The cytoplasmic sites of the snRNP protein complexes are punctate structures that are responsive to changes in metabolism and intracellular architecture

Five anti-Sm monoclonal antibodies, Y12, 7.13, KSm4, KSm6, and 128, stain similar discrete punctate structures distributed throughout the cytoplasm of hamster fibroblasts in addition to the expected intense nuclear staining. Several criteria suggest the cytoplasmic staining reflects the cytoplasmic pools of snRNP core proteins. The relative intensity of the cytoplasmic staining is similar to the 30% relative abundance of the cytoplasmic snRNP core proteins compared to the nuclear snRNP core proteins based on cell-fractionation studies. Moreover, the cytoplasmic staining is removed by the same extraction conditions that solubilize the pools of cytoplasmic snRNP core proteins. The cytoplasmic sites of staining are typically spherical but heterogeneous in diameter (0.2-0.5 microm). The larger particles greatly exceed the diameter of individual snRNP core particles and are likely to represent centers of many snRNP proteins or snRNP protein complexes. The staining, though punctate, is evenly dispersed throughout the cytoplasm with no evidence of major compartmentalization. The cytoplasmic staining pattern collapses into larger foci of intensely staining structures when cellular energy levels are depleted or when cells are exposed to hypertonic medium. Unlike the normal sites of snRNP protein cytoplasmic staining, these larger collapsed foci resist detergent extraction. These results suggest that the cytoplasmic staining identified with the anti-Sm monoclonal antibodies represents the large pools of snRNP core proteins in the cytoplasm.     

Regulation of cell volume via microvillar ion channels

A novel mechanism of cellular volume regulation is presented, which ensues from the recently introduced concept of transport and ion channel regulation via microvillar structures (Lange K, 1999, J Cell Physiol 180:19-35). According to this notion, the activity of ion channels and transporter proteins located on microvilli of differentiated cells is regulated by changes in the structural organization of the bundle of actin filaments in the microvillar shaft region. Cells with microvillar surfaces represent two-compartment systems consisting of the cytoplasm on the one side and the sum of the microvillar tip (or, entrance) compartments on the other side. The two compartments are separated by the microvillar actin filament bundle acting as diffusion barrier ions and other solutes. The specific organization of ion and water channels on the surface of microvillar cell types enables this two-compartment system to respond to hypo- and hyperosmotic conditions by activation of ionic fluxes along electrochemical gradients. Hypotonic exposure results in swelling of the cytoplasmic compartment accompanied by a corresponding reduction in the length of the microvillar diffusion barrier, allowing osmolyte efflux and regulatory volume decrease (RVD). Hypertonic conditions, which cause shortening of the diffusion barrier via swelling of the entrance compartment, allow osmolyte influx for regulatory volume increase (RVI). Swelling of either the cytoplasmic or the entrance compartment, by using membrane portions of the microvillar shafts for surface enlargement, activates ion fluxes between the cytoplasm and the entrance compartment by shortening of microvilli. The pool of available membrane lipids used for cell swelling, which is proportional to length and number of microvilli per cell, represents the sensor system that directly translates surface enlargements into activation of ion channels. Thus, the use of additional membrane components for osmotic swelling or other types of surface-expanding shape changes (such as the volume-invariant cell spreading or stretching) directly regulates influx and efflux activities of microvillar ion channels. The proposed mechanism of ion flux regulation also applies to the physiological main functions of epithelial cells and the auxiliary action of swelling-induced ATP release. Furthermore, the microvillar entrance compartment, as a finely dispersed ion-accessible peripheral space, represents a cellular sensor for environmental ionic/osmotic conditions able to detect concentration gradients with high lateral resolution. Volume regulation via microvillar surfaces is only one special aspect of the general property of mechanosensitivity of microvillar ionic pathways.     

The Role of Biomacromolecular Crowding,Ionic Strength,and Physicochemical Gradients in the Complexities of Life's Emergence

Summary: We have developed a general scenario of prebiotic physicochemical evolution during the Earth''s Hadean eon and reviewed the relevant literature. We suggest that prebiotic chemical evolution started in microspaces with membranous walls, where external temperature and osmotic gradients were coupled to free-energy gradients of potential chemical reactions. The key feature of this scenario is the onset of an emergent evolutionary transition within the microspaces that is described by the model of complex vectorial chemistry. This transition occurs at average macromolecular crowding of 20 to 30% of the cell volume, when the ranges of action of stabilizing colloidal forces (screened electrostatic forces, hydration, and excluded volume forces) become commensurate. Under these conditions, the macromolecules divide the interior of microspaces into dynamically crowded macromolecular regions and topologically complementary electrolyte pools. Small ions and ionic metabolites are transported vectorially between the electrolyte pools and through the (semiconducting) electrolyte pathways of the crowded macromolecular regions from their high electrochemical potential (where they are biochemically produced) to their lower electrochemical potential (where they are consumed). We suggest a sequence of tentative transitions between major evolutionary periods during the Hadean eon as follows: (i) the early water world, (ii) the appearance of land masses, (iii) the pre-RNA world, (iv) the onset of complex vectorial chemistry, and (v) the RNA world and evolution toward Darwinian thresholds. We stress the importance of high ionic strength of the Hadean ocean (short Debye''s lengths) and screened electrostatic interactions that enabled the onset of the vectorial structure of the cytoplasm and the possibility of life''s emergence.

Architecture is what ultimately distinguishes a living cell from a soup of the chemicals of which it is composed.—Franklin Harold (40)

     

The physicochemical nature of 37 degrees C cytoplasmic glucocorticoid receptors in HeLa S3 cells

The physicochemical properties of size, shape and surface charge have been determined for the soluble fraction of cytoplasmic glucocorticoid receptors which are located in the HeLa S3 cell cytoplasm after incubation of whole cells with glucocorticoid at 37 degrees C. Under hypotonic buffer conditions approximately 80% of the total recovered [3H]triamcinolone acetonide receptor complexes sedimented through a 5-20% density gradients to the tube bottom, and approximately 90% eluted from a Sephacryl S-300 gel exclusion column in the void volume. Increasing the [KCl] of the buffer in the sucrose density gradients, and gel exclusion columns to 0.15 M caused a reduction in the percentage of this large aggregate to approximately 64% and approximately 75%, respectively. Further increases in the [KCl] during analysis to 0.4 M reduced the percentage of rapidly sedimenting receptors to approximately 62%, and shifted the sedimentation coefficient of the slower sedimenting receptors from approximately 5.2 S to 3.9 S. These conditions also decreased the fraction of receptor in the void volume of gel exclusion columns to 67%. Ion exchange analysis of receptor binding to DEAE cellulose, hydroxylapatite, phosphocellulose, and DNA cellulose revealed heterogenous populations of receptor species; comprising both "unactivated" and "activated" receptor forms. The ratios of unactivated/activated receptors was highly dependent on the matrix employed and differed substantially among those evaluated. For example, by the criteria of DEAE cellulose and phosphocellulose chromatography approximately 60% of the total 37 degrees C cytoplasmic receptors were in the "activated" state. A large fraction of these receptors, however, failed to bind to DNA cellulose. These results demonstrate that the glucocorticoid receptors which remain in the HeLa S3 cytoplasm at 37 degrees C do not bind to ion exchange materials, which are used as indexes of receptor "activation," in a uniform manner. We hypothesize that the diminished DNA binding capability of these receptors accounts for their cellular localization in the HeLa S3 cell cytoplasm at 37 degrees C.     

A review comparing deoxyribonucleoside triphosphate (dNTP) concentrations in the mitochondrial and cytoplasmic compartments of normal and transformed cells

The deoxyribonucleoside triphosphate (dNTP) pools that support the replication of mitochondrial DNA are physically separated from the rest of the cell by the double membrane of the mitochondria. Perturbed homeostasis of mitochondrial dNTP pools is associated with a set of severe diseases collectively termed mitochondrial DNA depletion syndromes. The degree of interaction of the mitochondrial dNTP pools with the corresponding dNTP pools in the cytoplasm is currently not clear. We reviewed the literature on previously reported simultaneous measurements of mitochondrial and cytoplasmic deoxyribonucleoside triphosphate pools to investigate and quantify the extent of the influence of the cytoplasmic nucleotide metabolism on mitochondrial dNTP pools. We converted the reported measurements to concentrations creating a catalog of paired mitochondrial and cytoplasmic dNTP concentration measurements. Over experiments from multiple laboratories, dNTP concentrations in the mitochondria are highly correlated with dNTP concentrations in the cytoplasm in normal cells in culture (Pearson R = 0.79, p = 3 × 10(-7)) but not in transformed cells. For dTTP and dATP there was a strong linear relationship between the cytoplasmic and mitochondrial concentrations in normal cells. From this linear model we hypothesize that the salvage pathway within the mitochondrion is only capable of forming a concentration of approximately 2 μM of dTTP and dATP, and that higher concentrations require transport of deoxyribonucleotides from the cytoplasm.     

Low viscosity in the aqueous domain of cell cytoplasm measured by picosecond polarization microfluorimetry

Information about the rheological characteristics of the aqueous cytoplasm can be provided by analysis of the rotational motion of small polar molecules introduced into the cell. To determine fluid-phase cytoplasmic viscosity in intact cells, a polarization microscope was constructed for measurement of picosecond anisotropy decay of fluorescent probes in the cell cytoplasm. We found that the rotational correlation time (tc) of the probes, 2,7-bis-(2-carboxyethyl)-5-(and-6-)carboxyfluorescein (BCECF), 6-carboxyfluorescein, and 8-hydroxypyrene-1,3,6-trisulfonic acid (HPTS) provided a direct measure of fluid-phase cytoplasmic viscosity that was independent of probe binding. In quiescent Swiss 3T3 fibroblasts, tc values were 20-40% longer than those in water, indicating that the fluid-phase cytoplasm is only 1.2-1.4 times as viscous as water. The activation energy of fluid-phase cytoplasmic viscosity was 4 kcal/mol, which is similar to that of water. Fluid-phase cytoplasmic viscosity was altered by less than 10% upon addition of sucrose to decrease cell volume, cytochalasin B to disrupt cell cytoskeleton, and vasopressin to activate phospholipase C. Nucleoplasmic and peripheral cytoplasmic viscosities were not different. Our results establish a novel method to measure fluid-phase cytoplasmic viscosity, and indicate that fluid-phase cytoplasmic viscosity in fibroblasts is similar to that of free water.     

A Review Comparing Deoxyribonucleoside Triphosphate (dNTP) Concentrations in the Mitochondrial and Cytoplasmic Compartments of Normal and Transformed Cells

The deoxyribonucleoside triphosphate (dNTP) pools that support the replication of mitochondrial DNA are physically separated from the rest of the cell by the double membrane of the mitochondria. Perturbed homeostasis of mitochondrial dNTP pools is associated with a set of severe diseases collectively termed mitochondrial DNA depletion syndromes. The degree of interaction of the mitochondrial dNTP pools with the corresponding dNTP pools in the cytoplasm is currently not clear. We reviewed the literature on previously reported simultaneous measurements of mitochondrial and cytoplasmic deoxyribonucleoside triphosphate pools to investigate and quantify the extent of the influence of the cytoplasmic nucleotide metabolism on mitochondrial dNTP pools. We converted the reported measurements to concentrations creating a catalog of paired mitochondrial and cytoplasmic dNTP concentration measurements. Over experiments from multiple laboratories, dNTP concentrations in the mitochondria are highly correlated with dNTP concentrations in the cytoplasm in normal cells in culture (Pearson R = 0.79, p = 3 × 10^?7) but not in transformed cells. For dTTP and dATP there was a strong linear relationship between the cytoplasmic and mitochondrial concentrations in normal cells. From this linear model we hypothesize that the salvage pathway within the mitochondrion is only capable of forming a concentration of approximately 2 μM of dTTP and dATP, and that higher concentrations require transport of deoxyribonucleotides from the cytoplasm.     

Herpesvirus infection modifies adenovirus RNA metabolism in adenovirus type 5-transformed cells.

The effect of herpes simplex virus (HSV) infection of mRNA metabolism was examined in a system where the fate of specific RNA sequence can be assayed. Adenovirus type 5-transformed rat embryo cell line 107 synthesizes adenovirus-specific RNA (ad-RNA), which functions in the cytoplasm as mRNA. We have utilized ad-RNA as a model for mRNA metabolism, and in a preliminiary study we characterized ad-RNA in the nucleus and cytoplasm by hybridization to filter-bound adenovirus DNA. The results indicated the as-RNA accumulates in the nucleus and that cytoplasmic polyadenylic acid [poly(A)]-containing ad-RNA turns over with a half-life of a few hours. Pulse-chase experiments confirmed these observations and a half-life of about h was determined for the poly(A)-containing cytoplasmic ad-RNA. A second class of ad-RNA remains in the nucleus, where it turns over with a longer hlaf-life (about 24 h). The infection of 107 cells by HSV was restricted at 37 degree C, giving a burst size of 5 PFU per cell and allowing continued host DNA synthesis. Protein synthesis was inhibited greater than 50% by 7 h after infection, and total RNA synthesis was 50% inhibited by 4 h after infection. During the first 8 h after infection, HSV has little effect on the rate of synthesis of ad-RNA as determined by hybridization of nuclear RNA samples, but,during the same period, HSV inhibits the accumulation of poly(A)-containing ad-RNA in the cytoplasm. The degree of this inhibition increases steadily throughout this period and reaches 60% by 6.5 to 8 h after infection. Nosignificant effect was seen on the accumulation of total cellular poly(A)-containing RNA. It was concluded from these experiments that HSV infection alters the metabolism of ad-RNA so as to prevent the normal appearance of the poly(A)-containing mRNA in the cytoplasm. The result for ad-RNA may not represent the behavior of total cellular poly(A)-containing RNA under conditions where infection is restricted.     

THE PERMEABILITY OF THE AMPHIBIAN OOCYTE NUCLEUS, IN SITU

Ultralow temperature radioautography, suitable for the quantitative localization of diffusible solutes, was used to study the permeability of the nuclear envelope in the intact amphibian oocyte Sucrose-³H solutions were injected into mature oocytes, in volumes of 0 016–0 14% of that of the cell, and the subsequent movement of the solute was recorded. The resultant radioautographs show diffusion gradients in the cytoplasm and nucleus, and concentration gradients across the nuclear envelope Analysis of these gradients discloses that the nuclear envelope is as permeable as a comparable structure composed of cytoplasm, and is about 10⁸ times more permeable than the oocyte plasma membrane The diffusion coefficient of sucrose in cytoplasm is 2 x 10^-6 cm²/sec, or about one-third its diffusivity in pure water. This reduction can probably be accounted for by an effective lengthening of the diffusional path because of obstruction by cytoplasmic inclusions. The nuclear: cytoplasmic sucrose concentration ratio at diffusional equilibrium is about 3 05, or 1.6 times as great as expected from the water content of the two compartments This asymmetry is attributed to an unavailability of 36% of the cytoplasmic water as solvent Finally, sucrose entry into oocytes from a bathing solution was monitored by whole cell analysis and radioautography. These and the microinjection results are consistent with a model in which sucrose entry into the cell is entirely limited by the permeability of the plasma membrane. The results are inconsistent with cell models that hypothesize a short-circuit transport route from the extracellular compartment to the nucleus, and with models in which cytoplasmic diffusion is viewed as limiting the rate of solute permeation.     

Compartmentalization of metabolism of spatially-delimiting amino acid pools of yeast cells

The spatially different amino acid pools (i.e. cytoplasmic, vacuolar and mitochondrial) of yeast cells are metabolically compartmentalized. The accumulation of amino acids in these pools occurs at different rates; the highest rates are observed for glutamate and alanine. The former is predominantly accumulated in the cytoplasm, the latter--in the vacuoles. The renewal rates of the amino acid pools are also different. Each of them contains at least two subpools, readily convertible and relatively stable ones. The readily convertible subpools of the cytoplasmic and mitochondrial pools predominantly contain glutamate, aspartate, valine and alanine; that of the vacuolar pool--alanine. The bulk of the readily convertible alanine subpool (67%) is localized in the vacuoles, that of glutamate and aspartate (85 and 68%, respectively)--in the cytoplasm.     

Cytoplasmic DNA synthesis in rhinovirus type 14-inoculated KB cells.

Rhinovirus type 14 (RV14) incuced a transient statistically significant stimulation in synthesis of DNA which appeared between 0 and 3 h post-inoculation in the cytoplasm of high density monolayer cultures of KB cells. Newly synthesized DNA was measured by incorporation of [3H] thymidine into acid-insoluble DNAase-sensitive material and the cytoplasmic location established by cell fractionation and electron microscope radioautographic methods. A minimum of 10 plaque-forming units per cell of RV14 was required to stimulate DNA synthesis which did not occur above 34.5 degrees C, a temperature optimal for virus replication. Cytoplasmic DNA taken from RV14-infected or control cells could be differentiated from the bulk of cell (nuclear) DNA by several criteria, including: (1) RV14 induction of synthesis; (2) lower buoyant density and greater heterogeneity in CsCl and ethidium bromide/CsCl gradients; and (3) a different kinetic complexity upon reannealing. The Cot 1/2 value of cytoplasmic DNA, calclated as 50--100 from reassociation profiles, was about 10-fold less complex than the Cot 1/2 value of nuclear DNA (800-1000). These data rule out the possibility that cytoplasmic DNA arises by random breakage of nuclear DNA during cell disruption and extraction and are compatible with the hypothesis that inoculation of KB cells with RV14 results in stimulation of synthesis of a specific class of cell DNA which is detected in the cytoplasm.     

Cytoplasmic assembly of small nuclear ribonucleoprotein particles from 6 S and 20 S RNA-free intermediates in L929 mouse fibroblasts.

Newly transcribed small nuclear RNAs (snRNAs) appear transiently in the cytoplasm where they assemble with snRNP core proteins (B, D, E, F, and G) stored in large pools of snRNA-free intermediates before returning permanently to the nucleus. In this report, the cytoplasmic assembly of snRNP core particles in L929 mouse fibroblasts was investigated by kinetic analysis of assembly intermediates resolved on sucrose gradients. Immunoprecipitation of gradient fractions with anti-snRNP autoimmune antisera identify pools of 6 and 20 S snRNA-free snRNP protein intermediates. The snRNP B protein has a heterodisperse sedimentation from 4 to 20 S with peaks at 6 and 20 S, and the snRNP D protein is in a bimodal distribution at 6 and 20 S. At 6 S the D protein is assembled with the E, F, and G proteins into a RNA-free core particle with a stoichiometry of D4EFG. SnRNP assembly proceeds by snRNA assembling initially with the 6 S D4EFG particle and then two copies of the B protein to form an 11-15 S SnRNP particle. The 20 S forms of the D protein in the cytoplasm are less stable than the 6 S D4EFG particle. The U1-specific A and C proteins leak from isolated nuclei and appear in the cytoplasmic fractions where they sediment from 10 to 20 S and from 4 to 8 S, respectively.     

THE NUCLEAR PERMEABILITY, INTRACELLULAR DISTRIBUTION, AND DIFFUSION OF INULIN IN THE AMPHIBIAN OOCYTE

[³H]Inulin (mol wt ≈ 5,500) solutions are microinjected into the cytoplasm of mature oocytes of Rana pipiens and the subsequent movement of the solute recorded by quantitative ultralow temperature autoradiography. The autoradiographs show transient cellular diffusion gradients, the influence of the nucleus on these gradients, and the nuclear:cytoplasmic distribution of inulin. Analysis leads to the following conclusions: (a) Inulin diffuses in cytoplasm at about 3 x 10^-6 cm²/s, or one-fifth as rapidly as in water. Most of this decrease is attributable to the increased tortuosity of the diffusional path due to the presence of inclusions and macromolecules. (b) The nuclear envelope is very permeable to inulin; its resistance to inulin's passage is similar to that of cytoplasm. The envelope appears to play a negligible role in regulating the nucleocytoplasmic movement of solutes smaller than macromolecules, (c) Inulin concentrates in the nucleus to four times its cytoplasmic level; this is attributed to solute exclusion from cytoplasmic water. Evidence is presented that among hydrophilic solutes the degree of exclusion increases with molecular size. The potential significance of cytoplasmic exclusion processes to understanding secretion and the intracellular movement of macromolecules is briefly discussed.     

Effects of aluminum in red spruce (Picea rubens) cell cultures: Cell growth and viability, mitochondrial activity, ultrastructure and potential sites of intracellular aluminum accumulation

The effects of Al on red spruce ( Picea rubens Sarg.) cell suspension cultures were examined using biochemical, stereological and microscopic methods. Exposure to Al for 24–48 h resulted in a loss of cell viability, inhibition of growth and a significant decrease in mitochondrial activity. Soluble protein content increased in cells treated with Al. Using energy-dispersive X-ray microanalysis on sections of freeze-substituted cells that had no obvious disruption in cytoplasmic or cell wall structure, Al (always in the presence of P) was detected in dense regions in cell walls, cytoplasm, plastids and vacuoles after 48 h exposure to Al. Stereological quantification of spruce cell structure showed that, after 24 h of Al treatment, intact cells had increased vacuolar and total cell volume, but the nuclear volume did not change. In addition, Al treatment resulted in increased surface area of Golgi membranes and endoplasmic reticulum. The biochemical and ultrastructural alterations in red spruce cells, in combination with the presence of Al in cellular organelles of visually intact cells, suggest that Al movement occurred across the plasma membrane without major cellular disruption. Detailed short-term time course studies are needed to determine if intracellular Al in these cells results from its passage into cells through submicroscopic lesions in the plasma membrane or it is taken up into the symplast through the intact membrane by an active, but slow, process.     

Cytoplasmic deletion processes during spermatogenesis in mosses

Comparative ultrastructural observations reveal that cytoplasmic deletion during spermatogenesis in Sphagnum and other mosses (Bryopsida) has two distinct phases. In young spermatids, Golgi-derived vesicles produce the mucopolysaccharide sheaths in which the gametes are liberated. Golgi bodies, however, play no part in removal of cytoplasm during gamete maturation. Rounding off of the cells during this process results in a 50% reduction in volume. Mid-spermatid stages in Sphagnum are characterised by the sequential loss of Golgi bodies and endoplasmic reticulum (ER) but no further diminution of the cytoplasm. The final stages of nuclear metamorphosis and chromatin condensation, in late spermatids, are marked by the sudden appearance, in the otherwise featureless central cytoplasm, of a membrane vesicle complex (MVC) comprising cisternae, tubules, and smooth and coated vesicles. Following repositioning of the MVC beneath the plasma membrane, rapid shrinkage of the cytoplasm is associated with the presence of vesicle fusion profiles at the cell surface. The MVC is considered to be intimately involved in cytoplasmic breakdown and loss. Acid phosphatase activity can be detected throughout spermatogenesis. Spermatogenous cells and young spermatids possess relatively low levels of the enzyme, restricted to the ER and perinuclear space, but particularly high levels occur in the MVC region of late spermatids of Sphagnum. The deletion process in Bryopsida is much more gradual than that of Sphagnum. Mid-spermatids contain sheets of ER, Golgi with small vesicles, and irregular cisternae associated with coated vesicles. Vacuoles derived either from dilation of the ER or the coated vesicle complexes gradually increase in size and number at the expense of the cytoplasm. During the early stages of chromatin condensation, a large central vacuole opens onto the anterior face of the gametes. Further discharge of vesicles continues throughout gamete maturation. A comparative survey of spermatogenesis in land plants indicates that cytoplasmic deletion is achieved in different ways in different groups. We speculate that the spermatozoids of the common ancestor of archegoniate plants probably possessed large amounts of cytoplasm. The deletion mechanisms may have originated from a contractile vacuole apparatus.     

Spermiogenesis in the bullfrog (Rana catesbeiana): a study of cytoplasmic events including cell volume changes and cytoplasmic elimination

The process by which spermatid cytoplasmic volume is reduced and cytoplasm eliminated during spermiogenesis was investigated in the bullfrog Rana catesbeiana. At early phases of spermiogenesis, newly formed, rounded spermatids were found within spermatocysts. As acrosomal development, nuclear elongation, and chromatin condensation occurred, spermatid nuclei became eccentric within the cell. A cytoplasmic lobe formed from the caudal spermatid head and flagellum and extended toward the seminiferous tubule lumen. The cytoplasmic lobe underwent progressive condensation whereby most of its cytoplasm became extremely electron dense and contrasted sharply with numerous electron-translucent vesicles contained therein. At the completion of spermiogenesis, many spermatids with their highly condensed cytoplasm still attached were released from their Sertoli cell into the lumen of the seminiferous tubule. There was no evidence of the phagocytosis of residual bodies by Sertoli cells. Because spermatozoa are normally retained in the testis in winter and are not released until the following breeding season, sperm were induced to traverse the duct system with a single injection of hCG. Some spermatids remained attached to their cytoplasm during the sojourn through the testicular and kidney ducts; however, by the time the sperm reached the Wolffian duct, separation had occurred. The discarded cytoplasmic lobe (residual body) appeared to be degraded with the epithelium of the Wolffian duct. It was determined that the volume of the spermatid was reduced by 87% during spermiogenesis through a nuclear volume decrease of 76% and cytoplasmic volume decrease of 95.3%.     

Long-distance signal transmission and regulation of photosynthesis in characean cells

Photosynthetic electron transport in an intact cell is finely regulated by the structural flexibility of thylakoid membranes, existence of alternative electron-transport pathways, generation of electrochemical proton gradient, and continuous exchange of ions and metabolites between cell organelles and the cytoplasm. Long-distance interactions underlying reversible transitions of photosynthetic activity between uniform and spatially heterogeneous distributions are of particular interest. Microfluorometric studies of characean cells with the use of saturating light pulses and in combination with electrode micromethods revealed three mechanisms of distant regulation ensuring functional coordination of cell domains and signal transmission over long distances. These include: (1) circulation of electric currents between functionally distinct cell domains, (2) propagation of action potential along the cell length, and (3) continuous cyclical cytoplasmic streaming. This review considers how photosynthetic activity depends on membrane transport of protons and cytoplasmic pH, on ion fluxes associated with the electrical excitation of the plasmalemma, and on the transmission of photoinduced signals with streaming cytoplasm. Because of signal transmission with cytoplasmic flow, dynamic changes in photosynthetic activity can develop far from the point of photostimulus application and with a long delay (up to 100 s) after a light pulse stimulus is extinguished.     

Variations in ribulose 1,5-bisphosphate carboxylase protein levels,activities and subcellular distribution during photoautotrophic batch culture of Chlorogloeopsis fritschii

Ribulose 1,5-bisphosphate (RuBP) carboxylase is present in the cytoplasm and carboxysomes (polyhedral bodies) of the cyanobacterium Chlorogloeopsis fritschii. In vitro enzyme activities have been measured throughout photoautotrophic batch culture, together with RuBP carboxylase protein concentrations, determined by rocket immunoelectrophoresis. Enzyme activities and protein levels in the cytoplasmic and carboxysomal fractions varied in an apparently inverse manner during growth. The RuBP carboxylase activities per unit enzyme protein were maximal in late lag phase/early exponential phase for both cellular enzyme pools. Both rates per unit enzyme protein declined during exponential phase, cytoplasmic enzyme activity remaining consistently higher than that of the carboxysomal enzyme. Activities per unit cytoplasmic and carboxysomal enzyme protein showed very low, similar rates in late stationary phase and death phase. Dialysis experiments indicated that such changes were not due to interference in activity assays by soluble endogenous effectors. Major shifts in the subcellular distribution of RuBP carboxylase protein were found versus culture age, enzyme protein levels being predominantly carboxysomal in lag phase, mainly soluble in exponential phase and then mainly carboxysomal again in stationary/death phase. The data are discussed in terms of carboxysome function and the question of control of RuBP carboxylase synthesis in cyanobacteria.Abbreviations RuBP D-ribulose 1,5-bisphosphate - LTIB low Tris isolation buffer - HTIB high Tris isolation buffer - RIE rocket immunoelectrophoresis