flagellated green algae.

Ciliary coordination in newt lung cells requires microtubule-based linkages between basal bodies: A correlative light and EM study

1992 ◽  
Vol 50 (1) ◽  
pp. 570-571
Author(s):  
Robert Hard ◽  
Gerald Rupp ◽  
Matthew L. Withiam-Leitch ◽  
Lisa Cardamone
Keyword(s):  
Basal Body ◽  
Untreated Control ◽  
Beat Frequency ◽  
Primary Cultures ◽  
Living Cells ◽  
Power Stroke ◽  
Ultrastructural Changes ◽  
Lung Cells ◽  
Basal Bodies ◽  
Ciliated Cells

In a coordinated field of beating cilia, the direction of the power stroke is correlated with the orientation of basal body appendages, called basal feet. In newt lung ciliated cells, adjacent basal feet are interconnected by cold-stable microtubules (basal MTs). In the present study, we investigate the hypothesis that these basal MTs stabilize ciliary distribution and alignment. To accomplish this, newt lung primary cultures were treated with the microtubule disrupting agent, Colcemid. In newt lung cultures, cilia normally disperse in a characteristic fashion as the mucociliary epithelium migrates from the tissue explant. Four arbitrary, but progressive stages of dispersion were defined and used to monitor this redistribution process. Ciliaiy beat frequency, coordination, and dispersion were assessed for 91 hrs in untreated (control) and treated cultures. When compared to controls, cilia dispersed more rapidly and ciliary coordination decreased markedly in cultures treated with Colcemid (2 mM). Correlative LM/EM was used to assess whether these effects of Colcemid were coupled to ultrastructural changes. Living cells were defined as having coordinated or uncoordinated cilia and then were processed for transmission EM.

Structural Analysis of Basal Bodies of The Isolated Oral Apparatus of Tetrahymena Pyriformis

1970 ◽  
Vol 6 (3) ◽  
pp. 679-700
Author(s):  
J. WOLFE
Keyword(s):  
Structural Analysis ◽  
Cell Membrane ◽  
Basal Body ◽  
Structural Integrity ◽  
Tetrahymena Pyriformis ◽  
Basal Plate ◽  
Basal Bodies ◽  
The Third ◽  
Ionic Detergent

The oral apparatus of Tetrahymena pyriformis was isolated using a non-ionic detergent to disrupt the cell membrane. The mouth consists largely of basal bodies and microfilaments. Each basal body is attached to the mouth by a basal plate which is integrated into the meshwork of microfilaments that confers upon the oral apparatus its structural integrity. Each basal body is composed of 9 triplet microtubules. Two of the 3 tubules, subfibres ‘A’ and ‘B’ are composed of filamentous rows of globules with a spacing of 4.5nm. The third tubule, subfibre ‘C’, is only one-third the length of the basal body.

Definition of individual components within the cytoskeleton of Trypanosoma brucei by a library of monoclonal antibodies

1989 ◽  
Vol 93 (3) ◽  
pp. 491-500 ◽  
Author(s):  
A. Woods ◽  
T. Sherwin ◽  
R. Sasse ◽  
T.H. MacRae ◽  
A.J. Baines ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Trypanosoma Brucei ◽  
Immunofluorescence Microscopy ◽  
Complex Mixtures ◽  
Immunogold Electron Microscopy ◽  
Basal Bodies ◽  
Tubulin Isotypes ◽  
Definition Of ◽  
Attachment Zone

The detergent-insoluble T. brucei cytoskeleton consists of several morphologically distinct regions and organelles, many of which are detectable only by electron microscopy. We have produced a set of monoclonal antibodies that define each structural component of this highly ordered cytoskeleton. The monoclonal antibodies were selected by cloning of hybridomas produced from mice injected with complex mixtures of proteins of either the cytoskeleton itself or salt extracts thereof. Four antibodies define particular tubulin isotypes and locate the microtubules of the axoneme and sub-pellicular array; two antibodies recognize the flagellum attachment zone; one recognizes the paraflagellar rod and another the basal bodies. Finally, one antibody defines a detergent-insoluble component of the nucleus. The antigens detected by each monoclonal antibody have been analysed by immunofluorescence microscopy, immunogold electron microscopy and Western blotting.

scholarly journals Fungi Burger from Stale Bread? A Case Study on Perceptions of a Novel Protein-Rich Food Product Made from an Edible Fungus

Foods ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1112 ◽  
Author(s):  
Coralie Hellwig ◽  
Rebecca Gmoser ◽  
Magnus Lundin ◽  
Mohammad J. Taherzadeh ◽  
Kamran Rousta
Keyword(s):  
Information Dissemination ◽  
Food Product ◽  
Environmental Benefits ◽  
Edible Fungus ◽  
Neurospora Intermedia ◽  
Rate Characteristic ◽  
And Gender ◽  
Protein Rich ◽  
Novel Protein

The current study aims to assess how a novel fungi product made from the filamentous fungus Neurospora intermedia, cultivated on bread residuals, is perceived using questionnaires. Participants were asked to rate characteristic attributes of a fungi burger patty and state their preference when comparing it to Quorn and hamburger patties. The data were analyzed to assess whether gender or age was statistically associated with preference profiles. Neither age nor gender was associated with the preference profiles regarding the comparison of burger patties. Except for age and bitterness, age and gender were also not associated with the preference profiles regarding the sensory characteristics of the fungi burger patty. Most of the participants liked the characteristics of the fungi burger patty. The results indicate that fungi products from waste can become accepted products when information dissemination targets environmental benefits. Moreover, to be commercially accepted, the chewiness and bitterness of the product should be improved. Other improvements should target the overall taste in order to cater to people who prefer meat-based protein sources.

scholarly journals The ultrastructure of cell division in Euglena gracilis

1978 ◽  
Vol 31 (1) ◽  
pp. 25-35
Author(s):  
M.A. Gillott ◽  
R.E. Triemer
Keyword(s):  
Cell Division ◽  
Nuclear Envelope ◽  
Basal Body ◽  
Euglena Gracilis ◽  
Equatorial Region ◽  
Basal Bodies ◽  
Cleavage Furrow ◽  
Free Zone ◽  
Prophase Nucleus

The ultrastructure of mitosis in Euglena gracilis was investigated. At preprophase the nucleus migrates anteriorly and associates with the basal bodies. Flagella and basal bodies replicate at preprophase. Cells retain motility throughout division. The reservoir and the prophase nucleus elongate perpendicular to the incipient cleavage furrow. One basal body pair surrounded by a ribosome-free zone is found at each of the nuclear poles. The spindle forms within the intact nuclear envelope- Polar fenestrae are absent. At metaphase, the endosome is elongated from pole to pole, and chromosomes are loosely arranged in the equatorial region. Distinct, trilayered kinetochores are present. Spindle elongates as chromosomes migrate to the poles forming a dumb-bell shaped nucleus by telophase. Daughter nuclei are formed by constriction of the nuclear envelope. Cytokinesis is accomplished by furrowing. Cell division in Euglena is compared with that of certain other algae.

Utilization of proteins from AluProt-CGNA (a novel protein-rich lupin variety) in the development of oil-in-water multilayer emulsion systems

2015 ◽  
Vol 118 (7) ◽  
pp. 1104-1112 ◽  
Author(s):  
César Burgos-Díaz ◽  
Miguel Gallardo ◽  
Eduardo Morales ◽  
José A. Piornos ◽  
Ana M. Marqués ◽  
...  
Keyword(s):  
Oil In Water ◽  
Emulsion Systems ◽  
Protein Rich ◽  
Novel Protein

The distribution of the fiber cell intrinsic membrane proteins MP20 and connexin46 in the bovine lens

1992 ◽  
Vol 103 (1) ◽  
pp. 245-257 ◽  
Author(s):  
E. Tenbroek ◽  
M. Arneson ◽  
L. Jarvis ◽  
C. Louis
Keyword(s):  
Monoclonal Antibody ◽  
Fiber Cell ◽  
Immunogold Electron Microscopy ◽  
Lens Fiber ◽  
Lens Fiber Cell ◽  
Western Blots ◽  
Fiber Cells ◽  
Bovine Lens ◽  
Narrow Side ◽  
Lens Membranes

MP20 is an intrinsic membrane protein previously identified in mammalian lens fiber cells. To identify a possible role for this protein in the lens, the distribution of MP20 and connexin46 has now been examined. Western immunoblotting with an anti-peptide antibody generated to the C-terminal 8 amino acids of MP20 confirmed the presence of this protein in the lens of several different mammalian species. A monoclonal antibody 5H1 was prepared that, in Western blots of bovine lesn membranes, recognized the same component as an antibody to rat connexin46 (Cx46). The apparent molecular mass of this component decreased from 59 kDa to 55 kDa following treatment of lens membranes with alkaline phosphatase. A monoclonal antibody to connexin-related MP70 recognized a 70 kDa component in bovine lens membranes confirming the presence of these two different connexin proteins in bovine lens membranes. To localize MP20 and Cx46 in the bovine lens membrane, lens fiber cell bundles were immunofluorescently labeled with both the MP20 antibody, and the monoclonal antibody to Cx46. Cx46 was identified in large plaques on the broad faces of the lens fiber cells throughout the outer 1 mm of the lens cortex. MP20 colocalized with Cx46 only in a restricted area 0.5 mm to 1.0 mm into the lens. In other regions of the lens, MP20 appeared more diffusely distributed over the fiber cell surface, although apparently concentrated in the ball-and-socket regions at the corners of the narrow side of the inner cortical lens fiber cells. These inner cortical regions were devoid of Cx46. A difference in distribution of these two proteins was confirmed in studies of immunofluorescently labeled lens cryosections. Furthermore, immunogold electron microscopy of purified lens membranes identified MP20 in both junctional regions (with Cx46) and in single membranes. These results provide evidence for a role for MP20 in mammalian lens fiber cell junctional formation or organization.

Basal Body and Flagellar Development During the Vegetative Cell Cycle and the Sexual Cycle of Chlamydomonas Reinhardii

1974 ◽  
Vol 16 (3) ◽  
pp. 529-556 ◽  
Author(s):  
T. CAVALIER-SMITH
Keyword(s):  
Basal Body ◽  
Vegetative Cell ◽  
Amorphous Material ◽  
Sexual Cycle ◽  
Transitional Region ◽  
Basal Bodies ◽  
Vegetative Cells ◽  
Chlamydomonas Reinhardii ◽  
Body Development ◽  
Microtubular Roots

Basal body development and flagellar regression and growth in the unicellular green alga Chlamydomonas reinhardii were studied by light and electron microscopy during the vegetative cell cycle in synchronous cultures and during the sexual life cycle. Flagella regress by gradual shortening prior to vegetative cell division and also a few hours after cell fusion in the sexual cycle. In vegetative cells basal bodies remain attached to the plasma membrane by their transitional fibres and do not act as centrioles at the spindle poles during division. In zygotes the basal bodies and associated microtubular roots and cross-striated connexions all dissolve, and by 6.5 h after mating all traces of flagellar apparatus and associated structures have disappeared. They remain absent for 6 days throughout zygospore maturation and then are reassembled during zygospore germination, after meiosis has begun. Basal body assembly in developing zygospores occurs close to the plasma membrane (in the absence of pre-existing basal bodies) via an intermediate stage consisting of nine single A-tubules surrounding a central ‘cartwheel’. Assembly is similar in vegetative cells (and occurs prior to cell division), except that new basal bodies are physically attached to old ones by amorphous material. In vegetative cells, amorphous disks, which may possibly be still earlier stages in basal-body development occur in the same location as 9-singlet developing basal bodies. After the 9-singlet structure is formed, B and C fibres are added and the basal body elongates to its mature length. Microtubular roots, striated connexions and flagella are then assembled. Both flagellar regression and growth are gradual and sequential, the transitional region at the base of the flagellum being formed first and broken down last. The presence of amorphous material at the tip of the axoneme of growing and regressing flagella suggests that the axoneme grows or shortens by the sequential assembly or disassembly at its tip. In homogenized cells basal bodies remain firmly attached to each other by their striated connexions. The flagellar transitional region, and parts of the membrane and of the 4 microtubular roots, also remain attached; so also do new developing basal bodies, if present. These structures are well preserved in homogenates and new fine-structural details can be seen. These results are discussed, and lend no support to the idea that basal bodies have genetic continuity. It is suggested that basal body development can be best understood if a distinction is made between the information needed to specify the structure of a basal body and that needed to specify its location and orientation.

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