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The significance of cAMP induced alterations in the cellular structure of Phycomyces.

Effect of cyclic AMP (cAMP) on Phycomyces blakesleeanus was studied by growing sporangiospores on glucose-asparagine agar or liquid medium containing three different levels of cAMP (10, 20 and 40 micronM) in addition to the control (no cAMP added). The response of Phycomyces to the exogenous cAMP concentration in the medium is as follows: (1) the time required for germ tube emergence is reduced; (2) the diameter of the mycelium is increased (sometimes more than 10 times) and frequency of branching is also increased; (3) the cell wall of the mycelium is thickened (in some cases more than 5 times); (4) the glycogen in the cytoplasm is decreased as visualized in thin sections and also demonstrated in biochemical quantitation; and (5) the distribution of intercalated membranous particles (Imp) on plasma membrane is altered and this can be easily detected in freeze-fractured replica. Such a change in Imp is seen in the formation of small clusters of aggregated particles on the plasmic half (PF) and craters on the complementary exoplasmic half (EF) of the plasma membrane. Although the mechanism of cAMP action requires further exploration, it is possible that the addition of cAMP to the culture medium leads to degradation of glycogen and enhancement of chitin synthesis since the cell wall is largely composed of chitin. The alteration in Imp may be related to a change in the activity of chitin synthetase which is a plasma membrane-bound enzyme.

Cyclic AMP↗

Alteration of skeletal muscle cellular structures by potassium depletion.

After rats had been fed a low-potassium diet for 4 to 8 weeks, skeletal muscle showed ultrastructural changes involving membranous organelles. Mitochondria were often swollen, condensed, or disintegrated. The transverse tubules were disoriented, focally dilated, and tortuous. The sarcoplasmic reticulum showed various degrees of dilatation. Vacuoles of different sizes occurred frequently. Whirls of membranes were closely associated with any of the membranous organelles, especially near an orifice of a transverse tubule. Supplementation of potassium reversed these changes. These findings are very similar to those in patients with hypokalemic periodic paralysis. The vacuolar myopathy in these patients may be secondary to the electrolyte alteration in skeletal muscles, and the chronic weakness of some patients may be due to excitation-contraction uncoupling as a result of the involvement of sarcotubular systems.

Animals↗

The effects of new cytochalasins from Phomopsis sp. and the derivatives on cellular structure and actin polymerization.

The effects of ten 10-phenyl-[11]cytochalasins produced by Phomopsis sp. including novel compounds having 5,7- or 6,7-glycol structures and their derivatives, on the cell morphology and actin distribution in C3H-2K cells, as well as on lymphocyte capping and actin polymerization, were examined. The structure-activity relationship reported in the previous papers has been confirmed. The novel glycol type compounds showed little or no activity, suggesting the importance of the perhydroisoindol-1-one nucleus for the manifestation of the cytochalasin actions.

Actins↗

The nuclear matrix is a thermolabile cellular structure.

Heat shock sensitizes cells to ionizing radiation, cells heated in S phase have increased chromosomal aberrations, and both Hsp27 and Hsp70 translocate to the nucleus following heat shock, suggesting that the nucleus is a site of thermal damage. We show that the nuclear matrix is the most thermolabile nuclear component. The thermal denaturation profile of the nuclear matrix of Chinese hamster lung V79 cells, determined by differential scanning calorimetry (DSC), has at least 2 transitions at Tm = 48 degrees C and 55 degrees C with an onset temperature of approximately 40 degrees C. The heat absorbed during these transitions is 1.5 cal/g protein, which is in the range of enthalpies for protein denaturation. There is a sharp increase in 1-anilinonapthalene-8-sulfonic acid (ANS) fluorescence with Tm = 48 degrees C, indicating increased exposure of hydrophobic residues at this transition. The Tm = 48 degrees C transition has a similar Tm to those predicted for the critical targets for heat-induced clonogenic killing (Tm = 46 degrees C) and thermal radiosensitization (Tm = 47 degrees C), suggesting that denaturation of nuclear matrix proteins with Tm = 48 degrees C contribute to these forms of nuclear damage. Following heating at 43 degrees C for 2 hours, Hsc70 binds to isolated nuclear matrices and isolated nuclei, probably because of the increased exposure of hydrophobic domains. In addition, approximately 25% of exogenous citrate synthase also binds, indicating a general increase in aggregation of proteins onto the nuclear matrix. We propose that this is the mechanism for increased association of nuclear proteins with the nuclear matrix observed in nuclei Isolated from heat-shocked cells and is a form of indirect thermal damage.

Animals↗

Extracellular serine-proteinases isolated from Streptomyces alboniger: partial characterization and effect of aprotinin on cellular structure.

Streptomyces alboniger ATCC 12461 grown in brain heart infusion (BHI) medium produced two extracellular serine-proteinases, denoted SP I and SP II, which were purified by ammonium sulfate precipitation and aprotinin-agarose affinity chromatography. SP I was purified 88,9-fold and SP II 66,7- fold, with 33.4% and 10.4% yield, respectively. The optimum pH for the proteinases activity, using a-N-p-tosyl-L-arginine-methyl ester (TAME) as substrate, was 9-10 and the optimum temperature was 37 degrees C. The proteolytic activity of SP I and SP II was inhibited by aprotinin and SP I was partially inhibited by leupeptin, both serine-proteinase inhibitors. S. alboniger growth in BHI-liquid medium decreased when 5 mg/ml, 10 mg/ml of aprotinin was used, being completely inhibited with 20 mg/ml and 40 mg/ml. At the ultrastructural level, aprotinin-treated S. alboniger cells showed swelling of the bacterial body and condensation of the genetic material, probably related to the inhibition of its growth.

Aprotinin↗

Fluorescence microscopy with antisera against specific cellular structure: double photography method for cell identification in populations of multiple cell types.

Immunofluorescent staining techniques using antitubulin antibody have been difficult to apply to meiotic tissue (testis) because of the large number of cell types present. Such techniques customarily use a fluorescent dye to counterstain nuclei, and this counterstain is hard to distinguish because of the fluorescence of the antitubulin. By counterstaining with dilute hematoxylin, we can photograph the same field using UV and then conventional illumination. This double photography allows us to identify precisely the many types of cells present, and it will be a useful tool for reexamining the staging of spermatogenesis.

Animals↗

[Autoradiographic study of cellular structure of the epithelium of the duct of testicular epididymis of rats].

Mature rats were given intraperitoneal injections of H-3-thymidine (1 mkk/g 1-32 hours before being killed. Labelled and non-labelled mitoses and interphase cells of different types were counted in each zone of the epididymis autographs. The diurnal fluctuatiof the mitoticindex (Im) was found: form 0,19% in the day-time to 0.33% in the night at and morning hours (psmaller than 0.05). The average diurnal Im was equal to .23%-0.03. The fist wave of labelled mitoses of the epithelial cells was observed during 32 hours, tg-2 (3-5 hours) and ts(13-14 hours) were graphically calculated. The time tg-2-tm-ts was equal to 19-20 hours. Therprietal (0.87%), basal (1.87%) and oreolar (2.20%) cells of the epidermis duct labelled 1 hour after ijection of H-3-thymidine. The apical cells (3.%) were labelled 8 hours later, while the light ones were not labelled during the whole period of observation. On these grounds, the parietal, basal and oreolar cells are considered to be proliferative cells, while the light and apical ones-to be their derivatives in the epidemis epithelium. Besides, the oreolar cells may be regareded as a foreign element in the epidermis according to their morphological features and ability to migrate throughout the total depth of the epithelial layer.

Animals↗

[Isolation and characteristics of the cellular structures of Candida tropicalis].

Using the method of radioactive indicators, the specific content of proteins, lipids, total phosphorus, and lipid phosphorus was estimated in the morphological cell fractions of Candida tropicalis after growth in a liquid mineral medium containing n-octadecane as a source of carbon. The morphological cell fractions were produced by means of differential centrifuging. The following comparatively pure fractions were obtained simultaneously: soluble fraction, microsomes, mitochondria, large membranes, and cell walls.

Candida↗