Charles K. Pallaghy

The Effect of Sub-Lethal Concentrations of Mercury and Zinc on Chlorella: II. Photosynthesis and Pigment Composition

Summary The effects of either 1 mM ZnCl 2 , 1 μm HgCl 2 or 0.1 μM phenyl mercuric acetate solutions on photosynthesis, pigment composition and respiration are described for cultures of Chlorella . Although photosynthesis and respiration are initially inhibited in young cultures, there is a recovery towards control values as the cultures proceed in their growth. In young cultures the inhibition in photosynthesis is associated with a loss in chlorophyll. In cultures grown for several days in metal solutions, photosynthesis per unit mass of chlorophyll may exceed control values - in phenyl mercuric acetate solutions in particular. Mercury solutions markedly enhance protochlorophyll levels in young cultures, while decreasing chlorophyll a/b and pheophytin a/b ratios. These observations are interpreted in terms of the action of metals on the oxidative and reductive steps in the biosynthetic pathway of pigments.

The Effect of Sub-Lethal Concentrations of Mercury and Zinc on Chlorella I. Growth Characteristics and Uptake of Metals

Summary The effects of either 1 mM ZnCl 2 , 1 μM HgCl 2 or 0.1 μM phenylmercuric acetate solutions on a variety of growth parameters are described for cultures of the Emerson strain of Chlorella . A significant finding is the comparative gain in the net carbon balance of heavy metal treated cells due to the inhibition of export of carbon compounds such as glycollate from the cells. There is also a stimulation of RNA, DNA and protein levels in HgCl 2 treated cells. After an initial rapid uptake of mercury and zinc by cells, it appears that the daughter cells are able to regulate and prevent the further entry of these heavy metals into the biomass.

The Effect of Sub-Lethal Concentrations of Mercury and Zinc on Chlorella: III. Development and Possible Mechanisms of Resistance to Metals

Summary Chlorella cultures maintained continuously in replenished solutions of heavy metals (1 mM ZnCl 2 , 1 μM HgCl 2 or 0.1 μm phenyl mercuric acetate) initially exhibit symptoms of metal toxicity, which are characterised by a sharp reduction in the pigment content and in the rates of cell division and metabolic activity. There is a concurrent gain in the net carbon balance of the culture due to the inhibition of export of photosynthates from the cells by heavy metals ( De Filippis and Pallaghy, 1976a , De Filippis and Pallaghy, 1976b ). After a period of 40 days (approximating 40 series of cell division) the cultures develop resistance and almost fully regain the rates of cell division displayed by control cultures. In attempting to elucidate the mechanism of resistance, we studied the fate of the heavy metals in cultures of sensitive and resistant cells. Evidence from 65 Zn uptake studies indicate that resistance is accompanied by an inhibition of the temperature sensitive component of zinc uptake and by a reduction in the number of exchange sites available for zinc in the free space (cell walls) per unit dry weight of the cells. Tolerance is therefore characterised by the development of a typical exclusion mechanism. Mercury resistance is primarily associated with an increased capacity for resistant cells to volatilize mercury out of solution by enzymic means. The ethylene generated by metal stressed cultures (up to 1.3 pmol/mg dry wt/hr) is sufficient to account for approximately 25%, and a mercury-reducing enzyme system for the remainder of the mercury volatilized.

Metabolic aspects of stomatal opening and ion accumulation by guard cells in Vicia faba

Summary Double labelling experiments employing 42K and 36Cl (using 10 mM KCl and 0.1 mM CaCl2 solutions) have shown that guard cells in floating epidermal strips of Vicia faba accumulate approximately 3 K ions for every Cl ion during stomatal opening. Light or CO2-free air stimulated stomatal opening and the uptake of 42K and 36Cl by the guard cells. These results suggest organic acid anions (approximately 100 mM if divalent) acting as counter ions for K. DCMU and FCCP inhibited stomatal opening, and 42K and 36Cl accumulation in similar proportions so that the K content of guard cells almost always reflected the extent of stomatal opening. The effects of other inhibitors are also discussed.

Unerwartete Kinetik des Efflux' und der Aufnahme von Ionen bei verschiedenen Pflanzengeweben

Summary A discontinuity of the ion efflux curve similar to that described by Pallaghy et al. (1970) for short maize root segments is also observed with more homogeneous materials such as root cortex cells, Mnium leaflets and others. The time course of 42K+ uptake by aged segments of isolated root cortex tissue also shows a discontinuity. The shoulder of the efflux curve cannot, therefore, be due to two overlapping efflux processes, i. e. via the cortex and the xylem respectively of the root segments. Although, at this stage, no model explaining all experimental findings can be presented, it seems clear that the three-compartmental model free space-cytoplasm-vacuole in series does not apply under all circumstances.

PRESENCE AND DISTRIBUTION OF BROMINE IN THYSANOCLADIA DENSA (SOLIERIACEAE, GIGARTINALES), A MARINE RED ALGA FROM THE GREAT BARRIER REEF1

Electron‐probe X‐ray microanalysis of freeze dried sections of growing branch tips of Thysanocladia densa Sander shows high concentrations of bromine in the region of the medulla. Indirect evidence infers that the Br is associated with intercellular, granular deposits which accumulate within the mucilage of the medulla. The concentration of Br decreases in older regions of the thalli.

X-ray microanalysis of calcium in fixed and in shock-frozen hydrated green algal cells: Mougeotia, Spirogyra and Zygnema

Abstract X-ray microanalysis, widely used for ionic detection in cell tissues, was performed on 0.5 μm thick glutaraldehyde-osmiumtetroxide fixed cell sections. Calcium in a water-insoluble state was found in membrane bound vesicles of about 0.5 μm in diameter. No other cell compartment showed a comparable calcium accumulation. In vivo staining by the fluorescent calcium indicator dye chlorotetracycline revealed a vesicular calcium pattern in Mougeotia matching the results by X-ray microanalysis: a dense population of calcium vesicles is located close to the chloroplast edge where the actin filaments are attached. The data find further support from X-ray microanalysis of shock-frozen hydrated Mougeotia cells and of the closely related green algal cells Spirogyra and Zygnema (Zygnemaceae). X-ray microanalysis as performed in this study seems a reliable tool for the detection of water-insoluble calcium stored by membrane-covered cytoplasmic compartments.

Rapid and economical multiple base site-directed mutagenesis of double stranded plasmid DNA

This method for the mutagenesis of ds-DNA, utilizing the best features of previously published protocols, incorporates a fragmentation procedure on the plasmid, thermostable enzymes and two transformations in E. coli. Screening of positive clones can begin after about two days. Insertions, deletions and substitutions of up to 50 bp are routinely obtained with 90-95% of clones positive as proven by sequencing. The cost is about one half to one third of equiv-alent commercial procedures.