Lipoxygenase activity and lipid composition of cotyledons and oil bodies of two sunflower hybrids

Lipoxygenase (linoleate: oxygen oxidoreductase, EC 1.13.11.12) and lipid fatty acid composition of cotyledons and oil bodies of two near-isogenic high and low oleic acid sunflower hybrids (Helianthus annuus L.) were investigated during seedling growth. Triacylglycerol fatty acid content decreased strongly in cotyledons of both hybrids between days 1 and 7 of seedling growth. Oil bodies contained mostly neutral lipids and small amounts of phospholipids and proteins. Between days 1 and 7 of germination the degradation rate of oil body neutral lipids was lower for the high oleic acid hybrid compared to the low oleic acid hybrid. From days 0 to 9 of germination, neutral lipids and phospholipids of cotyledons and oil bodies from the high oleic acid hybrid contained more oleic acid and less linoleic acid than those of the low oleic acid hybrid. The presence of an oil body lipoxygenase was shown enzymatically and by immunodetection. In both hybrids, lipoxygenase activity of oil bodies and whole cotyledons was lower in 7 than in 1 day-old seedlings. Moreover, the high oleic acid hybrid showed a lower oil body lipoxygenase activity than the low oleic acid hybrid, both at 1 and 7 days of germination. Immunoblot analysis indicated that differences in enzyme activity between the two hybrids were not due to changes in the amount of antigenic protein. Oleic acid was found to be a competitive inhibitor of oil body lipoxygenase. Based on these results it is concluded that oleic acid could act as a natural inhibitor of sunflower oil body lipoxygenase.This work was supported by Spanish DGICYT (Grant PB 94-0118-CO2-01)Peer Reviewe

Chloroplast Fe(III) chelate reductase activity is essential for seedling viability under iron limiting conditions

Photosynthesis, heme biosynthesis, and Fe-S cluster assembly all take place in the chloroplast, and all require iron. Reduction of iron via a membrane-bound Fe(III) chelate reductase is required before iron transport across membranes in a variety of systems, but to date there has been no definitive genetic proof that chloroplasts have such a reduction system. Here we report that one of the eight members of the Arabidopsis ferric reductase oxidase (FRO) family, FRO7, localizes to the chloroplast. Chloroplasts prepared from fro7 loss-of-function mutants have 75% less Fe(III) chelate reductase activity and contain 33% less iron per microgram of chlorophyll than wild-type chloroplasts. This decreased iron content is presumably responsible for the observed defects in photosynthetic electron transport. When germinated in alkaline soil, fro7 seedlings show severe chlorosis and die without setting seed unless watered with high levels of soluble iron. Overall, our results provide molecular evidence that FRO7 plays a role in chloroplast iron acquisition and is required for efficient photosynthesis in young seedlings and for survival under iron-limiting conditions

From geography to genes: evolutionary perspectives on salinity tolerance in the brackish water barnacle Balanus improvisus

How species respond to changes in their environment is a fundamental question in biology. This has become an increasingly important issue as anthropogenic effects of climate change and biological invasions have major impacts on marine ecosystems worldwide. In this thesis I investigated the role of salinity tolerance from an evolutionary perspective, using a wide range of techniques, spanning from population genetics and common-garden experiments to characterizing potential genes involved in osmoregulation in barnacles. I used the acorn barnacle species Balanus (Amphibalanus) improvisus, which displays a remarkably broad salinity tolerance, to investigate how this trait has influenced the species' potential to establish in new environments, and respond to projected near-future salinity reductions in coastal seas. I also examined physiological and molecular mechanisms that may be involved in osmoregulation in B. improvisus. I further analysed population genetic structure using microsatellites and mitochondrial DNA, and related the results to anthropogenic and natural dispersal dynamics on both global and regional (Baltic Sea) scales. I found high genetic diversity in most populations, with many shared haplotypes between distant populations. This supports the hypothesis that maritime shipping is an important vector for the dispersal of the cosmopolitan species B. improvisus. Nonetheless, natural larval dispersal is also important on smaller geographical scales, such as within the Baltic Sea. Marked genetic differentiation between northern and southern Baltic Sea populations raises the question whether there is restricted gene flow within the Baltic Sea, creating potential for local adaptations to evolve. To investigate the extent to which the broad distribution of B. improvisus along the Baltic Sea salinity gradient is explained by local adaptation versus physiological plasticity, I performed a common-garden experiment in which multiple populations were exposed to different salinities and multiple fitness-related phenotypic traits were recorded. The experiment confirmed that phenotypic plasticity, rather than local adaptation, explained the broad distribution of the species along the salinity gradient. Interestingly, all populations of B. improvisus performed best at low and intermediate salinities in many fitness-related traits (survival, growth and reproduction), although other traits (e.g. shell strength an juvenile growth) indicated higher costs associated with low salinity. A candidate gene approach was used to investigate the molecular basis of broad salinity tolerance in B. improvisus by characterizing the Na+/K+ ATPase (NAK) of B. improvisus – an ion transporter commonly involved in active osmoregulation in many species. We identified two main gene variants in B. improvisus (NAK1 and NAK2), and found that NAK1 mRNA existed in two isoforms that were differentially expressed in different life stages and adult tissues, suggesting an active role in osmoregulation. Lastly, I summarise current knowledge about salinity tolerance in barnacles and outline new research directions to further our understanding of the physiological and molecular mechanisms involved in salinity tolerance in barnacles

Chelation by histidine inhibits the vacuolar sequestration of nickel in roots of the hyperaccumulator Thlaspi caerulescens

* The mechanisms of enhanced root to shoot metal transport in heavy metal hyperaccumulators are incompletely understood. Here, we compared the distribution of nickel (Ni) over root segments and tissues in the hyperaccumulator Thlaspi caerulescens and the nonhyperaccumulator Thlaspi arvense, and investigated the role of free histidine in Ni xylem loading and Ni transport across the tonoplast. * Nickel accumulation in mature cortical root cells was apparent in T. arvense and in a high-Ni-accumulating T. caerulescens accession, but not in a low-accumulating T. caerulescens accession. * Compared with T. arvense, the concentration of free histidine in T. caerulescens was 10-fold enhanced in roots, but was only slightly higher in leaves, regardless of Ni exposure. Nickel uptake in MgATP-energized root- and shoot-derived tonoplast vesicles was almost completely blocked in T. caerulescens when Ni was supplied as a 1 : 1 Ni-histidine complex, but was uninhibited in T. arvense. Exogenous histidine supply enhanced Ni xylem loading in T. caerulescens but not in T. arvense. * The high rate of root to shoot translocation of Ni in T. caerulescens compared with T. arvense seems to depend on the combination of two distinct characters, that is, a greatly enhanced root histidine concentration and a strongly decreased ability to accumulate histidine-bound Ni in root cell vacuoles