Actin in <i>Tetrahymena paravorax</i>: Ultrastructural Localization of HMM‐Binding Filaments in Glycerinated Cells1,2

Méténier, Guy

doi:10.1111/j.1550-7408.1984.tb02950.x

Cited by 26 publications

(9 citation statements)

References 33 publications

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“…While the bulk of this filament system has been established as centrin (Beisson et al 2001), this does not necessarily preclude association of centrin filaments with actin, as we can show. Recall that widely different affinity stains for actin, including heavy meromyosin, have resulted in cortical labeling in Paramecium (Tiggemann and Plattner 1981;Kersken et al 1986a,b), as well as in Tetrahymena (Méténier 1984). Theoretically, previous LM and preembedding-EM localization studies could have faced the problem of sol- Figure 5 Postembedding immunogold labeling in the cell cortex.…”

Section: Background From Previous Workmentioning

confidence: 99%

Immunolocalization of Actin in Paramecium Cells

Kissmehl

Sehring

Wagner

et al. 2004

J Histochem Cytochem.

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S U M M A R YWe have selected a conserved immunogenic region from several actin genes of Paramecium , recently cloned in our laboratory, to prepare antibodies for Western blots and immunolocalization. According to cell fractionation analysis, most actin is structurebound. Immunofluorescence shows signal enriched in the cell cortex, notably around ciliary basal bodies (identified by anti-centrin antibodies), as well as around the oral cavity, at the cytoproct and in association with vacuoles (phagosomes) up to several m in size. Subtle strands run throughout the cell body. Postembedding immunogold labeling/EM analysis shows that actin in the cell cortex emanates, together with the infraciliary lattice, from basal bodies to around trichocyst tips. Label was also enriched around vacuoles and vesicles of different size including "discoidal" vesicles that serve the formation of new phagosomes. By all methods used, we show actin in cilia. Although none of the structurally welldefined filament systems in Paramecium are exclusively formed by actin, actin does display some ordered, though not very conspicuous, arrays throughout the cell. F-actin may somehow serve vesicle trafficking and as a cytoplasmic scaffold. This is particularly supported by the postembedding/EM labeling analysis we used, which would hardly allow for any largescale redistribution during preparation. A ctin is a highly flexible cytoskeletal component that participates in many static and dynamic functions in eukaryotic cells (Pollard et al. 2000). This includes reversible self-assembly of monomeric G-actin to F-actin filaments. Also generally known is that these filaments may be more or less bundled and can serve different functions, such as structural enforcement and restructuring of the cell cortex, rearrangement of cortical components during intracellular signaling, organelle dynamics and transport, etc. The latter includes wellestablished functions such as phagosome formation and plasma streaming, i.e., cyclosis (Shimmen and Yokota 2004). However, quite recent results highlight a much broader functional spectrum of F-actin than previously assumed. This applies to early steps of exocytosis, including dense core vesicle docking (Morales et al. 2000;Pendleton and Koffer 2001;Manneville et al. 2003;Gasman et al. 2004), late steps of endocytosis (Engqvist-Goldstein and Drubin 2003;Guilherme et al. 2004), exo-endocytosis coupling (Valentijn et al. 1999), endo-phagosome interaction (Kjeken et al. 2004), delivery of endocytosed receptors to lysosomes for degradation (Stoorvogel et al. 2004), vacuole fusion in yeast (Merz and Wickner 2004), and positioning of the nucleus (Starr and Han 2003). Some aspects are still poorly understood, particularly, e.g., the role of actin in flagella of algae (Mitchell 2000;Hayashi et al. 2001;Hirono et al. 2003), whereas its occurrence in cilia has remained a matter of debate. Another line of experiments concerns the potential role of actin in mediating the connection between cortical Ca 2 ϩ -stores and the plasma membrane...

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Section: Background From Previous Workmentioning

confidence: 99%

Immunolocalization of Actin in Paramecium Cells

Kissmehl

Sehring

Wagner

et al. 2004

J Histochem Cytochem.

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“…These sites include the oral apparatus (8,21,29,41,45), phagosomes (food vacuoles) (8, 31), the cytoproct (8, 28), basal bodies (29,40,41,64), and cilia (41, 46, 64) of both Tetrahymena (reviewed in reference 15) and Paramecium, and the division furrow (22, 28) of Tetrahymena. However, despite the numerous places where actin has been found, its role in the life of these cells is incompletely understood.…”

mentioning

confidence: 99%

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

et al. 2006

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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 ACT1⌬ 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 ACT1⌬ 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 ACT1⌬ 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.Actin, although not abundant (26), has been detected at numerous intracellular sites within ciliates. These sites include the oral apparatus (8,21,29,41,45), phagosomes (food vacuoles) (8, 31), the cytoproct (8, 28), basal bodies (29,40,41,64), and cilia (41, 46, 64) of both Tetrahymena (reviewed in reference 15) and Paramecium, and the division furrow (22, 28) of Tetrahymena. However, despite the numerous places where actin has been found, its role in the life of these cells is incompletely understood.Perhaps the best-established function of ciliate actin is associated with phagocytosis, as cytochalasins and other drugs that affect actin polymerization (9) inhibit formation of food vacuoles in Tetrahymena pyriformis (49, 50), T. vorax (23), T. thermophila (72), and Paramecium tetraurelia (8,14). Cytochalasin B also binds to isolated oral apparatuses of T. thermophila (21), and inhibits the formation of this organelle, but does so at a much higher concentration than is sufficient to inhibit feeding (20). A recent study also has shown that latrunculin, an inhibitor of actin polymerization, interferes with the posteriorly directed movement of phagosomes within this cell (31).From past work it is unclear whether perturbing actin function interferes with cell division. Cytochalasin B did not prevent cell division of...

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“…The detached vesicle fuses with lysosome membranes to form a mature food vacuole (Kitajima and Thompson 1977; Rasmussen 1976). Although actin has been localized to the food vacuole‐forming region of the Tetrahymena oral apparatus (Méténier 1984), the precise involvement of actin in this process remains obscure. In the present study, both the treatment of cells with cytochalasin and the disruption of MYO1 transcription resulted in inhibition of food vacuole formation, an indication that Myolp‐actin interaction is required for endocytosis in Tetrahymena.…”

Section: Discussionmentioning

confidence: 99%

MYO1, a Novel, Unconventional Myosin Gene Affects Endocytosis and Macronuclear Elongation inTetrahymena thermophila

Williams

Hosein

Garcés

et al. 2000

J Eukaryotic Microbiology

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Targeted gene disruption was used to investigate the function of MYO1, an unconventional myosin gene in Tetrahymena thermophila. Phenotypic analysis of a transformed strain that lacked a functional MYO1 gene was conducted at both 20 degrees C and 35 degrees C. At either temperature the delta MYO1 strain had a smaller cytoplasm/nucleus ratio than wild type. At 20 degrees C, delta MYO1 populations had a longer doubling time than wild type, lower saturation density, and a reduced rate of food vacuole formation. However, at 35 degrees C, these characteristics were comparable to wild type. Although micronuclear division and cytokinesis appeared normal in delta MYO1 cells, failure of the macronucleus to elongate properly resulted in unequal segregation of macronuclear DNA in cells maintained at either 20 degrees C or 35 degrees C.

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Actin in Tetrahymena paravorax: Ultrastructural Localization of HMM‐Binding Filaments in Glycerinated Cells1,2

Cited by 26 publications

References 33 publications

Immunolocalization of Actin in Paramecium Cells

Immunolocalization of Actin in Paramecium Cells

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

MYO1, a Novel, Unconventional Myosin Gene Affects Endocytosis and Macronuclear Elongation inTetrahymena thermophila

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