SUMMARYWe discuss and illustrate recent advances that have been made to image the distribution of metabolites among cells and tissues of plants using different mass spectrometry technologies. These technologies include matrixassisted laser desorption ionization, desorption electrospray ionization, and secondary ion mass spectrometry. These are relatively new technological applications of mass spectrometry and they are providing highly spatially resolved data concerning the cellular distribution of metabolites. We discuss the advantages and limitations of each of these mass spectrometric methods, and provide a description of the technical barriers that are currently limiting the technology to the level of single-cell resolution. However, we anticipate that advances in the next few years will increase the resolving power of the technology to provide unprecedented data on the distribution of metabolites at the subcellular level, which will increase our ability to decipher new knowledge concerning the spatial organization of metabolic processes in plants.

High-spatial resolution and high-mass resolution techniques are developed and adopted for the mass spectrometric imaging of epicuticular lipids on the surface of Arabidopsis thaliana. Single cell level spatial resolution of approximately 12 mum was achieved by reducing the laser beam size by using an optical fiber with 25 mum core diameter in a vacuum matrix-assisted laser desorption ionization-linear ion trap (vMALDI-LTQ) mass spectrometer and improved matrix application using an oscillating capillary nebulizer. Fine chemical images of a whole flower were visualized in this high spatial resolution showing substructure of an anther and single pollen grains at the stigma and anthers. The LTQ-Orbitrap with a MALDI ion source was adopted to achieve MS imaging in high mass resolution. Specifically, isobaric silver ion adducts of C29 alkane (m/z 515.3741) and C28 aldehyde (m/z 515.3377), indistinguishable in low-resolution LTQ, can now be clearly distinguished and their chemical images could be separately constructed. In the application to roots, the high spatial resolution allowed molecular MS imaging of secondary roots and the high mass resolution allowed direct identification of lipid metabolites on root surfaces.

The ontogeny of seed structure and the accumulation of seed storage substances is the result of a determinant genetic program. Using RNA interference, the synthesis of soybean (Glycine max) glycinin and conglycinin storage proteins has been suppressed. The storage protein knockdown (SP2) seeds are overtly identical to the wild type, maturing to similar size and weight, and in developmental ontogeny. The SP2 seeds rebalance the proteome, maintaining wild-type levels of protein and storage triglycerides. The SP2 soybeans were evaluated with systems biology techniques of proteomics, metabolomics, and transcriptomics using both microarray and next-generation sequencing transcript sequencing (RNA-Seq). Proteomic analysis shows that rebalancing of protein content largely results from the selective increase in the accumulation of only a few proteins. The rebalancing of protein composition occurs with small alterations to the seed's transcriptome and metabolome. The selectivity of the rebalancing was further tested by introgressing into the SP2 line a green fluorescent protein (GFP) glycinin allele mimic and quantifying the resulting accumulation of GFP. The GFP accumulation was similar to the parental GFPexpressing line, showing that the GFP glycinin gene mimic does not participate in proteome rebalancing. The results show that soybeans make large adjustments to the proteome during seed filling and compensate for the shortage of major proteins with the increased selective accumulation of other proteins that maintains a normal protein content.

Purpose – The purpose of this paper is to examine the relationships among organizational learning, absorptive capacity, imitation and innovation in the Chinese context. Design/methodology/approach – Based on the organizational learning theory and innovation theory, the paper presents a framework linking organizational learning, absorptive capacity, imitation and innovation. Using a key informant technique, a survey questionnaire was designed and sent to the middle or top management managers of 115 firms located in Peking, People’s Republic (PR) of China. Structural equation modeling (SEM) with the maximum likelihood (ML) estimation procedures was applied to test the hypotheses developed in the research. Findings – The empirical results show that both organizational learning and absorptive capacity have positive impacts on innovation; imitation has a positive impact on absorptive capacity; absorptive capacity mediates the relationship between imitation and innovation. Practical implications – This study has implications for firms aiming to enhance innovation by organizational learning, absorptive capacity and imitation. Originality/value – Despite the number of studies concerning organizational learning, absorptive capacity, imitation and innovation, research that encompasses the interrelationships between the four concepts simultaneously remains scarce. The paper provides a framework linking organizational learning, imitation, absorptive capacity and innovation, and it advances the argument that absorptive capacity is an important factor in predicting the Chinese firms’ transition from imitation to innovation.

Colloidal silver laser desorption/ionization (LDI) mass spectrometry (MS) was employed to directly profile and image epicuticular wax metabolites on a variety of different surfaces of Arabidopsis thaliana leaves and flowers. Major cuticular wax compounds, such as very long-chain fatty acids, alcohols, alkanes, and ketones, were successfully detected as silver adduct ions. The surface metabolites of different flower organs (carpels, petals, and sepals) were profiled for the first time at a spatial resolution of approximately 100 microm. In addition, mass spectral profiles and images were collected from wild type and a mutant strain, which carried alleles that affect the surface constituents of this organism. One of these mutant alleles (cer2-2) is in a gene whose biochemical functionality is still unclear, although its effect on normal epicuticular wax deposition was the characteristic that led to its original identification. Variations of wax products between different spatial locations for wild type and for a mutant strain were investigated by normalizing the ion intensities to a reference peak ([(107)Ag + (109)Ag](+)). The spatially resolved surface metabolite profiling data of this mutant has provided new insights into the complexity of epicuticular wax deposition at the cellular-resolution scale. This MS-based metabolite imaging technology has the potential to provide valuable data for dissecting metabolism in multicellular organism at the level of single cells.

HighlightWe uncovered significant structural and compositional differences between leaf cuticles of drought-tolerant and drought-sensitive wheat cultivars. Specific MYB factors drive cuticular responses to drought and may underlie genotypes with contrasting drought tolerance.

While it is widely accepted that genetic diversity determines the potential of adaptation, the role that gene expression variation plays in adaptation remains poorly known. Here we show that gene expression diversity could have played a positive role in the adaptation of Miscanthus lutarioriparius. RNA-seq was conducted for 80 individuals of the species, with half planted in the energy crop domestication site and the other half planted in the control site near native habitats. A leaf reference transcriptome consisting of 18,503 high-quality transcripts was obtained using a pipeline developed for de novo assembling with population RNA-seq data. The population structure and genetic diversity of M. lutarioriparius were estimated based on 30,609 genic single nucleotide polymorphisms. Population expression (E p ) and expression diversity (E d ) were defined to measure the average level and the magnitude of variation of a gene expression in the population, respectively. It was found that expression diversity increased while genetic diversity decreased after the species was transplanted from the native habitats to the harsh domestication site, especially for genes involved in abiotic stress resistance, histone methylation, and biomass synthesis under water limitation. The increased expression diversity could have enriched phenotypic variation directly subject to selections in the new environment.

Sterol methyltransferase (SMT) from Saccharomyces cerevisiae was purified from Escherichia coli BL21(DE3) and labeled with the mechanism-based irreversible inhibitor [3-3H]26,27-dehydrozymosterol (26,27-DHZ). A 5-kDa tryptic digest peptide fragment containing six acidic residues at positions Glu-64, Asp-65, Glu-68, Asp-79, Glu-82, and Glu-98 was determined to contain the substrate analog covalently attached to Glu-68 by Edman sequencing and radioanalysis using C18 reverse phase high performance liquid chromatography. Site-directed mutagenesis of the six acidic residues to leucine followed by activity assay of the purified mutants confirmed Glu-68 as the only residue to participate in affinity labeling. Equilibration studies indicated that zymosterol and 26,27-DHZ were bound to native and the E68L mutant with similar affinity, whereas S-adenosylmethionine was bound only to the native SMT, K(d) of about 2 microm. Analysis of the incubation products of the wild-type and six leucine mutants by GC-MS demonstrated that zymosterol was converted to fecosterol, 26,27-DHZ was converted to 26-homo-cholesta-8(9),23(24)E,26(26')-trienol as well as 26-homocholesta-8(9),26(26')-3beta,24beta-dienol, and in the case of D79L and E82L mutants, zymosterol was also converted to a new product, 24-methylzymosta-8,25(27)-dienol. The structures of the methylenecyclopropane ring-opened olefins were determined unambiguously by a combination of (1)H and (13)C NMR techniques. A K(m) of 15 microm, K(cat) of 8 x 10(-4) s(-1), and partition ratio of 0.03 was established for 26,27-DHZ, suggesting that the methylenecyclopropane can serve as a lead structure for a new class of antifungal agents. Taken together, partitioning that leads to loss of enzyme function is the result of 26,27-DHZ catalysis forming a highly reactive cationic intermediate that interacts with the enzyme in a region normally not occupied by the zymosterol high energy intermediate as a consequence of less than perfect control. Alternatively, the gain in enzyme function resulting from the production of a delta(25(27))-olefin originates with the leucine substitution directing substrate channeling along different reaction channels in a similar region at the active site.