Evidence for extensive heterotrophic metabolism, antioxidant action, and associated regulatory events during winter hardening in Sitka spruce.

BACKGROUND: Cold acclimation in woody perennials is a metabolically intensive process, but coincides with environmental conditions that are not conducive to the generation of energy through photosynthesis. While the negative effects of low temperatures on the photosynthetic apparatus during winter have been well studied, less is known about how this is reflected at the level of gene and metabolite expression, nor how the plant generates primary metabolites needed for adaptive processes during autumn. RESULTS: The MapMan tool revealed enrichment of the expression of genes related to mitochondrial function, antioxidant and associated regulatory activity, while changes in metabolite levels over the time course were consistent with the gene expression patterns observed. Genes related to thylakoid function were down-regulated as expected, with the exception of plastid targeted specific antioxidant gene products such as thylakoid-bound ascorbate peroxidase, components of the reactive oxygen species scavenging cycle, and the plastid terminal oxidase. In contrast, the conventional and alternative mitochondrial electron transport chains, the tricarboxylic acid cycle, and redox-associated proteins providing reactive oxygen species scavenging generated by electron transport chains functioning at low temperatures were all active. CONCLUSIONS: A regulatory mechanism linking thylakoid-bound ascorbate peroxidase action with "chloroplast dormancy" is proposed. Most importantly, the energy and substrates required for the substantial metabolic remodeling that is a hallmark of freezing acclimation could be provided by heterotrophic metabolism.

Changes in RNA Splicing in Developing Soybean (Glycine max) Embryos.

Developing soybean seeds accumulate oils, proteins, and carbohydrates that are used as oxidizable substrates providing metabolic precursors and energy during seed germination. The accumulation of these storage compounds in developing seeds is highly regulated at multiple levels, including at transcriptional and post-transcriptional regulation. RNA sequencing was used to provide comprehensive information about transcriptional and post-transcriptional events that take place in developing soybean embryos. Bioinformatics analyses lead to the identification of different classes of alternatively spliced isoforms and corresponding changes in their levels on a global scale during soybean embryo development. Alternative splicing was associated with transcripts involved in various metabolic and developmental processes, including central carbon and nitrogen metabolism, induction of maturation and dormancy, and splicing itself. Detailed examination of selected RNA isoforms revealed alterations in individual domains that could result in changes in subcellular localization of the resulting proteins, protein-protein and enzyme-substrate interactions, and regulation of protein activities. Different isoforms may play an important role in regulating developmental and metabolic processes occurring at different stages in developing oilseed embryos.

Metabolic and Transcriptional Reprogramming in Developing Soybean (Glycine max) Embryos.

Soybean (Glycine max) seeds are an important source of seed storage compounds, including protein, oil, and sugar used for food, feed, chemical, and biofuel production. We assessed detailed temporal transcriptional and metabolic changes in developing soybean embryos to gain a systems biology view of developmental and metabolic changes and to identify potential targets for metabolic engineering. Two major developmental and metabolic transitions were captured enabling identification of potential metabolic engineering targets specific to seed filling and to desiccation. The first transition involved a switch between different types of metabolism in dividing and elongating cells. The second transition involved the onset of maturation and desiccation tolerance during seed filling and a switch from photoheterotrophic to heterotrophic metabolism. Clustering analyses of metabolite and transcript data revealed clusters of functionally related metabolites and transcripts active in these different developmental and metabolic programs. The gene clusters provide a resource to generate predictions about the associations and interactions of unknown regulators with their targets based on "guilt-by-association" relationships. The inferred regulators also represent potential targets for future metabolic engineering of relevant pathways and steps in central carbon and nitrogen metabolism in soybean embryos and drought and desiccation tolerance in plants.

Mining and visualization of microarray and metabolomic data reveal extensive cell wall remodeling during winter hardening in Sitka spruce (Picea sitchensis).

Microarray gene expression profiling is a powerful technique to understand complex developmental processes, but making biologically meaningful inferences from such studies has always been challenging. We previously reported a microarray study of the freezing acclimation period in Sitka spruce (Picea sitchensis) in which a large number of candidate genes for climatic adaptation were identified. In the current paper, we apply additional systems biology tools to these data to further probe changes in the levels of genes and metabolites and activities of associated pathways that regulate this complex developmental transition. One aspect of this adaptive process that is not well understood is the role of the cell wall. Our data suggest coordinated metabolic and signaling responses leading to cell wall remodeling. Co-expression of genes encoding proteins associated with biosynthesis of structural and non-structural cell wall carbohydrates was observed, which may be regulated by ethylene signaling components. At the same time, numerous genes, whose products are putatively localized to the endomembrane system and involved in both the synthesis and trafficking of cell wall carbohydrates, were up-regulated. Taken together, these results suggest a link between ethylene signaling and biosynthesis, and targeting of cell wall related gene products during the period of winter hardening. Automated Layout Pipeline for Inferred NEtworks (ALPINE), an in-house plugin for the Cytoscape visualization environment that utilizes the existing GeneMANIA and Mosaic plugins, together with the use of visualization tools, provided images of proposed signaling processes that became active over the time course of winter hardening, particularly at later time points in the process. The resulting visualizations have the potential to reveal novel, hypothesis-generating, gene association patterns in the context of targeted subcellular location.

Effects of drought on gene expression in maize reproductive and leaf meristem tissue revealed by RNA-Seq.

Drought stress affects cereals especially during the reproductive stage. The maize (Zea mays) drought transcriptome was studied using RNA-Seq analysis to compare drought-treated and well-watered fertilized ovary and basal leaf meristem tissue. More drought-responsive genes responded in the ovary compared with the leaf meristem. Gene Ontology enrichment analysis revealed a massive decrease in transcript abundance of cell division and cell cycle genes in the drought-stressed ovary only. Among Gene Ontology categories related to carbohydrate metabolism, changes in starch and Suc metabolism-related genes occurred in the ovary, consistent with a decrease in starch levels, and in Suc transporter function, with no comparable changes occurring in the leaf meristem. Abscisic acid (ABA)-related processes responded positively, but only in the ovaries. Related responses suggested the operation of low glucose sensing in drought-stressed ovaries. The data are discussed in the context of the susceptibility of maize kernel to drought stress leading to embryo abortion and the relative robustness of dividing vegetative tissue taken at the same time from the same plant subjected to the same conditions. Our working hypothesis involves signaling events associated with increased ABA levels, decreased glucose levels, disruption of ABA/sugar signaling, activation of programmed cell death/senescence through repression of a phospholipase C-mediated signaling pathway, and arrest of the cell cycle in the stressed ovary at 1 d after pollination. Increased invertase levels in the stressed leaf meristem, on the other hand, resulted in that tissue maintaining hexose levels at an "unstressed" level, and at lower ABA levels, which was correlated with successful resistance to drought stress.