Regulation of chloroplast metabolism in leaves: Evidence that NADP-dependent glyceraldehydephosphate dehydrogenase,but not ferredoxin-NADP reductase,controls electron flow to phosphoglycerate in the dark-light transition

P₇₀₀ is rapidly, but only transiently photooxidized upon illuminating dark-adapted leaves. Initial oxidation is followed by a reductive phase even under far-red illumination which excites predominantly photosystem (PS) I. In this phase, oxidized P₇₀₀ is reduced by electrons coming from PSII. Charge separation in the reaction center of PSI is prevented by the unavailability of electron acceptors on the reducing side of PSI. It is subsequently made possible by the opening of an electron gate which is situated between PSI and the electron acceptor phosphoglycerate. Electron acceptors immediately available for reduction while the gate is closed corresponded to 10 nmol · (mg chlorophyll)^–1 electrons in geranium leaves, 16 nmol · (mg chlorophyll)^–1 in sunflower and 22 nmol · (mg chlorophyll)^–1 in oleander. Reduction of NADP during the initial phase of P₇₀₀ oxidation showed that the electron gate was not represented by ferredoxin-NADP reductase. Availability of ATP indicated that electron flow was not hindered by deactivation of the thylakoid ATP synthetase. It is concluded that NADP-dependent glyceraldehydephosphate dehydrogenase is completely deactivated in the dark and activated in the light. The rate of activation depends on the length of the preceding dark period. As chloroplasts contain both NAD- and NADP-dependent glyceraldehydephosphate dehydrogenases, deactivation of the NADP-dependent enzyme disconnects chloroplast NAD and NADP systems and prevents phosphoglycerate reduction in the dark at the expense of NADPH and ATP which are generated by glucose-6-phosphate oxidation and glycolytic starch breakdown, respectively.Abbreviations Chl chlorophyll - P₇₀₀ electron donor pigment in the reaction center of photosystem I Cooperation of the Institute of Botany of the University of Würzburg with the Institute of Astrophysics and Atmospheric Physics of the Estonian Academy of Sciences in Tartu was supported by the Deutsche Forschungsgemeinschaft and the Estonian Academy of Sciences. This work was performed within the Sonderforschungsbereich 251 of the University of Würzburg.     

Analysis of pollen-tube growth in cultured maize silks

Summary In vitro pollen-tube growth in maize was studied using an in vitro pollination system. In the cut-silk method, ovaries with silks were placed on medium in vitro, whereafter the silk was cut and the upper part of the silk was pollinated. Pollen tubes were not able to bridge the space between the two silk parts. Even when silk parts were tightly connected, pollen tubes still were not able to pass the cut ends and reach the lower silk part. Pollen-tube growth rates and the direction of tube growth were not influenced by the presence or absence of an ovary. Prepollination did not have any influence on pollen-tube growth rate. Measurements of pollen-tube growth rate also showed that there was no population effect, i.e. growth rate was not stimulated by pollination with an excess of pollen grains. We found that the direction in which maize pollen grew was determined only by the positioning of the silk hairs.     

Effects of selective cholecystokinin antagonists L364,718 and L365,260 on food intake in rats

The selective type A and B cholecystokinin (CCK) receptor antagonists L364,718 and L365,260 were used to identify the receptor subtype that mediates the satiety effect of endogenous CCK. Male rats (n = 12–13/group), fed ground rat chow ad lib, received L364,718 (0, 1, 10, 100, or 1000 μg/kg IP) or L365,260 (0, 0.1, 1, 10, 100, 1000, or 10,000 μg/kg IP) 2 h after lights off, and food intake was measured 1.5, 3.5, and 5.5 h later. L364,718 significantly stimulated 1.5-h food intake by more than 40% at 10 μg/kg and higher doses; cumulative intake at 3.5 and 5.5 h remained elevated by about 20% at 1000 and 100 μg/kg of L364,718, respectively. In contrast, L365,260 had no significant stimulatory effect on feeding at any dose. The potency of L365,260 for antagonizing gastrin-stimulated gastric acid secretion was examined in unanesthetized rats. Male rats (n = 14), prepared with gastric and jugular vein cannulas, received doubling doses of gastrin (G-17I) (0.16–5 nmol/kg/h IV), each dose for 30 min, and gastric juice was collected for each 30-min period. G-17I stimulated gastric acid output dose dependently; the minimal effective dose was 0.16 nmol/kg/h, while maximal output (5-fold above basal) occurred at 5 nmol/kg/h. L365,260 (0, 1, 10, 100, 1000, or 10,000 μg/kg IV), administered 30 min before continuous infusion of G-17I (1.25 or 5 nmol/kg/h), significantly inhibited acid output only at 10,000 μg/kg; cumulative 60-min output was decreased by 60%. These results suggest that CCK acts at CCK-A receptors to produce satiety during the dark period in ad lib-feeding rats.     

Purification and characterization of G proteins from human brain: modification of GTPase activity upon phosphorylation

Summary Three G proteins from human brain membranes were purified to near homogeneity by conventional techniques including preparative electrophoresis. These G proteins were characterized by their ability to bind GTP, GDP and GTP analogs. Two of these proteins have molecular weights of 50,000 (G₅₀) and 36,000 (G₃₆), as determined on SDS-gels. G₃₆ was ADP-ribosylated by pertussis toxin. Thus, G₅₀ could represent a G_sα subunit, whereas G₃₆ could be G_iα or G_oα. G₅₀ was phosphorylated by cAMP dependent protein kinase and protein kinase C. G₃₆ was phosphorylated by a protein kinase independent of calcium and phospholipid, a proteolytic product of protein kinase C, analogous to protein kinase M. Phosphorylation of G₃₆ by this protein kinase induced a dramatic decrease in its GTPase activity. The third G protein, of molecular weight 22,000 probably belongs to the group of monomeric G proteins possessing functional similarities withras gene products. The regulation of G proteins involving calcium-dependent and independent pathways is delineated.     

Water and solute regulation in Procephalothrix spiralis Coe and Clitellio arenarius (Müller) III. Long-term acclimation to diluted seawaters and effect of putative neuroendocrine structures

When acutely transferred to diluted seawater (SW), Procephalothrix spiralis and Clitellio arenarius regulate water content (g H₂O/g solute free dry wt = s.f.d.w.) via loss of Na and Cl (µmoles/g.s.f.d.w.). The present study extends these observations to a greater range of salinities and determines the effects of long-term, stepwise acclimation to diluted seawaters. Final exposure to a given experimental seawater (70, 50, 30, 15%) was 48 hours. Osmolality (mOsm/kg H₂O) and Na, K, and Cl ion concentrations (mEq/l) were determined in total tissue water and in the extracellular fluid of C. arenarius. Extracellular volume was determined as the ¹⁴C-polyethylene glycol space. Both species behaved as hyperosmotic conformers in diluted seawaters. However, reduction of the osmotic gradient between worm and medium occurred in P. spiralis, but not C. arenarius, in 30 and 15% SW. In both species, osmolality and Na, Cl, and K concentrations in total tissue water decreased with increased dilution of the SW. Water content increased with dilution of the medium but was lower than that which would be predicted based on approximation of the van't Hoff relation. This indicated the occurrence of regulatory volume decrease (RVD). In P. spiralis, in 70 or 50% SW, RVD was accompanied by loss of Na and Cl contents. However, in 30 or 15% SW, Na and Cl contents increased and in worms in 15% SW K content decreased. The latter movements of Na, Cl and K are indicative of cellular hysteresis and were associated with decreased viability, indicating the lower limits of regulatory ability in this species. In comparison, RVD in C. arenarius occurred in all diluted seawaters and was accompanied by loss of Na and Cl contents. In C. arenarius, evidence for reduced viability was absent. Removal of the supra- and subesophageal ganglia of C. arenarius resulted in retention of water, Na and Cl (g H₂O or µmoles/g s.f.d.w.) in worms acclimated to 70% SW. Removal of the cerebral ganglia and cephalic glands of P. spiralis did not significantly influence regulation of water content.     

Experimental manipulations of the phytoplankton periodicity in large limnetic enclosures in Blelham Tarn,English Lake District

This paper reviews the results of experimental manipulations, carried out during the period 1977–1983, on the phytoplankton maintained in the limnetic enclosures at Blelham Tarn, English Lake District. Three categories of manipulations are considered.The effects of variation in the scale and frequency of phosphorus loading (range: 0.3 to 2.5 g P m^–2 a^–1) upon the mean phytoplankton biomass, its seasonal distribution and specific dominance are shown to conform to well-established patterns and relationships observed in natural lakes. Much of the seasonal variability in species dominance occurred independently of nutrient ratios, though carbon availability has been critical at times. Attempts to manipulate the rates of removal of phytoplankton by grazing have confirmed that they act selectively against certain smaller species only, that they alter the rate of successional change, rather than its direction, and that they have little lasting influence upon the total phytoplankton standing crop. Attempts to manipulate rates of sinking loss through artificial enlargement of the epilimnetic circulation also regulated the light-conditions experienced by suspended phytoplankton.Growth-rate relationships to an index of light exposure and to temperature fluctuation are also derived for several species and are related to morphological and physiological characters of the organisms concerned. These interpretations are briefly reviewed in relation to periodic cycles in natural lakes.     

Separate,Ca2+-activated K+ and Cl− transport pathways in Ehrlich ascites tumor cells

Summary The net loss of KCl observed in Ehrlich ascites cells during regulatory volume decrease (RVD) following hypotonic exposure involves activation of separate conductive K⁺ and Cl^– transport pathways. RVD is accelerated when a parallel K⁺ transport pathway is provided by addition of gramicidin, indicating that the K⁺ conductance is rate limiting. Addition of ionophore A23187 plus Ca²⁺ also activates separate K⁺ and Cl^– transport pathways, resulting in a hyperpolarization of the cell membrane. A calculation shows that the K⁺ and Cl^– conductance is increased 14-and 10-fold, respectively. Gramicidin fails to accelerate the A23187-induced cell shrinkage, indicating that the Cl^– conductance is rate limiting. An A23187-induced activation of⁴²K and³⁶Cl tracer fluxes is directly demonstrated. RVD and the A23187-induced cell shrinkage both are: (i) inhibited by quinine which blocks the Ca²⁺-activated K⁺ channel. (ii) unaffected by substitution of NO ₃ ^– or SCN^– for Cl^–, and (iii) inhibited by the anti-calmodulin drug pimozide. When the K⁺ channel is blocked by quinine but bypassed by addition of gramicidin, the rate of cell shrinkage can be used to monitor the Cl^– conductance. The Cl^– conductance is increased about 60-fold during RVD. The volume-induced activation of the Cl^– transport pathway is transient, with inactivation within about 10 min. The activation induced by ionophore A23187 in Ca²⁺-free media (probably by release of Ca²⁺ from internal stores) is also transient, whereas the activation is persistent in Ca²⁺-containing media. In the latter case, addition of excess EGTA is followed by inactivation of the Cl^– transport pathway. These findings suggest that a transient increase in free cytosolic Ca²⁺ may account for the transient activation of the Cl^– transport pathway. The activated anion transport pathway is unselective, carrying both Cl^–, Br^–, NO ₃ ^– , and SCN^–. The anti-calmodulin drug pimozide blocks the volume- or A23187-induced Cl^– transport pathway and also blocks the activation of the K⁺ transport pathway. This is demonstrated directly by⁴²K flux experiments and indirectly in media where the dominating anion (SCN^–) has a high ground permeability. A comparison of the A23187-induced K⁺ conductance estimated from⁴²K flux measurements at high external K⁺, and from net K^– flux measurements suggests single-file behavior of the Ca²⁺-activated K⁺ channel. The number of Ca²⁺-activated K⁺ channels is estimated at about 100 per cell.