Genetic relatedness can alter the strength of plant-soil interactions.

PREMISE: Intraspecific variation may play a key role in shaping the relationships between plants and their interactions with soil microbial communities. The soil microbes of individual plants can generate intraspecific variation in the responsiveness of the plant offspring, yet have been much less studied. To address this need, we explored how the relatedness of seedlings from established clones of Solidago altissima altered the plant-soil interactions of the seedlings. METHODS: Seedlings of known parentage were generated from a series of 24 clones grown in a common garden. Seedlings from these crosses were inoculated with soils from maternal, paternal, or unrelated clones and their performance compared to sterilized control inocula. RESULTS: We found that soil inocula influenced by S. altissima clones had an overall negative effect on seedling biomass. Furthermore, seedlings inoculated with maternal or paternal soils tended to experience larger negative effects than seedlings inoculated with unrelated soils. However, there was much variation among individual crosses, with not all responding to relatedness. CONCLUSIONS: Our data argue that genetic relatedness to the plant from which the soil microbial inoculum was obtained may cause differential impacts on establishing seedlings, encouraging the regeneration of non-kin adjacent to established clones. Such intraspecific variation represents a potentially important source of heterogeneity in plant-soil microbe interactions with implications for maintaining population genetic diversity.

Contribution of fungal and invertebrate communities to wood decay in tropical terrestrial and aquatic habitats.

Wood is a major carbon input into aquatic ecosystems and is thought to decay slowly, yet surprisingly little terrestrial carbon accumulates in marine sediments. A better mechanistic understanding of how habitat conditions and decomposer communities influence wood decay processes along the river-estuary-ocean continuum can address this seeming paradox. We measured mass loss, wood element, and polymer concentrations, quantified invertebrate-induced decay, and sequenced fungal communities associated with replicate sections of Guazuma branch wood submerged in freshwater, estuarine, and near-shore marine habitats and placed on the soil surface in nearby terrestrial habitats in three watersheds in the tropical eastern Pacific. Over 15 months, we found that wood decayed at similar rates in estuarine, marine, and terrestrial sites, reflecting the combined activity of invertebrate and microbial decomposers. In contrast, in the absence of shipworms (Teredinidae), which accounted for ~40% of wood mass loss in the estuarine habitats, decay proceeded more slowly in freshwater. Over the experiment, wood element chemistry diverged among freshwater, estuarine, and marine habitats, due to differences in both nutrient losses (e.g., potassium and phosphorus) and gains (e.g., calcium and aluminum) through decay. Similarly, we observed changes in wood polymer content, with the highest losses of cellulose, hemicellulose, and lignin moieties in the marine habitat. Aquatic fungal communities were strongly dominated by ascomycetes (88-99% of taxa), compared to terrestrial communities (55% ascomycetes). Large differences in fungal diversity were also observed across habitats with threefold higher richness in terrestrial than freshwater habitats and twofold higher diversity in freshwater than estuarine/marine habitats. Divergent decay trajectories across habitats were associated with widespread order-level differences in fungal composition, with distinct communities found in freshwater, estuarine and marine habitats. However, few individual taxa that were significantly associated with mass loss were broadly distributed, suggesting a high level of functional redundancy. The rapid processing of wood entering tropical rivers by microbes and invertebrates, comparable to that on land, indicates that estuaries and coastal oceans are hotspots not just for the processing of particulate and dissolved organic carbon, but also for woody debris and for the breakdown of lignin, the most recalcitrant polymer in plant tissue.

Wood decomposition in aquatic and terrestrial ecosystems in the tropics: contrasting biotic and abiotic processes.

Wood decomposition, a critical process in carbon and nutrient cycles, is influenced by environmental conditions, decomposer communities and substrate composition. While these factors differ between land and stream habitats, across-habitat comparisons of wood decay processes are rare, limiting our ability to evaluate the context- dependency of the drivers of decay. Here we tracked wood decomposition of three tree species placed in stream and terrestrial habitats in a lowland tropical forest in Panama. At 3 and 11 months we measured mass loss, wood nitrogen and wood polymer concentrations, and sampled wood-associated fungal and bacterial communities. After 11 months of decay we found that mass loss occurred 9% faster in streams than on land, but loss of cellulose, hemicellulose and lignin did not differ between habitats. We also observed large differences in microbial decomposer communities between habitats. Overall, we found faster mass loss of wood in water, but no differences in biotic decay processes between habitats despite distinct microbial communities in streams and on land. Our research challenges the assumption that wood decays relatively slowly in water reflecting unfavorable environmental conditions and a limited capacity of aquatic microbial communities to effectively degrade wood polymers.

Pretreatment of Hardwood and Miscanthus with Trametes versicolor for Bioenergy Conversion and Densification Strategies.

The pretreatment of plant biomass negatively impacts the economics of many bioenergy and bioproduct processes due to the thermochemical requirements for deconstruction of lignocelluluose. An effective strategy to reduce these severity requirements is to pretreat the biomass with white-rot fungi, such as Trametes versicolor, which have the innate ability to deconstruct lignocellulose with a suite of specialized enzymes. In the present study, the effects of 12 weeks of pretreatment with a wild-type strain (52J) and a cellobiose dehydrogenase-deficient strain (m4D) of T. versicolor on hardwood and Miscanthus were explored. Both strains of T. versicolor led to significant decreases of insoluble lignin and significant increases of soluble lignin after acid hydrolysis, which suggests improved lignin extractability. The glucose yields after saccharification using an enzyme cocktail containing chitinase were similar or significantly higher with 52J-treated biomass compared to untreated hardwood and Miscanthus, respectively. The fungal treated biomass, regardless of the strain used, also showed significant increases in energy content and compressive strength of pellets. Overall, the use of T. versicolor as a pretreatment agent for hardwood and Miscanthus could be an environmentally friendly strategy for conversion technologies that require delignification and saccharification, and/or processes that require densification and transport.

Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response.

The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to the hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that transgenic expression of microbial cell wall modifying enzymes induces transcriptome responses similar to those observed in defense responses.

Soil microbial communities alter leaf chemistry and influence allelopathic potential among coexisting plant species.

While both plant-soil feedbacks and allelochemical interactions are key drivers of plant community dynamics, the potential for these two drivers to interact with each other remains largely unexplored. If soil microbes influence allelochemical production, this would represent a novel dimension of heterogeneity in plant-soil feedbacks. To explore the linkage between soil microbial communities and plant chemistry, we experimentally generated soil microbial communities and evaluated their impact on leaf chemical composition and allelopathic potential. Four native perennial old-field species (two each of Aster and Solidago) were grown in pairwise combination with each species' soil microbial community as well as a sterilized inoculum. We demonstrated unequivocally that variation in soil microbial communities altered leaf chemical fingerprints for all focal plant species and also changed their allelopathic potential. Soil microbes reduced allelopathic potential in bioassays by increasing germination 25-54% relative to sterile control soils in all four species. Plants grown with their own microbial communities had the lowest allelopathic potential, suggesting that allelochemical production may be lessened when growing with microbes from conspecifics. The allelopathic potential of plants grown in congener and confamilial soils was indistinguishable from each other, indicating an equivalent response to all non-conspecific microbial communities within these closely related genera. Our results clearly demonstrated that soil microbial communities cause changes in leaf tissue chemistry that altered their allelopathic properties. These findings represent a new mechanism of plant-soil feedbacks that may structure perennial plant communities over very small spatial scales that must be explored in much more detail.

Microbial community structure of a freshwater system receiving wastewater effluent.

Despite our dependency on treatment facilities to condition wastewater for eventual release to the environment, our knowledge regarding the effects of treated water on the local watershed is extremely limited. Responses of lotic systems to the treated wastewater effluent have been traditionally investigated by examining the benthic macroinvertebrate assemblages and community structure; however, these studies do not address the microbial diversity of the water systems. In the present study, planktonic and benthic bacterial community structure were examined at 14 sites (from 60 m upstream to 12,100 m downstream) and at two time points along an aquatic system receiving treated effluent from the Charleston Wastewater Treatment Plant (Charleston, IL). Total bacterial DNA was isolated and 16S rRNA sequences were analyzed using a metagenomics platform. The community structure in planktonic bacterial communities was significantly correlated with dissolved oxygen concentration. Benthic bacterial communities were not correlated with water quality but did have a significant geographic structuring. A local restructuring effect was observed in both planktonic and benthic communities near the treated wastewater effluent, which was characterized by an increase in abundance of sphingobacteria. Sites further downstream from the wastewater facility appeared to be less influenced by the effluent. Overall, the present study demonstrated the utility of targeted high-throughput sequencing as a tool to assess the effects of treated wastewater effluent on a receiving water system, and highlighted the potential for this technology to be used for routine monitoring by wastewater facilities.

Differential metabolite profiles and salinity tolerance between two genetically related brown-seeded and yellow-seeded Brassica carinata lines.

Brassica carinata (Ethiopian mustard) has previously been identified as a potential crop species suitable for marginal land in the North American prairies due to its relatively high salt tolerance. Two genetically related B. carinata lines with brown-seeded (BS) and yellow-seeded (YS) phenotypes were assessed for their tolerance to sodium sulfate. Specifically, each line was greenhouse-grown under 0, 50 and 100mM of salt, and analyzed after four weeks and eight weeks of treatment. Generally, the height of the BS line was greater than the YS line under both salt treatments, indicating enhanced salt tolerance of the BS line. NMR-based metabolite profiling and PCA analyses indicated a more pronounced shift in key stem metabolites after four weeks of treatment with the YS line compared to the BS line. For example, tryptophan and formate levels increased in the YS line after four weeks of 100mM salt treatment, while proline and threonine levels varied uniquely compared to other metabolites of the lines. Together, the data indicate that the brown-seeded line has greater sodium tolerance than the yellow-seed line, provide clues to the biochemical underpinnings for the phenotypic variation, and highlight the utility of B. carinata as a biorefinery crop for saline land.

Constitutive expression of a fungal glucuronoyl esterase in Arabidopsis reveals altered cell wall composition and structure.

A family 15 carbohydrate esterase (CE15) from the white-rot basidiomycete, Phanerochaete carnosa (PcGCE), was transformed into Arabidopsis thaliana Col-0 and was expressed from the constitutive cauliflower mosaic virus 35S promoter. Like other CE15 enzymes, PcGCE hydrolyzed methyl-4-O-methyl-d-glucopyranuronate and could target ester linkages that contribute to lignin-carbohydrate complexes that form in plant cell walls. Three independently transformed Arabidopsis lines were evaluated in terms of nine morphometric parameters, total sugar and lignin composition, cell wall anatomy, enzymatic saccharification and xylan extractability. The transgenic lines consistently displayed a leaf-yellowing phenotype, as well as reduced glucose and xylose content by as much as 30% and 35%, respectively. Histological analysis revealed 50% reduction in cell wall thickness in the interfascicular fibres of transgenic plants, and FT-IR microspectroscopy of interfascicular fibre walls indicated reduction in lignin cross-linking in plants overexpressing PcGCE. Notably, these characteristics could be correlated with improved xylose recovery in transgenic plants, up to 15%. The current analysis represents the first example whereby a fungal glucuronoyl esterase is expressed in Arabidopsis and shows that the promotion of glucuronoyl esterase activity in plants can alter the extent of intermolecular cross-linking within plant cell walls.

Isolation and characterization of galactinol synthases from hybrid poplar.

The raffinose family of oligosaccharides (RFOs) serve as transport carbohydrates in the phloem, storage compounds in sink tissues, and putative biological agents to combat both abiotic and biotic stress in several plant species. To investigate further the functional roles of this class of compounds in trees, two cDNAs encoding galactinol synthase (GolS, EC 2.4.1.123), which catalyses the first step in the biosynthesis of RFOs, were identified and cloned from hybrid poplar (Populus alba×grandidentata). Phylogenetic analyses of the Populus GolS isoforms with other known GolS proteins suggested a putative role for these enzymes during biotic or abiotic stress in hybrid poplar. The predicted protein sequences of both isoforms (Pa×gGolSI and Pa×gGolSII) showed characteristics of GolS proteins from other species, including a serine phosphorylation site and the ASAAP pentapeptide hydrophobic domain. Kinetic analyses of recombinant Pa×gGolSI and Pa×gGolSII resulted in K(m) values for UPD-galactose of 0.80 and 0.65 mM and V(max) values of 657.5 and 1245 nM min(-1), respectively. Pa×gGolSI inherently possessed a broader pH and temperature range when compared with Pa×gGolSII. Interestingly, spatial and temporal expression analyses revealed that Pa×gGolSII transcript levels varied seasonally, while Pa×gGolSI did not, implying temperature-regulated transcriptional control of this gene in addition to the observed thermosensitivity of the respective enzyme. This evidence suggested that Pa×gGolSI may be involved in basic metabolic activities such as storage, while Pa×gGolSII is probably involved in seasonal mobilization of carbohydrates.

Biological pretreatment with a cellobiose dehydrogenase-deficient strain of Trametes versicolor enhances the biofuel potential of canola straw.

The use of Trametes versicolor as a biological pretreatment for canola straw was explored in the context of biofuel production. Specifically, the effects on the straw of a wild-type strain (52J) and a cellobiose dehydrogenase (CDH)-deficient strain (m4D) were investigated. The xylose and glucose contents of the straw treated with 52J were significantly reduced, while only the xylose content was reduced with m4D treatment. Lignin extractability was greatly improved with fungal treatments compared to untreated straw. Saccharification of the residue of the m4D-treated straw led to a significant increase in proportional glucose yield, which was partially attributed to the lack of cellulose catabolism by m4D. Overall, the results of this study indicate that CDH facilitates cellulose access by T. versicolor. Furthermore, treatment of lignocellulosic material with m4D offers improvements in lignin extractability and saccharification efficacy compared to untreated biomass without loss of substrate due to fungal catabolism.

Transcriptomic responses of the softwood-degrading white-rot fungus Phanerochaete carnosa during growth on coniferous and deciduous wood.

To identify enzymes that could be developed to reduce the recalcitrance of softwood resources, the transcriptomes of the softwood-degrading white-rot fungus Phanerochaete carnosa were evaluated after growth on lodgepole pine, white spruce, balsam fir, and sugar maple and compared to the transcriptome of P. carnosa after growth on liquid nutrient medium. One hundred fifty-two million paired-end reads were obtained, and 63% of these reads were mapped to 10,257 gene models from P. carnosa. Five-hundred thirty-three of these genes had transcripts that were at least four times more abundant during growth on at least one wood medium than on nutrient medium. The 30 transcripts that were on average over 100 times more abundant during growth on wood than on nutrient medium included 6 manganese peroxidases, 5 cellulases, 2 hemicellulases, a lignin peroxidase, glyoxal oxidase, and a P450 monooxygenase. Notably, among the genes encoding putative cellulases, one encoding a glycosyl hydrolase family 61 protein had the highest relative transcript abundance during growth on wood. Overall, transcripts predicted to encode lignin-degrading activities were more abundant than those predicted to encode carbohydrate-active enzymes. Transcripts predicted to encode three MnPs represented the most highly abundant transcripts in wood-grown cultivations compared to nutrient medium cultivations. Gene set enrichment analyses did not distinguish transcriptomes resulting from softwood and hardwood cultivations, suggesting that similar sets of enzyme activities are elicited by P. carnosa grown on different wood substrates, albeit to different expression levels.

Sucrose phosphate synthase expression influences poplar phenology.

The objective of this study was to manipulate the intracellular pools of sucrose, and investigate its role in regulating plant growth, phenology (leaf senescence and bud break) and fibre development. This objective was achieved by differentially expressing an Arabidopsis (Arabidopsis thaliana L. Heynh.) sucrose phosphate synthase (SPS) gene in hybrid poplar (Populus alba L.xPopulus grandidentata Michx.), a model system for tree biology with substantial industrial relevance in the context of short rotation forestry and a target bioenergy crop. Phenotypic differences were evident in the transgenic trees, as both the timing of bud flush and leaf senescence were altered compared to wild-type (WT) trees. Tree height and stem diameter were similar in WT and in the AtSPS transgenic trees, however, there were differences in the length of xylem fibres. Elevated concentrations of intracellular sucrose in both leaf and stem tissue of the transgenic trees are associated with a prolonged onset of senescence and an advancement in bud flush in the following spring. The association among sucrose content, tree phenology and elevated SPS gene expression implicates both enzyme and product in regulating poplar developmental processes.

Heterologous expression and functional characterization of two hybrid poplar cell-wall invertases.

The expression of two hybrid poplar cell-wall invertases (EC 3.2.1.26; PaxgINV1 and PaxgINV2) were previously shown to be spatially and temporally regulated in the vegetative tissues. The expression of PaxgINV1 was linked to processes relating to dormancy, while PaxgINV2 expression was prominent in tissues undergoing growth and expansion. In an effort to further elucidate the physiological roles of these key cell wall enzymes, PaxgINV1 and PaxgINV2 were heterologously expressed in the methylotrophic yeast Pichia pastoris. Three-dimensional predictive models of the poplar invertases revealed a structural channel containing both the conserved beta-fructofuranosidase and cell-wall invertase motifs, suggesting that this channel is the putative active site of these enzymes. Recombinant PaxgINV1 and PaxgINV2 had pH optima of 4.8 and 5.6 and temperature optima of 45 and 40 degrees C, respectively. Functional characterization revealed the ability for both enzymes to hydrolyze the fructose residue of sucrose, raffinose, stachyose and verbascose, with PaxgINV2 having higher specific activity for each of the substrates tested. The K(m) values of sucrose/raffinose/stachyose were 1.7/1.8/5.0 mM for PaxgINV1 and 1.6/1.7/1.9 mM for PaxgINV2, respectively. Activity analyses in the presence of various metal cations showed that PaxgINV2 was strongly inhibited by Cu(2+), Zn(2+) and Hg(2+), while PaxgINV1 was only weakly inhibited by these cations. The results from this study, coupled with previous expression data, suggest that PaxgINV1 and PaxgINV2 have distinct roles with respect to the physiology and development of hybrid poplar, specifically phloem unloading and processes related to dormancy and bud break.

Spatial and temporal expression profiling of cell-wall invertase genes during early development in hybrid poplar.

Cell-wall invertase genes are spatially and temporally regulated in several plant species, including Daucus carota L., Lycopersicon esculentum L. and Solanum tuberosum L. However, few studies of cell-wall invertase genes of trees have been conducted, despite the importance of trees as a source of lignocellulosic biopolymers. We identified three putative cell-wall invertase genes in hybrid poplar (Populus alba L. x grandidentata Michx.) that showed higher homology to each other than to cell-wall invertases of other dicotyledonous species, with two of the genes (PaxgINV2 and PaxgINV3) appearing as a genomic tandem repeat. These genes are more similar to each other than to tandemly repeated cell-wall invertases of other plants, perhaps indicating parallel evolution of a duplication event with cell-wall invertases in dicotyledons. Spatial and temporal expression analyses throughout a complete annual cycle indicated that PaxgINV1 and PaxgINV2 are highly regulated in vegetative tissues during three distinct growth phases: early growth, dormancy and post-dormancy. Expression of the third gene (PaxgINV3) appears to be tightly regulated and may represent a floral-specific cell-wall invertase. Of the two genes expressed in vegetative tissues, PaxgINV1 appears to be exclusively involved in processes related to dormancy, whereas PaxgINV2 appears to encode an enzyme involved in phloem unloading and in providing actively growing tissues, such as developing xylem, with the energy and carbon skeletons necessary for respiration and cell wall biosynthesis.

Over-expression of an arabidopsis family A sucrose phosphate synthase (SPS) gene alters plant growth and fibre development.

The objective of this study was to manipulate the intracellular pools of sucrose by differentially expressing exogenous sucrose phosphate synthase (SPS) and investigating its role in regulating plant growth and fibre development. Tobacco (Nicotiana tabacum cv. Xanthi) plants were transformed with an arabidopsis SPS gene under the regulation of the ubiquitously expressed tandem repeat of the 35S cauliflower mosaic virus promoter, and subject to growth trials and fibre characterization. It was apparent that over-expression of SPS resulted in substantially elevated concentrations of sink sucrose pools compared to wild-type plants, while source tissue sucrose pools remained the same. All transformed plants had significantly increased stem height, which was ascribed to internode elongation, and greater stem diameters, longer fibers and increased total dry biomass relative to the control plants. Difference in the chemical composition of either the storage or structural carbohydrates of the wild-type and SPS transgenic lines were only minor. The correlation between increased stem sucrose content and plant phenotypes with elevated SPS gene expression confirm a role for sucrose availability in controlling plant growth and fibre elongation.

Over-expression of UDP-glucose pyrophosphorylase in hybrid poplar affects carbon allocation.

The effects of the over-expression of the Acetobacter xylinum UDP-glucose pyrophosphorylase (UGPase) under the control of the tandem repeat Cauliflower Mosaic Virus promoter (2x35S) on plant metabolism and growth were investigated in hybrid poplar (Populus albaxgrandidentata). Transcript levels, enzyme activity, growth parameters, leaf morphology, structural and soluble carbohydrates, and soluble metabolite levels were quantified in both transgenic and wild-type trees. Transgenic 2x35S::UGPase poplar showed impaired growth rates, displaying reduced height growth and stem diameter. Morphologically, 2x35S::UGPase trees had elongated axial shoots, and leaves that were substantially smaller in size when compared with wild-type trees at equivalent developmental stages. Biochemical analysis revealed significant increases in soluble sugar, starch, and cellulose contents, and concurrent decreases in lignin content. Lignin monomer composition was altered in favour of syringyl moieties. Detailed soluble metabolite analysis revealed that 2x35S::UGPase trees had as much as a 270-fold increase in the salicylic acid 2-O-beta-D-glucoside (SAG), a compound typically associated with the stress response. These data suggest that while it is possible to alter the allocation of carbon in favour of cellulose biosynthesis, whole plant changes result in unexpected decreases in growth and an increase in defence metabolites.

Varied growth, biomass and cellulose content in tobacco expressing yeast-derived invertases.

The effects of the expression of yeast-derived apoplastic (AI) and cytosolic (CI) invertases (EC 3.2.1.26) on biomass and structural carbohydrate accumulation in tobacco (Nicotiana tabacum L. cv. Xanthi) were evaluated. Transgenic tobacco plants expressing AI or CI under the control of either a tandem repeat of the Cauliflower Mosaic Virus 35S promoter (2X35S), or a promoter that drives xylem-localized expression (Petroselinum crispum 4-coumarate:CoA ligase promoter; 4CL) were generated. Yeast-derived invertase transcript levels, invertase protein, enzyme activity, growth parameters as well as both structural and soluble carbohydrates of stem tissue of all transformed lines were quantified. Transgenic tobacco lines expressing invertase under the control of 4CL displayed severe growth retardation with both yeast-derived isogenes. Similarly, several transformed lines expressing either AI or CI regulated by the 2X35S promoter were also shorter than wild-type (WT) plants. Despite the decreases in height, some transformed lines had significant increases in biomass. One line (2X35S::AI-1) had a biomass/height increase of 88% and an increase in stem diameter of over 40%, while a second line (2X35S::CI-5) had a biomass/height increase of 21%. A separate line (2X35S::AI-2) had a 36% increase in cellulose content, while two others (4CL::AI-2 and 4CL::AI-3) displayed significant decreases in cellulose content. The observed phenotypes can be in part explained by the levels of foreign invertase present, subcellular localization and the carbohydrate status of the tissues.