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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    The journal of membrane biology 52 (1980), S. 147-159 
    ISSN: 1432-1424
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary MDCK cells form monolayers which have many of the properties usually found in transporting epithelia. The present article is devoted to the study of the structural and functional polarization of MDCK cells, which is one of the central features of transporting epithelia. The results show: (i) that MDCK monolayers transport 2.6 μmol hr−1 cm−2 of sodium in the apical to basolateral direction; (ii) the passive flux of this ion is relatively large (20.3 mole hr−1 cm−2), which is a characteristic of leaky epithelia; (iii) a large fraction of the penetration of sodium into the cells proceeds through an amiloride-sensitive channel, and the exit is operated mainly by a ouabain-sensitive pump; (iv) the net transport of sodium from the apical to the basolateral side agrees with the asymmetric labeling of the pumps with3H-ouabain; (v) this asymmetric labeling agrees, in turn, with a higher concentration of intramembrane particles (IMPs) in freeze-fracture replicas of the basolateral side of the plasma membrane; (vi) the structural polarization of confluent MDCK cells is also revealed by the location of microvilli, occluding junctions, and pinocytotic vesicles; and (vii) the presence of a continuous ring formed by actin microfilaments visualized by immunofluorescence under the lateral aspect of the plasma membrane that may be related to the distribution of the occluding junctions, which act as barriers separating apical from basolateral membrane components.
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  • 2
    Electronic Resource
    Electronic Resource
    [s.l.] : Nature Publishing Group
    Nature 248 (1974), S. 486-490 
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] The mRNA molecules from Xenopus laevis embryos represent polyribonucleotide chains of heterogeneous size transcribed from unique DNA sequences, each carrying a poly (A) stretch at the 3′ terminus and at the 5′ terminus, a fragment of 50 to 60 nucleotides transcribed from repeated DNA ...
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  • 3
    Electronic Resource
    Electronic Resource
    [s.l.] : Nature Publishing Group
    Nature 217 (1968), S. 179-180 
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] The technique involved placing the lens regenerates on agar cubes into which culture medium had been incorporated. The two media used were medium 199 with 15 per cent foetal bovine serum or Puck's N-16 (Microbiological Associates, Bethesda, Maryland). Penicillin was added at a concentration of 200 ...
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  • 4
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    The @journal of eukaryotic microbiology 41 (1994), S. 0 
    ISSN: 1550-7408
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: . Large percentages of Toxoplasma gondii tachyzoites could be induced to display two types of movement associated with active invasive behavior by exposing them for 1 min to 0.002% trypsin in phosphate-buffered saline (PBS). The motile activity, consisting of clockwise rotation around the posterior end (about 20 revolutions per min) and twirling-gliding over a poly-L-lysine substrate (1.2 ± 0.2 μm/s standard deviation), was observed and recorded by video-enhanced contrast microscopy. The number of active tachyzoites reached a maximum 1 min after trypsinization; the motile response of the population lasted for about 5 min. Activation was prevented by soybean trypsin-inhibitor, and could not be induced again in previously treated specimens. Electron-microscopy of trypsinized tachyzoites fixed in the presence of ruthenium-red revealed discrete discontinuities of the plasma membrane, which sealed within 90 min after washing with PBS. Treated tachyzoites were able to invade cultured epithelial cells with a higher relative infectivity than that of untreated parasites. Perfusion of trypsinized tachyzoites with 1 mM of either CaCl2 or MgCl2 and 1 mM ATP increased the number of activated parasites to over 60%; on the other hand, all induced motility was inhibited or blocked by agents that chelate divalent cations. The present preparation, which provided the first serial illustrations of T. gondii movements induced by a defined chemical stimulus, may offer a useful experimental model for the study of motility in this parasite.
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  • 5
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    The @journal of eukaryotic microbiology 41 (1994), S. 0 
    ISSN: 1550-7408
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: . Parasitic amebas propagate among hosts through cysts, the resistant forms in their life cycle. In spite of their key role in infection, little is known about the encystation process and the mechanisms involved in reaching this stage. Two features drastically affected by encystation are motility and cell shape, both of which are determined by the cytoskeleton, composed mainly of actin in these organisms. Therefore, we studied the occurrence and relative levels of actin and actin synthesis during encystation of Entamoeba invadens. Using a cDNA actin probe obtained from a library of E. histolytica and a monoclonal antibody against actin, we found that, while the total actin levels sharply decrease as encystation proceeds, the levels of actin mRNA are reduced only in mature cysts. Moreover, actin synthesis does not take place in precysts and the later stages of cyst formation. In contrast, the levels of other proteins remain stable in trophozoites, precysts and cysts, and stage specific peptides are actively synthesized in precysts. The results indicate that encystation is accompanied by a preferential down-regulation of actin synthesis and a decrease in actin levels. The reorganization of the cytoskeleton occurring as trophozoites transform into round, quiescent cells, could be a regulatory factor in the observed changes.
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  • 6
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    The @journal of eukaryotic microbiology 44 (1997), S. 0 
    ISSN: 1550-7408
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: . Actin mRNA levels were measured in Entamoeba histolytica trophozoites after experimentally inducing changes in the organization of the cytoskeleton. The treatment of trophozoites with forskolin, N6,2′-O-dibutyryl-adenosine 3′:5′-cyclic monophosphate, and phorbol myristate acetate induced the organization of actin into multiple dots and defined structures with a concomitant increase in F-actin content. Cytochalasin D elicited polarization of the structured actin and formation of aggregates, as well as an increment in F-actin. Simultaneously, up-regulation of actin mRNA levels was produced by all the drugs. De novo synthesis of actin mRNA, as measured by nuclear run-ons, showed increased transcription of actin mRNA. On the other hand, treatment of cells with actinomycin D blocked the elevation of actin mRNA synthesis induced by forskolin, dibutyryl cyclic AMP, or cytochalasin D whereas, the increment induced by PMA was not affected. These data indicate a regulatory control of actin mRNA synthesis at the transcriptional level by forskolin, dibutyryl cyclic AMP and cytochalasin D, and transcriptional as well as post-transcriptional controls by phorbol myristate acetate. The experiments presented here suggest the possibility that, regulation of actin mRNA transcription in E. histolytica trophozoites is linked to growth conditions, that are accompanied by reorganization of the actin cytoskeleton and thus, related to the motility and invasiveness of the parasite.
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  • 7
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    The @journal of eukaryotic microbiology 36 (1989), S. 0 
    ISSN: 1550-7408
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Characterization of a cytochalasin D-resistant mutant of the human parasite Entamoeba histolytica capable of growing at 10 μM cytochalasin is described. The mutant cells also show resistance to 5 mM colchicine and 100 μM cytochalasin B, drugs proved deleterious for wild type trophozoites. The mutants show increased osmotic fragility and electric mobility but reduced phagocytic activity, and agglutination by Concanavalin A. On the other hand pinocytic activity remains unaltered when compared with the wild type cells. Polymerized actin, seen by staining with phalloidin, often appears polarized to one end of the trophozoites and forms few of the endocytic invaginations found in wild type amebas. An altered distribution of part of the actin could explain the differences in surface properties and motility observed in the mutant amebas.
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  • 8
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 12 (1994), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Ribosomal DNA sequences in several species of the genus Entamoeba are highly repeated and display restriction fragment-length polymorphism (RFLP), which has been used to identify species and differentiate strains. However, the continuous variability of the non-transcribed repeat sequences in the ribosomal episome hinders an accurate typification. Looking for more reliable markers, we used DNA probes containing conserved sequences in the ribosomal episome — coding regions for the 16S and 5.8S rRNAs and transcribed spacers flanking the rDNA sequences, and the coding region for the 3 end of the 26S rRNA — to analyse hybridization patterns from five cloned pathogenic strains of Entamoeba histoiytica, two strains of the also pathogenic Entamoeba invadens and the non-pathogenic Laredo strain of Entamoeba moshkovskii. Our results provide reliable bases for the differentation of clones, strains and species of Entamoeba and the reconstruction of E. histolytica episomes. Differences in the number and length of rDNA-containing DNA fragments, previously observed by other investigators and confirmed by us, can be better defined by the present analysis.
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  • 9
    ISSN: 1573-2657
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Cultured MDCK cell monolayers respond to a low level of extracellular calcium ([Ca2+]e≤5μm) with a loss of transepithelial electrical resistance and transport function, and changes in position of a circumferential ring of actin filaments tethered to the plasma membrane at the zonula adhaerens. Keeping this cytoskeletal structure in place seems necessary to preserve the architecture of the tight junctions and therefore their sealing capacity. All three effects are reversible upon restituting normal [Ca2+]e. Recent work provided evidence of actin–myosin interactions at the filament ring, thus suggesting a contraction process involved in the alteration of the actin cytoskeleton. We now report that active contraction does occur and causes an extensive morphological transformation of MDCK cells. A marked increase in cell height simultaneous with a decrease in width and area of contact to the substratum was seen within 10min of removal of [Ca2+]e; recovery began immediately after replacing calcium, although it took longer for completion. Conventional and confocal epifluorescence studies showed actin colocalized with myosin II at various planes of resting or contracted cells, in particular at the ring level. Electron-micrographs revealed the circumferential actin ring associated with the plasma membrane in a waist-like constriction when Ca2+ was removed from the cultures. Contraction, as well as relaxation, in response to [Ca2+]e variations were inhibited by cytochalasin-D (an actin-filament disrupting drug), by okadaic acid (an inhibitor of myosin light-chain dephosphorylation), and by 2,3-butanedione monoxime (a blocker of myosin II ATPase activity). Similarly, no response was observed in cells previously depleted of metabolic energy by 2,4-dinitrophenol and 2-deoxy-D-glucose preincubation. The actin–myosin mediated reversible structural transformation of MDCK cells in response to [Ca2+]e poses new questions for the interpretation of in vitro experiments, as well as for the understanding of epithelial function.
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  • 10
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 32 (1995), S. 37-45 
    ISSN: 0886-1544
    Keywords: Entamoeba histolytica ; adhesion plates ; cytoskeleton ; fibronectin binding and degradation ; signaling ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Entamoeba histolytica trophozoites are pleiomorphic and highly motile cells. Although scarce fibrous material can be identified in the cytoplasm as elements of an organized cytoskeleton, clearly defined actin-containing structures are formed at the sites of cell-matrix contact upon the interaction of trophozoites with fibronectin (FN) and other extracellular matrix substrates. These structures are reminiscent of the adhesion plaques or focal contacts found in higher eukaryotic cells, where actin filament bundles insert into specialized regions of the plasma membrane and function as signal transduction organelles. Thus, the formation of adhesion plates in this parasitic ameba could be related to specific signaling responses involved in its invasive behavior. Here, we report the isolation of amebic adhesion plates and the results of their structural and molecular analyses. Filaments, with the characteristic diameter of F-actin, radiating from an electrondense matrix, are the main feature. Actin is one of the main protein components of the plate; other proteins identified are a FN-binding protein - previously reported as a “putative” FN receptor - the actin-binding proteins myosin II, myosin I, α-actinin, vinculin, and tropomyosin. The presence in the isolated plates of several proteases and protein kinases, in particular pp125FAK, is also demonstrated. Our results suggest that adhesion plates in amebas are dynamic membrane-cytoskeletal complexes participating not only in the attachment to FN substrates but also providing the structural basis for their involvement in parasite locomotion and invasiveness. © 1995 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
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