Evidence for multiple sites of ferricyanide reduction in chloroplasts

Various sites of ferricyanide reduction were studied in spinach chloroplasts. It was found that in the presence of dibromothymoquinone a fraction of ferricyanide reduction was dibromothymoquinone sensitive, implying that ferricyanide can be reduced by photosystem I as well as photosystem II. To separate ferricyanide reduction sites in photosystem II, orthophenanthroline and dichlorophenyl dimethylurea inhibitions were compared at various pH's. It was noted that at low pH ferricyanide reduction was not completely inhibited by orthophenanthroline. At high pH's, however, inhibition of ferricyanide reduction by orthophenanthroline was complete. It was found that varying concentration of orthophenanthroline at a constant pH showed different degrees of inhibition. In the study of ferricyanide reduction by photosystem II various treatments affecting plastocyanin were performed. It was found that Tween-20 or KCN treatments which inactivated plastocyanin did not completely inactivate ferricyanide reduction. These data support the conclusion that ferricyanide accepts electrons both before and after plastoquinone in photosystem II.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyurea - MV methyl viologen - DBMIB 2,5-dibromothymoquinone - DMBQ 2,6-dimethyl benzoquinone - OP 1,10-orthophenanthroline - TMPD tetramethyl-p-phenylenediamine - PS 1 photosystem I - PS II photosystem II - SN sucrose-sodium chloride chloroplasts Supported by NSF Grant BMS 74-19689.     

Inhibition of energy-transducing functions of chloroplast membranes by lipophilic iron chelators

Lipophilic metal chelators inhibit various energy-transducing functions of chloroplasts. The following observations were made.1. Photophosphorylation coupled to any known mode of electron transfer, i.e. whole-chain noncyclic, the partial noncyclic Photosystem I or Photosystem II reactions, or cyclic, is inhibited by several lipophilic chelators, but not by hydrophilic chelators.2. The light- and dithioerythritol-dependent Mg²⁺-ATPase was also inhibited by the lipophilic chelators.3. Electron transport through either partial reaction, Photosystem I or Photosystem II was not inhibited by lipophilic chelators. Whole-chain coupled electron transport was inhibited by bathophenanthroline, and the inhibition was not reversed by uncouplers. The diketone chelators diphenyl propanedione and nonanedione inhibited the coupled, whole-chain electron transport and the inhibition was reversed by uncouplers, a pattern typical of energy transfer inhibitors.The electron transport inhibition site is localized in the region of plastoquinone → cytochrome f. This inhibition site is consistent with other recent work (Prince et al. (1975) FEBS Lett. 51, 108 and Malkin and Aparicio (1975) Biochem. Biophys. Res. Commun. 63, 1157) showing that a non-heme iron protein is present in chloroplasts having a redox potential near +290 mV. A likely position for such a component to function in electron transport would be between plastoquinone and cytochrome f, just where our data suggests there to be a functional metalloprotein.4. Some of the lipophilic chelators induce H⁺ leakiness in the chloroplast membrane, making interpretation of their phosphorylation inhibition difficult. However, 1–3 mM nonanedione does not induce significant H⁺ leakiness, while inhibiting ATP formation and the Mg²⁺-ATPase. Nonanedione, at those concentrations, causes a two- to four-fold increase in the extent of H⁺ uptake.5. These results are consistent with, but do not prove, the involvement of a non-heme iron or a metalloprotein in chloroplast energy transduction.     

Decrease of NADH in HeLa cells in the presence of transferrin or ferricyanide

The short-term incubation of HeLa cells in the presence of diferric transferrin or ferricyanide, which are reduced externally by the transplasma membrane reductase, produces a stoichiometric decrease in NADH and increase in NAD+, which is stimulated by insulin. The NADP/NADPH ratio does not change during 15 min incubation with the oxidants. The total pyridine nucleotide pool of HeLa cells is not affected. Incubation with apotransferrin and ferrocyanide, which cannot act as oxidants for transmembrane electron transport, does not change the pyridine nucleotide concentrations in the cells. Our results show that NADH can act as the internal electron donor for the reduction of external oxidants by the transmembrane reductase. It appears that oxidation of NADH by the transmembrane electron transport using ferricyanide or iron transferrin as external electron acceptors is sufficient to stimulate growth in HeLa cells.     

Diferric transferrin reduction stimulates the Na+/H+ antiport of HeLa cells

Proton release from HeLa cells is stimulated by external oxidants for the transplasmalemma electron transport enzymes. These oxidants, such as ferricyanide and diferric transferrin, also stimulate cell growth. We now present evidence that proton release associated with the reduction of ferricyanide and diferric transferrin is through the Na+/H+ antiport. The stoichiometry of H+/e- release with diferric transferrin is over 50 to 1, which is greater than expected for oxidation of a protonated transmembrane electron carrier. Diferric transferrin induced proton release depends on external sodium and is inhibited by amiloride. Proton release is also inhibited when diferric transferrin reduction is inhibited by apotransferrin. A tightly coupled association between the redox system and the antiport is shown by sodium dependence and amiloride inhibition of diferric transferrin reduction. The results indicate a new role for ferric transferrin in growth stimulation by activation of the sodium-proton antiport.     

Reconstitution into liposomes of glucose active transport from the rabbit renal proximal tubule. Characteristics of the system

This paper describes the characteristics of Na⁺-dependent d-glucose transport into liposomes made from soybean phospholipids into which have been reconstituted detergent-solubilized components from the rabbit renal proximal tubular brush border membrane. Conditions for optimal and quantitative reconstitution of glucose carriers are defined. Na⁺-dependent d-glucose uptake occurs via a saturable system with a $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ of 0.125–0.135 mM, is responsive to the volume of the internal liposomal space, and shows ‘overshoot’ as seen in natural membranes. The rate of Na⁺-dependent d-glucose uptake and the magnitude of the ‘overshoot’ are proportional to the concentration of protein used in reconstitution.     

Long‐term mammal and nocturnal bird trends are influenced by vegetation type,weather and climate in temperate woodlands

Many studies have documented the individual effects of variables such as vegetation, long‐term climate and short‐term weather on biodiversity. Few, however, have explicitly explored how interactions among these major drivers can influence species abundance. We used data from a 15‐year study (2002–2017) in the endangered temperate woodlands of south‐eastern Australia to test hypotheses associated with the effects of vegetation type, long‐term climate and short‐term weather on population trajectories of seven species of (largely) nocturnal mammals and birds. Despite prolonged drought conditions, there was a significant increase in the abundance of some species over time (e.g. the Eastern Grey Kangaroo). It is possible that destocking of domestic livestock may have reduced competition with Kangaroos, thereby facilitating increases in abundance. The Common Brushtail Possum and Common Ringtail Possum were significantly less likely to occur in replanted woodlands, possibly because of the paucity of nesting sites. We found no evidence that replanted woodlands are refuges for exotic pest species like the European Rabbit and Red Fox. Short‐ and long‐term rainfall and vegetation type had important independent and combined effects on animal abundance. That is, responses to periods of high short‐term rainfall were dependent on vegetation type and whether sites occurred in long‐term climatically wet versus climatically dry locations. For example, the Red Fox responded positively to high levels of short‐term rainfall, but only at climatically dry sites. Our results highlight the complementary value of different vegetation types across the landscape and the context‐specific responses of animals to short‐term fluctuations in moisture availability. They also underscore the value of long‐term monitoring at a landscape scale for examining how multiple interacting factors influence trends in animal abundance.