scholarly journals Microtubule glycylation promotes basal body attachment to the cell cortex

2019 ◽  
Author(s):  
Anthony D. Junker ◽  
Adam W. J. Soh ◽  
Eileen T. O’Toole ◽  
Janet B. Meehl ◽  
Mayukh Guha ◽  
...  
Keyword(s):  
Light Microscopy ◽  
Tetrahymena Thermophila ◽  
Hydrodynamic Flow ◽  
Cell Cortex ◽  
Basal Bodies ◽  
Post Translational Modifications ◽  
Summary Statement ◽  
Motile Cilia ◽  
Em Tomography ◽  
Cortical Cytoskeleton

ABSTRACTMotile cilia generate directed hydrodynamic flow that is important for the motility of cells and extracellular fluids. To optimize directed hydrodynamic flow, motile cilia are organized and oriented into a polarized array. Basal bodies (BB) nucleate and position motile cilia at the cell cortex. Cytoplasmic BB-associated microtubules are conserved structures that extend from BBs. Using the ciliate, Tetrahymena thermophila, combined with EM-tomography and light microscopy, we show that BB-appendage microtubules assemble coincident with new BB assembly and are attached to the cell cortex. These BB-appendage microtubules are specifically marked with post translational modifications of tubulin, including glycylation. Mutations that prevent glycylation shorten BB-appendage microtubules and disrupt BB positioning and cortical attachment. Consistent with the attachment of BB-appendage microtubules to the cell cortex for BB positioning, mutations that disrupt the cellular cortical cytoskeleton similarly disrupt the cortical attachment and positioning of BBs. In summary, BB-appendage microtubules promote the organization of ciliary arrays through attachment to the cell cortex.SUMMARY STATEMENTBasal bodies position motile cilia at the cell cortex. This study finds tubulin glycylation to promote BB-associated microtubule elongation and structural attachment of basal bodies to the cell’s cortical cytoskeleton.

Mutational analysis of patterning of oral structures in Tetrahymena

Development ◽  
1984 ◽  
Vol 82 (1) ◽  
pp. 67-95
Author(s):  
Joseph Frankel ◽  
E. Marlo Nelsen ◽  
Julita Bakowska ◽  
Leslie M. Jenkins
Keyword(s):  
Mutational Analysis ◽  
Tetrahymena Thermophila ◽  
The State ◽  
The Other ◽  
Basal Bodies ◽  
Ciliated Protozoan ◽  
Vertebrate Limb ◽  
Spatial Configurations ◽  
Oral Development ◽  
Mutant Cells

The ciliary arrays of the oral apparatus of the ciliated protozoan Tetrahymena thermophila each have their own unique ‘pattern signature’, which varies little so long as the number of arrays remains the same. In this study, we analyse the consequence of increases in the number of these arrays (membranelles) brought about by certain mutations. In oral apparatuses of mutant cells, the addition of a membranelle is associated with specific alterations in at least one of the other membranelles. The features that are altered include the relative lengths of membranelles, the state of ciliation of basal bodies located at specific positions within these membranelles, and the spatial configurations resulting from displacement of ciliary units during late oral development. The final organization of each membranelle depends upon its relativeposition along the length of the oral apparatus. This indicates that the membranelles are not individually ‘named’ by the organism, and suggests that the unit of pattern organizationis the membranelle field as a whole. In the Discussion, we consider means for testing whether thesame underlying idea might also apply to multicellular systems, such as the vertebrate limb, in which spatially ordered differences appear to be superimposed upon a fundamental repeating pattern.

scholarly journals Asymmetrically localized proteins stabilize basal bodies against ciliary beating forces

2016 ◽  
Vol 215 (4) ◽  
pp. 457-466 ◽  
Author(s):  
Brian A. Bayless ◽  
Domenico F. Galati ◽  
Anthony D. Junker ◽  
Chelsea B. Backer ◽  
Jacek Gaertig ◽  
...  
Keyword(s):  
Basal Body ◽  
Mechanical Force ◽  
Basal Bodies ◽  
Mechanical Forces ◽  
Ciliary Beating ◽  
Motile Cilia ◽  
Molecular Components

Basal bodies are radially symmetric, microtubule-rich structures that nucleate and anchor motile cilia. Ciliary beating produces asymmetric mechanical forces that are resisted by basal bodies. To resist these forces, distinct regions within the basal body ultrastructure and the microtubules themselves must be stable. However, the molecular components that stabilize basal bodies remain poorly defined. Here, we determine that Fop1 functionally interacts with the established basal body stability components Bld10 and Poc1. We find that Fop1 and microtubule glutamylation incorporate into basal bodies at distinct stages of assembly, culminating in their asymmetric enrichment at specific triplet microtubule regions that are predicted to experience the greatest mechanical force from ciliary beating. Both Fop1 and microtubule glutamylation are required to stabilize basal bodies against ciliary beating forces. Our studies reveal that microtubule glutamylation and Bld10, Poc1, and Fop1 stabilize basal bodies against the forces produced by ciliary beating via distinct yet interdependent mechanisms.

scholarly journals The Two SAS-6 Homologs in Tetrahymena thermophila Have Distinct Functions in Basal Body Assembly

2009 ◽  
Vol 20 (6) ◽  
pp. 1865-1877 ◽  
Author(s):  
Brady P. Culver ◽  
Janet B. Meehl ◽  
Thomas H. Giddings ◽  
Mark Winey
Keyword(s):  
Basal Body ◽  
Structural Component ◽  
Tetrahymena Thermophila ◽  
Basal Bodies ◽  
Hub And Spoke ◽  
Structural Role ◽  
Higher Eukaryotes ◽  
Centriole Assembly ◽  
Cilia And Flagella

Cilia and flagella are structurally and functionally conserved organelles present in basal as well as higher eukaryotes. The assembly of cilia requires a microtubule based scaffold called a basal body. The ninefold symmetry characteristic of basal bodies and the structurally similar centriole is organized around a hub and spoke structure termed the cartwheel. To date, SAS-6 is one of the two clearly conserved components of the cartwheel. In some organisms, overexpression of SAS-6 causes the formation of supernumerary centrioles. We questioned whether the centriole assembly initiation capacity of SAS-6 is separate from or directly related to its structural role at the cartwheel. To address this question we used Tetrahymena thermophila, which expresses two SAS-6 homologues, TtSAS6a and TtSAS6b. Cells lacking either TtSAS6a or TtSAS6b are defective in new basal body assembly. TtSas6a localizes to all basal bodies equally, whereas TtSas6b is enriched at unciliated and assembling basal bodies. Interestingly, overexpression of TtSAS6b but not TtSAS6a, led to the assembly of clusters of new basal bodies in abnormal locations. Our data suggest a model where TtSAS6a and TtSAS6b have diverged such that TtSAS6a acts as a structural component of basal bodies, whereas TtSAS6b influences the location of new basal body assembly.

scholarly journals The Actin Gene ACT1 Is Required for Phagocytosis, Motility, and Cell Separation of Tetrahymena thermophila

2006 ◽  
Vol 5 (3) ◽  
pp. 555-567 ◽  
Author(s):  
Norman E. Williams ◽  
Che-Chia Tsao ◽  
Josephine Bowen ◽  
Gery L. Hehman ◽  
Ruth J. Williams ◽  
...  
Keyword(s):  
Cell Separation ◽  
Tetrahymena Thermophila ◽  
Cell Cortex ◽  
Actin Gene ◽  
Wild Type ◽  
Daughter Cells ◽  
Genetic Constructs ◽  
Mutant Cells ◽  
Phagocytic Uptake ◽  
Cell Cycle Length

ABSTRACT A previously identified Tetrahymena thermophila actin gene (C. G. Cupples and R. E. Pearlman, Proc. Natl. Acad. Sci. USA 83:5160-5164, 1986), here called ACT1, was disrupted by insertion of a neo3 cassette. Cells in which all expressed copies of this gene were disrupted exhibited intermittent and extremely slow motility and severely curtailed phagocytic uptake. Transformation of these cells with inducible genetic constructs that contained a normal ACT1 gene restored motility. Use of an epitope-tagged construct permitted visualization of Act1p in the isolated axonemes of these rescued cells. In ACT1Δ mutant cells, ultrastructural abnormalities of outer doublet microtubules were present in some of the axonemes. Nonetheless, these cells were still able to assemble cilia after deciliation. The nearly paralyzed ACT1Δ cells completed cleavage furrowing normally, but the presumptive daughter cells often failed to separate from one another and later became reintegrated. Clonal analysis revealed that the cell cycle length of the ACT1Δ cells was approximately double that of wild-type controls. Clones could nonetheless be maintained for up to 15 successive fissions, suggesting that the ACT1 gene is not essential for cell viability or growth. Examination of the cell cortex with monoclonal antibodies revealed that whereas elongation of ciliary rows and formation of oral structures were normal, the ciliary rows of reintegrated daughter cells became laterally displaced and sometimes rejoined indiscriminately across the former division furrow. We conclude that Act1p is required in Tetrahymena thermophila primarily for normal ciliary motility and for phagocytosis and secondarily for the final separation of daughter cells.

scholarly journals Sfr1, a Tetrahymena thermophila Sfi1 Repeat Protein, Modulates the Production of Cortical Row Basal Bodies

mSphere ◽  
2016 ◽  
Vol 1 (6) ◽  
Author(s):  
Westley Heydeck ◽  
Alexander J. Stemm-Wolf ◽  
Janin Knop ◽  
Christina C. Poh ◽  
Mark Winey
Keyword(s):  
Basal Body ◽  
Binding Proteins ◽  
Tetrahymena Thermophila ◽  
Binding Protein ◽  
Extensive Study ◽  
Basal Bodies ◽  
Content Type ◽  
Inhibitory Role ◽  
Body Production

ABSTRACT Basal bodies and centrioles are structurally similar and, when rendered dysfunctional as a result of improper assembly or maintenance, are associated with human diseases. Centrins are conserved and abundant components of both structures whose basal body and centriolar functions remain incompletely understood. Despite the extensive study of centrins in Tetrahymena thermophila, little is known about how centrin-binding proteins contribute to centrin’s roles in basal body assembly, stability, and orientation. The sole previous study of the large centrin-binding protein family in Tetrahymena revealed a role for Sfr13 in the stabilization and separation of basal bodies. In this study, we found that Sfr1 localizes to all Tetrahymena basal bodies and complete genetic deletion of SFR1 leads to overproduction of basal bodies. The uncovered inhibitory role of Sfr1 in basal body production suggests that centrin-binding proteins, as well as centrins, may influence basal body number both positively and negatively. Basal bodies are essential microtubule-based structures that template, anchor, and orient cilia at the cell surface. Cilia act primarily in the generation of directional fluid flow and sensory reception, both of which are utilized for a broad spectrum of cellular processes. Although basal bodies contribute to vital cell functions, the molecular contributors of their assembly and maintenance are poorly understood. Previous studies of the ciliate Tetrahymena thermophila revealed important roles for two centrin family members in basal body assembly, separation of new basal bodies, and stability. Here, we characterize the basal body function of a centrin-binding protein, Sfr1, in Tetrahymena. Sfr1 is part of a large family of 13 proteins in Tetrahymena that contain Sfi1 repeats (SFRs), a motif originally identified in Saccharomyces cerevisiae Sfi1 that binds centrin. Sfr1 is the only SFR protein in Tetrahymena that localizes to all cortical row and oral apparatus basal bodies. In addition, Sfr1 resides predominantly at the microtubule scaffold from the proximal cartwheel to the distal transition zone. Complete genomic knockout of SFR1 (sfr1Δ) causes a significant increase in both cortical row basal body density and the number of cortical rows, contributing to an overall overproduction of basal bodies. Reintroduction of Sfr1 into sfr1Δ mutant cells leads to a marked reduction of cortical row basal body density and the total number of cortical row basal bodies. Therefore, Sfr1 directly modulates cortical row basal body production. This study reveals an inhibitory role for Sfr1, and potentially centrins, in Tetrahymena basal body production. IMPORTANCE Basal bodies and centrioles are structurally similar and, when rendered dysfunctional as a result of improper assembly or maintenance, are associated with human diseases. Centrins are conserved and abundant components of both structures whose basal body and centriolar functions remain incompletely understood. Despite the extensive study of centrins in Tetrahymena thermophila, little is known about how centrin-binding proteins contribute to centrin’s roles in basal body assembly, stability, and orientation. The sole previous study of the large centrin-binding protein family in Tetrahymena revealed a role for Sfr13 in the stabilization and separation of basal bodies. In this study, we found that Sfr1 localizes to all Tetrahymena basal bodies and complete genetic deletion of SFR1 leads to overproduction of basal bodies. The uncovered inhibitory role of Sfr1 in basal body production suggests that centrin-binding proteins, as well as centrins, may influence basal body number both positively and negatively.

Evidence for the presence of DNA at basal body sites in Tetrahymena pyriformis

1965 ◽  
Vol 162 (989) ◽  
pp. 473-491 ◽  
Keyword(s):  
Cell Division ◽  
Basal Body ◽  
Mitotic Cycle ◽  
Tritiated Thymidine ◽  
Tetrahymena Pyriformis ◽  
Cell Cortex ◽  
Basal Bodies ◽  
Temperature Shock ◽  
Large Numbers ◽  
Order Of Magnitude

The reasons that have led to a search for DNA in the basal body of Tetrahymena pyriformis are twofold: the well-known property of proliferation of this organelle and the possibility that basal body DNA might be involved in its morphogenesis. After a brief review of earlier work the methods employed in this paper are described. To ensure large numbers of cells in a particular state of development organisms were grown in synchronized culture. Animals required for autoradiographic studies were appropriately treated with tritiated thymidine. All investigations were made on the cell cortex or 'ghost’ in order to avoid confusion from cell contents. In addition to autoradiography of ghosts, tests were made with acridine orange in the fluorescence microscope. It is concluded from fluorescence tests that basal bodies of T. pyriformis strain S contain DNA . This DNA is not detectable for the first 2h of the temperature-shock cycle, but is detect­able thereafter until cell division. The presence of DNA is confirmed by the autoradiography experiments. The amount of DNA per basal body is estimated very roughly in order of magnitude as 2 × 10 -16 g. The origin of basal body DNA is discussed and the possibilities and consequences of the existence of DNA in the homologous centriole are examined in terms of the mitotic cycle, the amoeba-flagellate transformation in Naegleria , and artificial parthenogenesis. The paper concludes with a brief discussion of the genetic implications of basal body DNA .

scholarly journals Mcidas mutant mice reveal a two-step process for the specification and differentiation of multiciliated cells in mammals

2018 ◽  
Author(s):  
Hao Lu ◽  
Priyanka Anujan ◽  
Feng Zhou ◽  
Yiliu Zhang ◽  
Yan Ling Chong ◽  
...  
Keyword(s):  
Basal Body ◽  
Target Genes ◽  
Initial Step ◽  
Mutant Mice ◽  
Respiratory Disorder ◽  
Basal Bodies ◽  
Cell Cycle Regulators ◽  
Multiciliated Cells ◽  
Motile Cilia ◽  
Body Production

ABSTRACTMotile cilia on multiciliated cells (MCCs) function in fluid clearance over epithelia. Studies with Xenopus embryos and patients with the congenital respiratory disorder reduced generation of multiple motile cilia, have implicated the nuclear protein MCIDAS (MCI), in the transcriptional regulation of MCC specification and differentiation. Recently, a paralogous protein, GMNC, was also shown to be required for MCC formation. Surprisingly, and in contrast to the presently held view, we find that Mci mutant mice can specify MCC precursors. However, these precursors cannot produce multiple basal bodies, and mature into single ciliated cells. We show that MCI is required specifically to induce deuterosome pathway components for the production of multiple basal bodies. Moreover, GMNC and MCI associate differentially with the cell-cycle regulators E2F4 and E2F5, which enables them to activate distinct sets of target genes (ciliary transcription factor genes versus genes for basal body generation). Our data establish a previously unrecognized two-step model for MCC development: GMNC functions in the initial step for MCC precursor specification. GMNC induces Mci expression, which then drives the second step of basal body production for multiciliation.SUMMARY STATEMENTWe show how two GEMININ family proteins function in mammalian multiciliated cell development: GMNC regulates precursor specification and MCIDAS induces multiple basal body formation for multiciliation.

scholarly journals Characterization of ASF1 and GCN5 in the ciliated protozoan tetrahymena thermophila

2021 ◽  
Author(s):  
Abdel Rahman Karsou
Keyword(s):  
Tetrahymena Thermophila ◽  
Histone H3 ◽  
Model Organism ◽  
Human Cells ◽  
Ciliated Protozoan ◽  
Yeast Saccharomyces Cerevisiae ◽  
Post Translational Modifications ◽  
Lysine Residues ◽  
Acetyl Group ◽  
H3 Acetylation

One method of regulating accessibility of DNA is chromatin remodelling via histone post-translational modifications (PTM). Adding an acetyl group to the lysine residues (K) on the core histone H3 is one of these chemical modifications. Acetylation of H3 on lysine 56 (H3K56ac) is an important histone alteration that is conserved among most if not all eukaryotes including humans. Several histone acetyl transferases (HAT) have been shown to be responsible for H3K56ac in different organisms including Gen5 and p300/CPB in human cells and Rtt109 in fungi including the yeast Saccharomyces cerevisiae. In addition the histone chaperone ASf1, is also required for these modifications in yeast and human cells. The ciliated protozoan Tetrahymena thermophila is an effective model organism for studying the function of histone PTMs in certain processes including meiosis and RNA interference. Here, I show that tGen5 has H3 acetylation activity and that tAsf1 binds Histone H3.

scholarly journals Characterization of ASF1 and GCN5 in the ciliated protozoan tetrahymena thermophila

2021 ◽  
Author(s):  
Abdel Rahman Karsou
Keyword(s):  
Tetrahymena Thermophila ◽  
Histone H3 ◽  
Model Organism ◽  
Human Cells ◽  
Ciliated Protozoan ◽  
Yeast Saccharomyces Cerevisiae ◽  
Post Translational Modifications ◽  
Lysine Residues ◽  
Acetyl Group ◽  
H3 Acetylation

One method of regulating accessibility of DNA is chromatin remodelling via histone post-translational modifications (PTM). Adding an acetyl group to the lysine residues (K) on the core histone H3 is one of these chemical modifications. Acetylation of H3 on lysine 56 (H3K56ac) is an important histone alteration that is conserved among most if not all eukaryotes including humans. Several histone acetyl transferases (HAT) have been shown to be responsible for H3K56ac in different organisms including Gen5 and p300/CPB in human cells and Rtt109 in fungi including the yeast Saccharomyces cerevisiae. In addition the histone chaperone ASf1, is also required for these modifications in yeast and human cells. The ciliated protozoan Tetrahymena thermophila is an effective model organism for studying the function of histone PTMs in certain processes including meiosis and RNA interference. Here, I show that tGen5 has H3 acetylation activity and that tAsf1 binds Histone H3.

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