Energy parameters of Acetabularia cell membrane]

The energy parameters of the mechanism involved in the action of Acetabularia cell membrane were studied. The resistance of the membrane is found to be near 3 kohm per sq. cm when resting and at the peak of excitation it is much lower (100 ohm/cm(2). The power of the mechanism regulating the action of the membrane during the peak. The total power of the membrane system of Acetabularia is 8 microwatt per sp. cm. reaches 6.5 microwatt/cm(2). The energy consumed by the regulatory mechanism during the action potential is minus 290 plus or minus 20 erg/cm(2) and the total energy dissipation is 20 erg/cm(2). The values of the energy consumed by the cell during the action potential are compared with its energy resource.     

An application of membrane theory to tip morphogenesis in Acetabularia

Focusing our attention on the cell wall and the plasmalemma (i.e. the cell membrane), we seek to show that the initiation of the regenerative, growing tip in the unicellular marine alga Acetabularia mediterranea, can be predicted using the techniques of thin-shell and elasticity theory. We build upon and extend the work of Goodwin & Trainor (1985, J. theor. Biol. 117, 79-106), Trainor & Goodwin (1986, Physica D. 21D, 137-145) and Brière & Goodwin (1988, J. theor. Biol. 131, 461-475) where the attention was focused on the calcium-regulated strain and stress fields of the cortical cytoplasm. Finally, we attempt to model the subsequent tip growth using a moving-boundary formulation, with cytosolic free calcium concentration and turgor pressure being the two variables responsible for the growth.     

The majority of cellular fatty acid acylated proteins are localized to the cytoplasmic surface of the plasma membrane

The BC3Hl muscle cell line was previously reported to contain a broad array of fatty acid acylated proteins [Olson, E. N., Towler, D. A., & Glaser, L. (1985) J. Biol. Chem. 260, 3784-3790]. Palmitate was shown to be attached to membrane proteins posttranslationally through thiol ester linkages, whereas myristate was attached cotranslationally, or within seconds thereafter, to soluble and membrane-bound proteins through amide linkages [Olson, E. N., & Spizz, G. (1986) J. Biol. Chem. 261, 2458-2466]. The temporal and subcellular differences between palmitate and myristate acylation suggested that these two classes of acyl proteins might follow different intracellular pathways to distinct subcellular membrane systems or organelles. In this study, we examined the subcellular localization of the major fatty acylated proteins in BC3Hl cells. Palmitate-containing proteins were localized to the plasma membrane, but only a subset of myristate-containing proteins was localized to this membrane fraction. The majority of acyl proteins were nonglycosylated and resistant to digestion with extracellular proteases, suggesting that they were not exposed to the external surface of the plasma membrane. Many proteins were, however, digested during incubation of isolated membranes with proteases, which indicates that these proteins face the cytoplasm. Two-dimensional gel electrophoresis of proteins labeled with [3H]palmitate and [3H]myristate revealed that individual proteins were modified by only one of the two fatty acids and did not undergo both N-linked myristylation and ester-linked palmitylation. Together, these results suggest that the majority of cellular acyl proteins are routed to the cytoplasmic surface of the plasma membrane, and they raise the possibility that fatty acid acylation may play a role in intracellular sorting of nontransmembranous, nonglycosylated membrane proteins.     

Contribution of whole cell and cytoplasmic polypeptides to apparent red cell membrane alterations

We have compared densitometric tracings of whole cell, cytoplasmic and membrane polypeptide electrophoretic patterns in an attempt to distinguish atypical partitioning from intrinsic membrane polypeptide changes occurring as a result of reticulocyte enrichment, metabolic depletion, $\text{N-}\text{ethylmaleimide}$ treatment and hereditary xerocytosis. We report that membrane alterations seen in a reticulocyte-enriched population of normal cells are present in the whole cells prior to membrane isolation. Some of the membrane alterations in metabolically depleted cells and all of those in $\text{N-}\text{ethylmaleimide-treated}$ cells are traced to modifications in the partitioning of polypeptides between membranes and supernatant (cytoplasm) at hemolysis.The power of this approach in resolving the sources of apparent red cell membrane protein alterations is demonstrated in studies with hereditary xerocytes. Suggested altered partitioning of these cells described earlier (Sauberman, N., Fortier, N.L., Fairbanks, G., O'Connor, R.J. and Snyder, L.M. (1979) Biochim. Biophys. Acta 556, 292–313) is further documented and found to be unrelated to the younger cell population or slight metabolic depletion that occurs during the washing of xerocytes prior to hemolysis.     

Rna transport from the nucleus to the cytoplasm in Acetabularia mediterranea]

The biosynthesis of nuclear RNA, its transport from the nucleus to the cytoplasm and distribution in the cytoplasm were studied in Acetabularia mediterranea under different light conditions. It was shown that the nuclear RNA incorportate 3H-uracil more rapidly in the darkness and the transport of labeled RNA from the nucleus slowed down after the transfer of plants in the cold medium in the darkness. To study the distribution of nuclear RNA in the cytoplasm, the 3H-uracil labeled nuclei were transplanted in the rhizoids of unlabeled plants, the dikaryons obtained were kept for different time in the light and in the darkness and the content of 3H-RNA was determined in different stem regions. It was shown that the transport of 3H-RNA in the cytoplasm is slowed down in the darkness and it is distributed by the basal-apical gradient. RNA is rapidly accumulated in the apical stem zone in the light and redistributed afterwards in the basal stem zones. The problem of relationship between the polarity and nuclear RNA distribution in Acetabularia is discussed.     

Damage to the cytoplasmic membrane and cell death caused by lycopene in Candida albicans

Lycopene, an acyclic carotenoid found in tomatoes (Lycopersicon esculentum) and a number of fruits, has shown various biological properties, but its antifungal effects remain poorly understood. The current study investigated the antifungal activity of lycopene and its mode of action. Lycopene showed potent antifungal effects toward pathogenic fungi, tested in an energy-independent manner, with low hemolytic effects against human erythrocytes. To confirm the antifungal effects of lycopene, its effects on the dimorphism of Candida albicans induced by fetal bovine serum (FBS), which plays a key role in the pathogenesis of a host invasion, were investigated. The results showed that lycopene exerted potent antifungal activity on the serum-induced mycelia of C. albicans. To understand the antifungal mode of action of lycopene, the action of lycopene against fungal cell membranes was examined by FACScan analysis and glucose and trehalose-release test. The results indicated that lycopene caused significant membrane damage and inhibited the normal budding process, resulting from the destruction of membrane integrity. The present study indicates that lycopene has considerable antifungal activity, deserving further investigation for clinical applications.     

Poly(3-hydroxybutyrate) granules at the early stages of formation are localized close to the cytoplasmic membrane in Caryophanon latum

Localization of newly synthesized poly(3hydroxybutyrate) (PHB) granules was determined by confocal laser scanning fluorescence microscopy of Nile red-stained cells and by transmission electron microscopy (TEM). PHB granules of Nile red-stained living cells of Caryophanon latum at the early stages of PHB accumulation were frequently found at or close to the cytoplasmic membrane. TEM analysis of the same culture revealed electron-translucent globular structures resembling PHB granules that were nonrandomly distributed in the cell lumen but were frequently found at or close to the cytoplasmic membrane. Immunogold labeling using PHB-specific antiserum confirmed that the electron-translucent structures represented PHB granules. Electron microscopy examination of PHB granules after cell lysis revealed that PHB granules were often associated with membrane vesicles. Nonrandom localization of PHB granules was also found in Beijerinckia indica. Cells of this species harbored one PHB granule at each cell pole. Our results show that newly synthesized PHB granules often are close to or even in physical contact with the cytoplasmic membrane. Possible explanations for this unexpected finding and a hypothetical model of PHB granule formation in C. latum are discussed.     

The TraB protein, which mediates the intermycelial transfer of the Streptomyces plasmid pSN22, has functional NTP-binding motifs and is localized to the cytoplasmic membrane

The traB gene on the Streptomyces conjugative plasmid pSN22 is required for intermycelial plasmid transfer and the mobilization of chromosomal markers (Cma). The predicted amino acid sequence of TraB contains one Walker type-A and two type-B NTP-binding motifs. Site-directed mutagenesis revealed that the type-A motif and one of the type-B motifs, 109 amino acid residues downstream of the type-A motif, were essential for both plasmid transfer and Cma. The second type-B sequence could be changed without any phenotypic effect. A modified traB gene was constructed, resulting in the production of a functional protein with an amino-terminal c-Myc epitope tag for immunological analysis. This protein was associated with the cytoplasmic membrane, suggesting that TraB is a membrane protein that uses energy from ATP hydrolysis to transport DNA between mycelia. The c-Myc tagging of TraB decreased the efficiency of intramycelial plasmid spread, suggesting that TraB is involved in both inter- and intramycelial transfer processes.     

Cu2+-induced permeability of the Escherichia coli cytoplasmic membrane]

Cu(2+)-induced permeability of cytoplasmic membranes of Escherichia coli for different cations and neutral molecules of saccharose was estimated by studying their effect on cell plasmolysis during uncharged exchange of cytoplasmic K+ ions by periplasmic space cations. The addition of copper resulted in the exchange of K+ ions by periplasmic Na+, Tris+, streptomycin2+, Cu2+, Ca2+, Mg2+, Cd2+, and Mn2+. It is concluded that Cu(2+)-induced conducting pathways in bacterial membranes are hydrophilic channels with a radius of approximately 0.5 nm and a nonselective permeability for different cations.     

Quantitative analysis of glomerular basement membrane glycosaminoglycans and evidence for their binding to adjacent cell membrane lipids

Glycosaminoglycans (GAG) were isolated from rat renal glomerular basement membranes subjected to extraction with detergents, and were quantitatively analyzed using a recently described competitive binding assay that is specific for and sensitive to microgram amounts of chondroitin and heparan sulfate. Whereas crude membranes prepared by osmotic lysis contained only 6 micrograms/mg dry wt, subsequent treatment with Triton X-100 or deoxycholate (DOC) increased measureable GAG to about 17 and 34 micrograms/mg, respectively. Repeated freezing and thawing of isolated glomeruli also augmented measurable GAG content in subsequently osmotically lysed membranes to levels observed in Triton-treated membranes. DOC solubilized approximately equal to 15-20% of membrane-associated GAG. Chondroitin sulfate comprised approximately equal to 30% of total GAG, and all of the chondroitin sulfate but only 10% of the heparan sulfate was extracted from the insoluble matrix by DOC. The findings indicate that GAG content of glomerular basement membrane is several-fold higher than previously estimated, and that a substantial portion is bound to cell membrane lipids. The results further suggest two populations of GAG in basement membrane; one that is intercalated with adjacent cell membranes, and another that remains as an integral component of the insoluble matrix after detergent extraction.     

SNAREs contribute to the specificity of membrane fusion

Intracellular membrane fusion is mediated by the formation of a four-helix bundle comprised of SNARE proteins. Every cell expresses a large number of SNARE proteins that are localized to particular membrane compartments, suggesting that the fidelity of vesicle trafficking might in part be determined by specific SNARE pairing. However, the promiscuity of SNARE pairing in vitro suggests that the information for membrane compartment organization is not encoded in the inherent ability of SNAREs to form complexes. Here, we show that exocytosis of norepinephrine from PC12 cells is only inhibited or rescued by specific SNAREs. The data suggest that SNARE pairing does underlie vesicle trafficking fidelity, and that specific SNARE interactions with other proteins may facilitate the correct pairing.     

Polar distribution of RNA in the cytoplasm of Acetabularia mediterranea]

The distribution of total RNA and its individual fractions in two regions of Acetabularia mediterranea stem during regeneration was investigated. During regeneration of both the nuclear and enucleated cells, RNA concentration increases in the cytoplasm of growth zone whereas it changes insignificantly in the central stem region. A study of the qualitative RNA composition in the same stem regions has shown that during regeneration high molecular weight RNA fractions (main peaks - 0,56-10(6) and 1.04-10(6) Dalton) are found in the growth zone and are practically absent from the central cell region. Low molecular weight RNA (supposedly, tRNA and products of RNA destruction) are present in both the cell regions under study.     

Relationships between plasma membrane depolarization,nuclear membrane breakdown,and the appearance of cytoplasmic factors during the first meiotic division in Rana pipiens oocytes

The present study examines the relationship among three events which occur in Rana pipiens oocytes following hormone-induced reinitiation of meiosis: (i) plasma membrane depolarization, (ii) nuclear membrane breakdown, and (iii) the appearance of specific cytoplasmic factors. Preventing plasma membrane depolarization with a voltage clamp did not prevent nuclear breakdown. Transfer of cytoplasm from hormone-induced oocytes into naive oocytes resulted in both depolarization and nuclear breakdown. Depolarization of injected oocytes was dependent on protein synthesis, whereas nuclear breakdown was not. These findings, together with other recent evidence, suggest that different cytoplasmic factors may initiate protein synthesis-independent nuclear membrane events and protein synthesis-dependent plasma membrane events.     

Species specificity, age factors, and various neurochemical correlates of the animal spontaneous behavior after exposure to electromagnetic field of the ultralow intensity

Behavioral and neurochemical reactions of small laboratory animals (mice and rats of different age) under exposure to ultralow-intensity electromagnetic fields (EMF, frequency of 4200 and 970 MHz, modulated by a quasistochastic signal in the range of 20-20,000 Hz, power density 15 microW/cm2, specific body absorption rate up to 4.5 mJ/kg) were studied. The EMF basically inhibited the locomotor and exploratory activity in the "open-field" test. The species- and age-specific features rather than radiation conditions dominated. However, decrease in the EMF frequency considerably intensified the observed effect. Change in animal behavior was accompanied by shifts in neurochemical processes, i.e., sharp activation of serotoninergic and inhibition of morepinephrinergic system.