Dimah Z. Habash

Two Cytosolic Glutamine Synthetase Isoforms of Maize Are Specifically Involved in the Control of Grain Production

The roles of two cytosolic maize glutamine synthetase isoenzymes (GS1), products of the Gln1-3 and Gln1-4 genes, were investigated by examining the impact of knockout mutations on kernel yield. In the gln1-3 and gln1-4 single mutants and the gln1-3 gln1-4 double mutant, GS mRNA expression was impaired, resulting in reduced GS1 protein and activity. The gln1-4 phenotype displayed reduced kernel size and gln1-3 reduced kernel number, with both phenotypes displayed in gln1-3 gln1-4. However, at maturity, shoot biomass production was not modified in either the single mutants or double mutants, suggesting a specific impact on grain production in both mutants. Asn increased in the leaves of the mutants during grain filling, indicating that it probably accumulates to circumvent ammonium buildup resulting from lower GS1 activity. Phloem sap analysis revealed that unlike Gln, Asn is not efficiently transported to developing kernels, apparently causing reduced kernel production. When Gln1-3 was overexpressed constitutively in leaves, kernel number increased by 30%, providing further evidence that GS1-3 plays a major role in kernel yield. Cytoimmunochemistry and in situ hybridization revealed that GS1-3 is present in mesophyll cells, whereas GS1-4 is specifically localized in the bundle sheath cells. The two GS1 isoenzymes play nonredundant roles with respect to their tissue-specific localization.

The importance of cytosolic glutamine synthetase in nitrogen assimilation and recycling

Glutamine synthetase assimilates ammonium into amino acids, thus it is a key enzyme for nitrogen metabolism. The cytosolic isoenzymes of glutamine synthetase assimilate ammonium derived from primary nitrogen uptake and from various internal nitrogen recycling pathways. In this way, cytosolic glutamine synthetase is crucial for the remobilization of protein-derived nitrogen. Cytosolic glutamine synthetase is encoded by a small family of genes that are well conserved across plant species. Members of the cytosolic glutamine synthetase gene family are regulated in response to plant nitrogen status, as well as to environmental cues, such as nitrogen availability and biotic/abiotic stresses. The complex regulation of cytosolic glutamine synthetase at the transcriptional to post-translational levels is key to the establishment of a specific physiological role for each isoenzyme. The diverse physiological roles of cytosolic glutamine synthetase isoenzymes are important in relation to current agricultural and ecological issues.

Genomic approaches for designing durum wheat ready for climate change with a focus on drought

Climate change is projected to have a significant impact on temperature and precipitation profiles in the Mediterranean basin. The incidence and severity of drought will become commonplace and this will reduce the productivity of rain-fed crops such as durum wheat. Genetic diversity is the material basis for crop improvement and plant breeding has exploited naturally occurring variation to deliver cultivars with improved resistance to abiotic stresses. The coupling of new genomic tools, technologies, and resources with genetic approaches is essential to underpin wheat breeding through marker-assisted selection and hence mitigate climate change. Improvements in crop performance under abiotic stresses have primarily targeted yield-related traits and it is anticipated that the application of genomic technologies will introduce new target traits for consideration in wheat breeding for resistance to drought. Many traits relating to the plants response and adaptation to drought are complex and multigenic, and quantitative genetics coupled with genomic technologies have the potential to dissect complex genetic traits and to identify regulatory loci, genes and networks. Full realization of our abilities to manipulate metabolism, transduction pathways, and transcription factors for crop improvement ultimately relies on our basic understanding of the regulation of plant networks at all levels of function.

New perspectives on glutamine synthetase in grasses

Members of the glutamine synthetase (GS) gene family have now been characterized in many crop species such as wheat, rice, and maize. Studies have shown that cytosolic GS isoforms are involved in nitrogen remobilization during leaf senescence and emphasized a role in seed production particularly in small grain crop species. Data from the sequencing of genomes for model crops and expressed sequence tag (EST) libraries from non-model species have strengthened the idea that the cytosolic GS genes are organized in three functionally and phylogenetically conserved subfamilies. Using a bioinformatic approach, the considerable publicly available information on high throughput gene expression was mined to search for genes having patterns of expression similar to GS. Interesting new hypotheses have emerged from searching for co-expressed genes across multiple unfiltered experimental data sets in rice. This approach should inform new experimental designs and studies to explore the regulation of the GS gene family further. It is expected that understanding the regulation of GS under varied climatic conditions will emerge as an important new area considering the results from recent studies that have shown nitrogen assimilation to be critical to plant acclimation to high CO(2) concentrations.

Wheat nitrogen metabolism during grain filling : comparative role of glumes and the flag leaf

The mobilization of nitrogen (N) compounds and the roles played by glumes and the flag leaf during grain filling were studied in bread wheat (Triticum aestivum L. cv. Florida) grown under field conditions. Glumes lost twice as much of their total N content as that lost by the flag leaf between the milk and early dough stages. In the flag leaf, glumes and grains, Glu, Asp, Ser and Ala accounted for 85% of all the reductions in the free amino acid pool. Principal component analysis of free amino acid pools separated grains from the glumes and the flag leaf, suggesting grain specific regulations in the use of free amino acids in protein synthesis. In all three organs, no decrease in Gln was detected, probably due to steady glutamine synthetase (GS; EC 6.3.1.2) activities per soluble protein in both the flag leaf and glumes. Compared with the flag leaf, glumes presented relatively smaller amounts of the chloroplast GS associated isoform. This we show is due to a lower relative number of mesophyll cells in glumes as supported by the different anatomy and the cellular pattern of the GS immunolocalization. We argue that cellular distribution plays a key role in supporting metabolism to enable the various functions undertaken by glume tissue.

Systems Responses to Progressive Water Stress in Durum Wheat

Durum wheat is susceptible to terminal drought which can greatly decrease grain yield. Breeding to improve crop yield is hampered by inadequate knowledge of how the physiological and metabolic changes caused by drought are related to gene expression. To gain better insight into mechanisms defining resistance to water stress we studied the physiological and transcriptome responses of three durum breeding lines varying for yield stability under drought. Parents of a mapping population (Lahn x Cham1) and a recombinant inbred line (RIL2219) showed lowered flag leaf relative water content, water potential and photosynthesis when subjected to controlled water stress time transient experiments over a six-day period. RIL2219 lost less water and showed constitutively higher stomatal conductance, photosynthesis, transpiration, abscisic acid content and enhanced osmotic adjustment at equivalent leaf water compared to parents, thus defining a physiological strategy for high yield stability under water stress. Parallel analysis of the flag leaf transcriptome under stress uncovered global trends of early changes in regulatory pathways, reconfiguration of primary and secondary metabolism and lowered expression of transcripts in photosynthesis in all three lines. Differences in the number of genes, magnitude and profile of their expression response were also established amongst the lines with a high number belonging to regulatory pathways. In addition, we documented a large number of genes showing constitutive differences in leaf transcript expression between the genotypes at control non-stress conditions. Principal Coordinates Analysis uncovered a high level of structure in the transcriptome response to water stress in each wheat line suggesting genome-wide co-ordination of transcription. Utilising a systems-based approach of analysing the integrated wheat’s response to water stress, in terms of biological robustness theory, the findings suggest that each durum line transcriptome responded to water stress in a genome-specific manner which contributes to an overall different strategy of resistance to water stress.

AIGO: Towards a unified framework for the Analysis and the Inter-comparison of GO functional annotations

BackgroundIn response to the rapid growth of available genome sequences, efforts have been made to develop automatic inference methods to functionally characterize them. Pipelines that infer functional annotation are now routinely used to produce new annotations at a genome scale and for a broad variety of species. These pipelines differ widely in their inference algorithms, confidence thresholds and data sources for reasoning. This heterogeneity makes a comparison of the relative merits of each approach extremely complex. The evaluation of the quality of the resultant annotations is also challenging given there is often no existing gold-standard against which to evaluate precision and recall.ResultsIn this paper, we present a pragmatic approach to the study of functional annotations. An ensemble of 12 metrics, describing various aspects of functional annotations, is defined and implemented in a unified framework, which facilitates their systematic analysis and inter-comparison. The use of this framework is demonstrated on three illustrative examples: analysing the outputs of state-of-the-art inference pipelines, comparing electronic versus manual annotation methods, and monitoring the evolution of publicly available functional annotations. The framework is part of the AIGO library (http://code.google.com/p/aigo) for the Analysis and the Inter-comparison of the products of Gene Ontology (GO) annotation pipelines. The AIGO library also provides functionalities to easily load, analyse, manipulate and compare functional annotations and also to plot and export the results of the analysis in various formats.ConclusionsThis work is a step toward developing a unified framework for the systematic study of GO functional annotations. This framework has been designed so that new metrics on GO functional annotations can be added in a very straightforward way.

Integrating Multiple Studies of Wheat Microarray Data to Identify Treatment-Specific Regulatory Networks

Microarrays have allowed biologists to better understand gene regulatory mechanisms. Wheat microarray data analysis is a complex and challenging topic and knowledge of gene regulation in wheat is still very superficial. However, understanding key mechanisms in this plant holds much potential for food security, especially with a changing climate. The purpose of this paper is to combine multiple microarray studies to automatically identify subnetworks that are distinctive to specific experimental conditions. For example, identifying a regulatory network of genes that only exists under certain types of experimental conditions will assist in understanding the nature of the mechanisms. We derive unique networks from multiple independent networks to better understand key mechanisms and how they change under different conditions. We compare the results with biclustering, detect the most predictive genes and validate the results based upon known biological mechanisms. We also explore how this pipeline performs on yeast microarray data.