Todd N. Rosenstiel

Increased CO2 uncouples growth from isoprene emission in an agriforest ecosystem.

The emission of isoprene from the leaves of forest trees is a fundamental component of biosphere–atmosphere interactions, controlling many aspects of photochemistry in the lower atmosphere. As almost all commercial agriforest species emit high levels of isoprene, proliferation of agriforest plantations has significant potential to increase regional ozone pollution and enhance the lifetime of methane, an important determinant of global climate. Here we show that growth of an intact Populus deltoides plantation under increased CO2 (800 µmol mol-1 and 1,200 µmol mol-1) reduced ecosystem isoprene production by 21% and 41%, while above-ground biomass accumulation was enhanced by 60% and 82%, respectively. Exposure to increased CO2 significantly reduced the cellular content of dimethylallyl diphosphate, the substrate for isoprene synthesis, in both leaves and leaf protoplasts. We identify intracellular metabolic competition for phosphoenolpyruvate as a possible control point in explaining the suppression of isoprene emission under increased CO2. Our results highlight the potential for uncoupling isoprene emission from biomass accumulation in an agriforest species, and show that negative air-quality effects of proliferating agriforests may be offset by increases in CO2.

Isoprene emission from terrestrial ecosystems in response to global change: minding the gap between models and observations

Coupled surface–atmosphere models are being used with increased frequency to make predictions of tropospheric chemistry on a ‘future’ earth characterized by a warmer climate and elevated atmospheric CO2 concentration. One of the key inputs to these models is the emission of isoprene from forest ecosystems. Most models in current use rely on a scheme by which global change is coupled to changes in terrestrial net primary productivity (NPP) which, in turn, is coupled to changes in the magnitude of isoprene emissions. In this study, we conducted measurements of isoprene emissions at three prominent global change experiments in the United States. Our results showed that growth in an atmosphere of elevated CO2 inhibited the emission of isoprene at levels that completely compensate for possible increases in emission due to increases in aboveground NPP. Exposure to a prolonged drought caused leaves to increase their isoprene emissions despite reductions in photosynthesis, and presumably NPP. Thus, the current generation of models intended to predict the response of isoprene emission to future global change probably contain large errors. A framework is offered as a foundation for constructing new isoprene emission models based on the responses of leaf biochemistry to future climate change and elevated atmospheric CO2 concentrations.

Dependence of photosynthesis and energy dissipation activity upon growth form and light environment during the winter

Two very distinctive responses of photosynthesis to winter conditions have been identified. Mesophytic species that continue to exhibit growth during the winter typically exhibit higher maximal rates of photosynthesis during the winter or when grown at lower temperatures compared to individuals examined during the summer or when grown at warmer temperatures. In contrast, sclerophytic evergreen species growing in sun-exposed sites typically exhibit lower maximal rates of photosynthesis in the winter compared to the summer. On the other hand, shaded individuals of those same sclerophytic evergreen species exhibit similar or higher maximal rates of photosynthesis in the winter compared to the summer. Employment of the xanthophyll cycle in photoprotective energy dissipation exhibits similar characteristics in the two groups of plants (mesophytes and shade leaves of sclerophytic evergreens) that exhibit upregulation of photosynthesis during the winter. In both, zeaxanthin + antheraxanthin (Z + A) are retained and PS II remains primed for energy dissipation only on nights with subfreezing temperatures, and this becomes rapidly reversed upon exposure to increased temperatures. In contrast, Z + A are retained and PS II remains primed for energy dissipation over prolonged periods during the winter in sun leaves of sclerophytic evergreen species, and requires days of warming to become fully reversed. The rapid disengagement of this energy dissipation process in the mesophytes and shade sclerophytes apparently permits a rapid return to efficient photosynthesis and increased activity on warmer days during the winter. This may be associated with a decreasing opportunity for photosynthesis in source leaves relative to the demand for photosynthesis in the plants sinks. In contrast, the sun-exposed sclerophytes – with a relatively high source to sink ratio – maintain PS II in a state primed for high levels of energy dissipation activity throughout much of the winter. Independent of whether photosynthesis was up- or downregulated, all species under all conditions exhibited higher levels of soluble carbohydrates during the winter compared to the summer. Thus downregulation of photosynthesis and of Photosystem II do not appear to limit carbohydrate accumulation under winter conditions. A possible signal communicating an altered source/sink balance, or that may be influencing the engagement of Z + A in energy dissipation, is phosphorylation of thylakoid proteins such as D1.

Effect of nitrogen limitation on foliar antioxidants in relationship to other metabolic characteristics.

Abstract. The long-term effect of limiting soil nitrogen (N) availability on foliar antioxidants, thermal energy dissipation, photosynthetic and respiratory electron transport, and carbohydrates was investigated in Spinacia oleracea L. Starch, sucrose, and glucose accumulated in leaves of N-limited spinach at predawn, consistent with a downregulation of chloroplast processes by whole-plant sink limitation in response to a limited supply of N-based macromolecules throughout the plant. On a leaf-area or dry-weight basis, levels of chlorophyll, carotenoid pools, photosynthetic electron transport capacity, as well as activities for the predominantly chloroplast-localized antioxidant enzymes ascorbate peroxidase (EC 1.11.1.11) and glutathione reductase (EC 1.6.4.2) were much lower in N-limited versus N-replete plants. When expressed on a chlorophyll basis, foliar levels of all of these parameters were similar in N-replete versus N-limited plants. However, on a total-protein basis, antioxidant enzyme activities were higher in N-limited plants. Nitrogen-limited spinach showed higher levels of thermal energy dissipation and of zeaxanthin and antheraxanthin at midday, as well as slightly higher ascorbate contents relative to chlorophyll. These results indicate that strong, long-term N limitation led not only to alterations in the balance between different processes but also to an overall downregulation of light collection, photosynthetic electron transport capacity, and chloroplast-based antioxidant enzymes. This is further supported by the finding that glucose-feeding of excised leaves led to strong concomitant decreases in photosynthetic electron transport capacity and ascorbate peroxidase activity. On a leaf-area basis, neither superoxide dismutase (EC 1.15.1.1) activity nor dark repiration rates showed a treatment effect. This indicates that overall mitochondrial electron transport activity does not decrease under long-term N limitation and is consistent with localization of an important fraction of foliar superoxide dismutase in mitochondria.

On-line Analysis of the 13CO2 Labeling of Leaf Isoprene Suggests Multiple Subcellular Origins of Isoprene Precursors.

Abstract. Isoprene (2-methyl-1,3-butadiene) is the most abundant biogenic hydrocarbon released from vegetation, and there is continuing interest in understanding its biosynthesis from photosynthetic precursors in leaf chloroplasts. We used on-line proton-transfer-reaction mass spectrometry (PTR–MS) to observe the kinetics of 13C-labeling of isoprene following exposure to 13CO2 and then the loss of 13C after a return to normal 12CO2 in oak (Quercus agrifolia Nee) and cottonwood (Populus deltoides Barr.) leaves. Assignments of labeled isoprene species were verified by gas chromatography–mass spectrometry. For the first time, it was possible to observe the half-lives of individually 13C-labeled isoprene species during these transitions, and to trace some of the label to a C3 fragment that contained the two isoprene carbons derived from pyruvate via the deoxyxylulose-5-phosphate (DOXP) pathway. At steady state (under 13CO2), approximately 80% of isoprene carbon was labeled, with fully labeled isoprene as the major species (approx. 60%). The source of the unlabeled C is suggested to be extrachloroplastic, but not from photorespiratory carbon. After a transfer to 12CO2, 13C-labeling persisted in one isoprene carbon for several hours; this persistence was much more pronounced in (i) leaves inhibited by fosmidomycin, a specific inhibitor of the DOXP pathway, and (ii) in sun leaves which have higher ratios of soluble sugars to starch. From the mass 41–44 fragment data, and labeling predicted from the DOXP pathway in chloroplasts, precursors may arise from cytosolic pyruvate/phosphoenolpyruvate equivalents transported into the chloroplast; this idea was supported by an indirect measure of pyruvate labeling. Other sources of cytosolic isoprene precursors (i.e. dimethylallyl diphosphate or pentose phosphate) could not be excluded. The data obtained shed light on the half-lives of photosynthetic metabolites, exchanges of carbon between cellular pools, and suggest multiple origins of isoprene precursors in leaves.

Differential Accumulation of Dimethylallyl Diphosphate in Leaves and Needles of Isoprene- and Methylbutenol-Emitting and Nonemitting Species

The biosynthesis and emission of volatile plant terpenoids, such as isoprene and methylbutenol (MBO), depend on the chloroplastic production of dimethylallyl diphosphate (DMAPP). To date, it has been difficult to study the relationship of cellular DMAPP levels to emission of these volatiles because of the lack of a sensitive assay for DMAPP in plant tissues. Using a recent DMAPP assay developed in our laboratories, we report that species with the highest potential for isoprene and MBO production also exhibit elevated light-dependent DMAPP production, ranging from 110% to 1,063%. Even species that do not produce significant amounts of volatile terpenoids, however, exhibit some potential for light-dependent production of DMAPP. We used a nonaqueous fractionation technique to determine the intracellular distribution of DMAPP in isoprene-emitting cottonwood (Populus deltoides) leaves; approximately 65% to 70% of the DMAPP recovered at midday occurred in the chloroplasts, indicating that most of the light-dependent production of DMAPP was chloroplastic in origin. The midday concentration of chloroplastic DMAPP in cottonwood leaves is estimated to be 0.13 to 3.0 mm, which is consistent with the relatively high K ms that have been reported for isoprene synthases (0.5–8 mm). The results provide support for the hypothesis that the light dependence of isoprene and MBO emissions is in part due to controls over DMAPP production.

Why only some plants emit isoprene

Isoprene (2-methyl-1,3-butadiene) is emitted from many plants and it appears to have an adaptive role in protecting leaves from abiotic stress. However, only some species emit isoprene. Isoprene emission has appeared and been lost many times independently during the evolution of plants. As an example, our phylogenetic analysis shows that isoprene emission is likely ancestral within the family Fabaceae (= Leguminosae), but that it has been lost at least 16 times and secondarily gained at least 10 times through independent evolutionary events. Within the division Pteridophyta (ferns), we conservatively estimate that isoprene emissions have been gained five times and lost two times through independent evolutionary events. Within the genus Quercus (oaks), isoprene emissions have been lost from one clade, but replaced by a novel type of light-dependent monoterpene emissions that uses the same metabolic pathways and substrates as isoprene emissions. This novel type of monoterpene emissions has appeared at least twice independently within Quercus, and has been lost from 9% of the individuals within a single population of Quercus suber. Gain and loss of gene function for isoprene synthase is possible through relatively few mutations. Thus, this trait appears frequently in lineages; but, once it appears, the time available for evolutionary radiation into environments that select for the trait is short relative to the time required for mutations capable of producing a non-functional isoprene synthase gene. The high frequency of gains and losses of the trait and its heterogeneous taxonomic distribution in plants may be explained by the relatively few mutations necessary to produce or lose the isoprene synthase gene combined with the assumption that isoprene emission is advantageous in a narrow range of environments and phenotypes.

Evidence of reduced arbuscular mycorrhizal fungal colonization in multiple lines of Bt maize

PREMISE OF THE STUDY Insect-resistant Bacillus thuringiensis (Bt) maize is widely cultivated, yet few studies have examined the interaction of symbiotic arbuscular mycorrhizal fungi (AMF) with different lines of Bt maize. As obligate symbionts, AMF may be sensitive to genetic changes within a plant host. Previous evaluations of the impact of Bt crops on AMF have been inconsistent, and because most studies were conducted under disparate experimental conditions, the results are difficult to compare. METHODS We evaluate AMF colonization in nine Bt maize lines, differing in number and type of engineered trait, and five corresponding near-isogenic parental (P) base hybrids in greenhouse microcosms. Plants were grown in 50% local agricultural soil with low levels of fertilization, and AMF colonization was evaluated at 60 and 100 d. Nontarget effects of Bt cultivation on AMF colonization were tested in a subsequently planted crop, Glycine max, which was seeded into soil that had been preconditioned for 60 d with Bt or P maize. KEY RESULTS We found that Bt maize had lower levels of AMF colonization in their roots than did the non-Bt parental lines. However, reductions in AMF colonization were not related to the expression of a particular Bt protein. There was no difference in AMF colonization in G. max grown in the Bt- or P-preconditioned soil. CONCLUSIONS These findings are the first demonstration of a reduction in AMF colonization in multiple Bt maize lines grown under the same experimental conditions and contribute to the growing body of knowledge examining the unanticipated effects of Bt crop cultivation on nontarget soil organisms.

Sex-specific volatile compounds influence microarthropod-mediated fertilization of moss

Sexual reproduction in non-vascular plants requires unicellular free-motile sperm to travel from male to female reproductive structures across the terrestrial landscape. Recent data suggest that microarthropods can disperse sperm in mosses. However, little is known about the chemical communication, if any, that is involved in this interaction or the relative importance of microarthropod dispersal compared to abiotic dispersal agents in mosses. Here we show that tissues of the cosmopolitan moss Ceratodon purpureus emit complex volatile scents, similar in chemical diversity to those described in pollination mutualisms between flowering plants and insects, that the chemical composition of C. purpureus volatiles are sex-specific, and that moss-dwelling microarthropods are differentially attracted to these sex-specific moss volatile cues. Furthermore, using experimental microcosms, we show that microarthropods significantly increase moss fertilization rates, even in the presence of water spray, highlighting the important role of microarthropod dispersal in contributing to moss mating success. Taken together, our results indicate the presence of a scent-based ‘plant–pollinator-like’ relationship that has evolved between two of Earth’s most ancient terrestrial lineages, mosses and microarthropods.

Field Evaluation of Arbuscular Mycorrhizal Fungal Colonization in Bacillus thuringiensis Toxin-Expressing (Bt) and Non-Bt Maize

ABSTRACT The cultivation of genetically engineered Bacillus thuringiensis toxin-expressing (Bt) maize continues to increase worldwide, yet the effects of Bt crops on arbuscular mycorrhizal fungi (AMF) in soil are poorly understood. In this field experiment, we investigated the impact of seven different genotypes of Bt maize and five corresponding non-Bt parental cultivars on AMF and evaluated plant growth responses at three different physiological time points. Plants were harvested 60 days (active growth), 90 days (tasseling and starting to produce ears), and 130 days (maturity) after sowing, and data on plant growth responses and percent AMF colonization of roots at each harvest were collected. Spore abundance and diversity were also evaluated at the beginning and end of the field season to determine whether the cultivation of Bt maize had a negative effect on AMF propagules in the soil. Plant growth and AMF colonization did not differ between Bt and non-Bt maize at any harvest period, but AMF colonization was positively correlated with leaf chlorophyll content at the 130-day harvest. Cultivation of Bt maize had no effect on spore abundance and diversity in Bt versus non-Bt plots over one field season. Plot had the most significant effect on total spore counts, indicating spatial heterogeneity in the field. Although previous greenhouse studies demonstrated that AMF colonization was lower in some Bt maize lines, our field study did not yield the same results, suggesting that the cultivation of Bt maize may not have an impact on AMF in the soil ecosystem under field conditions.