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A K Fok

Publications and source records attributed to A K Fok.

45 records · Page 3Linked to original sources
PubMed1981

Axenic Paramecium caudatum. II. Changes in fine structure with culture age.

The fine structure of axenically grown Paramecium caudatum undergoes characteristic changes which reflect the cell's nutritional and metabolic conditions. Cells in early log phase of growth contain large lipid accumulations and extensive foci of rough ER bordered by Golgi stacks, indicating an active metabolism. Lipid bodies and ER foci are reduced during mid log phase. By days 5 and 6 (late log phase) some rough ER and a few lipid bodies are found in the cortex while the endoplasm is filled with free ribosomes and small vesicles. Conjugating cells, observed at the onset of stationary phase (day 6), show an accumulation of secondary lysosomes concomitant with a lack of digestive vacuoles. Lipofuscin-like granules and autophagosomes appear on day 7 and increase in number throughout the stationary and death phases. Immature autophagosomes are smaller and are bounded by a "thin" membrane lacking a glycocalyx. Mature autophagosomes (autophagolysosomes) containing partially digested mitochondria have a "thicker" membrane with a glycocalyx, a characteristic of lysosomes. Fusion of lysosomes with autophagosomes is verified by the presence of the natural lysosomal marker, the paracrystalline sheets, in the autophagolysosomes. The residue remaining from the lipid fields on days 5 to 20 includes highly convoluted membrane-like patches with long non-membrane-bound cylinders. Concomitant with the decrease in the number of digestive vacuoles, lysosomes, autophagosomes and lipofuscin-like granules proliferate from day 7 and become the striking feature of death-phase cells. With increased culture age the cisternae of Golgi stacks become dilated and decrease in diameter, and the number of ribosomes on the ER at the cis-face of these stacks is reduced. Mitochondria sometimes increase in density, and some have multi-lamellar whorls extending from their outer membrane.

Cell Survival↗
PubMed1980

Membrane recycling and endocytosis in Paramecium confirmed by horseradish peroxidase pulse-chase studies.

Paramecium caudatum cells were pulse-chased in horseradish peroxidase (HRP) using pulses of 30 s to follow endosome (endocytotic vesicle) formation from defaecating digestive vacuoles and 3-min pulses to follow the movement and fate of these vesicles in the cell. Endosomes formed during the 30 s in HRP are heavily labelled with HRP reaction product and are mostly flattened. Some align along microtubules that point toward the cytopharynx. Diskoidal vesicles at the cytopharynx are unlabelled in unchased cells. Cells exposed to a 3-min HRP pulse contain varied amounts of HRP-labelled diskoidal vesicles at the cytopharynx but no labelled vesicles near their closed cytoprocts. Labelled vesicles are also found aligned along the cytopharyngeal microtubular ribbons. Diskoidal vesicles are no longer labelled after a 47-min chase. Use of pulse-chase HRP cytochemistry supports the hypothesis that the membrane of digestive vacuoles retrieved at the cytoproct is moved along microtubular ribbons directly to the cytopharynx where the membrane enters the diskoidal vesicle pool. This membrane appears to enter neither the Golgi nor lysosomal systems in its passage. HRP reaction product is also found in vesicles near the parasomal sacs, in some secondary lysosomes and in small spherical vesicles that are probably trichocyst membrane fragments. Possibly some membrane from parasomal sacs or condensing digestive vacuoles may also enter the diskoidal vesicle pool but apparently not membrane from trichocysts.

Animals↗
PubMed1979

Axenic Paramecium caudatum. I. Mass culture and structure.

To establish and grow Paramecium caudatum in mass axenic culture the culture medium of Soldo, Godoy & van Wagtendonk was modified by substituting phosphatidylethanolamine (PE) for TEM-4T and by a 10-fold increase in folic acid. Population densities of 4000 to 6000 cells/ml and a generation time of 20-26 h are regularly obtained. Optimal growth is obtained with PE-stigmasterol ratios between 40:1 to 400:1. Cells from 1-day-old axenic cultures have many lipid bodies aggregated in clumps (which disappear in 2 to 3 days) as well as foci of rough endoplasmic reticulum bordered by dictyosomes. The latter suggests a very active metabolism. Crystalline sheets found in both food vacuoles and lysosomes presumably play a role in digestion. Axenically grown cells also have abundant Golgi bodies (dictyosomes) and by late log phase become filled with lysosomes.

Animals↗
PubMed1975

Cytochemical localization of peroxisomes in Tetrahymena pyriformis.

Pronounced staining of Tetrahymena pyriformis peroxisomes was demonstrated when glutaraldehyde-fixed cells were incubated in an alkaline medium containing 3,3'-diaminobenzidine and hydrogen peroxide. A variable amount of electron-opague deposit was observed when cells were incubated in diaminobenzidine and H2O2 for 1 hr while an intense deposit followed incubation for 4 hr in the same medium. The staining was abolished completely when 3-amino-1,2,4-triazole, KCN or NaN3 was added to the incubation medium. Based on these cytochemical observations and the morphologic identification by size, shape and other ultrastructural details, it is suggested that this study presents evidence for a conclusive morphologic identification of Tetrahymena peroxisomes.

Amitrole↗
PubMedDate not supplied

Nonclathrin vesicle coats and filament networks in the transition zone and trans-Golgi region of the Golgi complex of Paramecium.

Understanding vesicle trafficking to and through the Golgi stack has been greatly elucidated recently, but the question of what holds the endoplasmic reticulum (ER) and Golgi stack together in many cell types and an explanation of anterograde trafficking in the ER-Golgi transitional zone have not yet been adequately explained. We have studied these problems using both the thin sectioning and the quick-freeze deep-etch (QF-DE) technique on Paramecium cells harvested at different culture ages. Although the Golgi apparatus of Paramecium is made up of many sets of more reduced stacks of cisternae than those of many mammalian cells, the stacks in Paramecium always bear a close relationship to a transitional element of the ER from which non-clathrin-coated transition vesicles arise. In QF-DE replicas two networks of filaments are clearly shown; one is in this ER-Golgi transition zone and the other is on the trans side of the Golgi stack. The network associated with the trans-Golgi region links a number of vesicular elements. The network in the transition zone spans the distance between the ER and the cis-cisterna of the Golgi stack and has branches extending to the coats of the enmeshed nonclathrin-coated transition vesicles. These coats consist of a layer of 11-nm globular elements (the same size as coatomer complexes) which surround the 40-nm-diameter transition vesicles. We conclude that the filamentous network holds the ER and Golgi stack together and prevents the dispersal of the transition vesicles away from this zone. This network may also delineate and stabilize the transitional element within the ER and, finally, help organize anterograde transition vesicle trafficking in this ER-Golgi transition zone.

Animals↗
PubMedDate not supplied

Monoclonal antibody to a bacterial endonuclear symbiont Holospora cross reacts with proteins of contractile vacuole radial canals of Paramecium species.

A monoclonal antibody (mAb) IR-2-1 was raised against a 67-kDa protein purified from the macronucleus-specific bacterial symbiont Holospora obtusa of Paramecium caudatum. The mAb was found to react with two bands (31 and 67-kDa) on gels of H. obtusa. Indirect immunofluorescence microscopy showed that these antigens were distributed inside the cells. However, unexpectedly, this mAb also cross reacted with the radial arms of the contractile vacuole in P. caudatum, P. tetraurelia, P. multimicronucleatum, P. jenningsi and P. bursaria as well as with their cytoplasm. Immunoelectron microscopy showed that the antigens were located on the decorated spongiome of the radial arms. In immunoblots, mAb IR-2-1 reacted with a band of 67 kDa in all Paramecium species examined. However, no band appeared in the immunoblot of isolated macronuclei of H. obtusa-free P. caudatum and no label was seen in the nuclear matrix of the macronucleus of air-dried P. caudatum. These results suggest that the 67-kDa antigen found in H. obtusa was not imported from the host macronucleus and the same antigen in the host contractile vacuoles and cytoplasm were not derived from the symbiont. These results also showed that an epitope on the decorated spongiome of the Paramecium species is shared by its bacterial symbiont. In contrast to the decorated tubule-specific mAb, DS-1, the antigens for IR-2-1 appeared to be loosely membrane bound as they were lost in paraformaldehyde fixed and acetone permeabilized Paramecium.

Animals↗
PubMedDate not supplied

The striated bands of Paramecium are immunologically distinct from the centrin-specific infraciliary lattice and cytostomal cord.

The pellicle of Paramecium has three two-dimensionally arrayed systems that occupy separate but closely paralleling planes. All three systems are now distinguishable by their differing immunological properties. This study focused on the two deeper systems. The infraciliary lattice lies innermost and labels with centrin-specific antibodies. The middle system, the striated bands, is specifically labeled with a monoclonal antibody that we have raised to a 110 kDa cortical antigen in P. multimicronucleatum. This antibody labels a similar geometric cortical pattern in at least two species, P. multimicronucleatum and P. tetraurelia. Centrin-specific structures appear to be net-like in the above two species but show a more interrupted pattern in P. caudatum. The cytostomal cord is an essentially unbranched extension of the net-like infraciliary lattice and, like it, is centrin-specific. The cord has a unique association with the alveolar sacs which suggest these calcium-storing compartments contribute to the calcium fluxes required for contraction of the cord. A structural rather than a contractile function is favored for the striated bands, based solely on their morphology.

Animals↗
PubMedDate not supplied

Phagosome formation in Paramecium: roles of somatic and oral cilia and of solid particles as revealed by video microscopy.

The roles of somatic and oral cilia and solid particles during digestive vacuole (DV) formation in Paramecium multimicronucleatum were investigated using video-enhanced and immunofluorescence microscopy. Membrane incorporation into DVs was found to increase linearly with increasing particle concentration. The rate of discoidal vesicle transport to the cytopharynx was not affected by particles, showing that particles are not required for membrane trafficking to the cytopharynx. However, the presence of particles leads to an increased membrane fusion between the cytopharyngeal membrane and the discoidal vesicles. When live cells lost their somatic cilia on the left-ventral side anterior to the oral region due to deciliation, membrane incorporation into newly formed DVs was strongly inhibited. Using video-enhanced microscopy, latex beads were seen to be loaded along the quadrulus on the dorsal surface of the buccal cavity, but few beads were seen next to the dorsal and ventral peniculi. Particle sequestration into a pre-formed nascent digestive vacuole (NDV) was studied in Triton X-100-permeabilized cells whose ciliary beating was reactivated by the addition of Mg-ATP. Both beat frequency and the percentage of cells containing bead-labeled NDV were dependent on the Mg-ATP concentration: the higher the beat frequency, the higher the percentage of cells with a bead-labeled NDV. These results suggest that ciliary beating is probably the only mechanism required for particle accumulation in the NDV, while a coordinated beating of the somatic cilia on the left-ventral side anterior to the oral region as well as the quadrulus moves particles into the NDV. The beating of the peniculi may somehow prevent the backward flow of particles out of the NDV.

Animals↗